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We have investigated the importance of carotenoids on the accumulation and function of the photosynthetic apparatus using a mutant of the green alga Chlamydomonas reinhardtii lacking carotenoids. The FN68 mutant is deficient in phytoene synthase, the first enzyme of the carotenoid biosynthesis pathway, and therefore is unable to synthesize any carotenes and xanthophylls. We find that FN68 is unable to accumulate the light-harvesting complexes associated with both photosystems as well as the RC subunits of photosystem II. The accumulation of the cytochrome b6f complex is also strongly reduced to a level approximately 10% that of the wild type. However, the residual fraction of assembled cytochrome b6f complexes exhibits single-turnover electron transfer kinetics comparable to those observed in the wild-type strain. Surprisingly, photosystem I is assembled to significant levels in the absence of carotenoids in FN68 and possesses functional properties that are very similar to those of the wild-type complex.
Carotenoids (Cars) are fundamental components of the photosynthetic apparatus (Young and Britton, 1993, and refs. therein). The vast majority of Cars are noncovalently bound to either the core or the antenna subunits of PSI or PSII (Siefermann-Harms, 1985; Bassi et al., 1993). The most abundant Car bound to the core subunits of both photosystems is β-carotene, which is found in the vast majority of oxygenic organisms (Siefermann-Harms, 1985; Bassi et al., 1993). The light-harvesting complexes (LHCs) that act as the outer antenna in plants and green algae bind a wider range of oxygenated Cars, known as xanthophylls, the most abundant of which is lutein (Siefermann-Harms, 1985; Bassi et al., 1993; Jennings et al., 1996). The stoichiometry of xanthophylls binding to LHC complexes depends on the particular complexes and often on the illumination conditions during the organism's growth (Siefermann-Harms, 1985; Demmig-Adams, 1990; Horton et al., 1996). Intriguingly, a molecule of β-carotene (as well as a molecule of chlorophyll [Chl] a) is found also in the cytochrome (Cyt) b6f complex (Kurisu et al., 2003; Stroebel et al., 2003).
Cars have multiple functions in the photosynthetic process; they act as light-harvesting pigments (Frank and Cogdell, 1993), enlarging the optical cross section to radiation that is poorly absorbed by Chl. Moreover, Cars play a crucial role in processes such as nonphotochemical quenching that control the efficiency of light harvesting in response to the intensity of the incident radiation (for review, see Demmig-Adams, 1990; Horton et al., 1996; Niyogi, 1999). Probably the most important role of Cars in photosynthesis is the quenching of the excited triplet state of Chl (for review, see Frank and Cogdell, 1993; Giacometti et al., 2007), preventing the formation of highly reactive singlet oxygen, which represents the principal species active under high light stress (Hideg et al., 1994; Krieger-Liszkay, 2005). The importance of Cars is demonstrated by the observation that disruption of their biosynthesis through mutation, or by inhibition of a key enzyme in the pathway, leads to either lethal phenotypes or to rapid photobleaching of the photosynthetic tissue (Claes, 1957; Faludi-Dániel et al., 1968, 1970; Bolychevtseva et al., 1995; Trebst and Depka, 1997).
Moreover, it has been shown that the presence of xanthophylls is absolutely necessary for refolding in vitro of LHC I and LHC II antenna complexes (Plumley and Schmidt, 1987; Paulsen et al., 1993; Sandonà et al., 1998). Such Cars, therefore, have a structural role, as well as their involvement in light harvesting, nonphotochemical quenching regulation, and the quenching of the Chl triplet state. Whether Cars also play a key structural role in the formation and stability of the core complexes of both PSI and PSII has not been systematically explored, since assembly of these complexes in vitro is not feasible. Studies in vivo using higher plants are complicated by the fact that Car deficiency is lethal and can be studied only during the early stages of greening and leaf development (Faludi-Dániel et al., 1968, 1970; Inwood et al., 2008). In these studies, it was shown that the accumulation of PSII complexes was greatly impaired in mutants of maize (Zea mays; Faludi-Dániel et al., 1968, 1970; Inwood et al., 2008), while the assembly of PSI appeared to be less sensitive to Car availability. In mutants of the cyanobacterium Synechocystis sp. PCC 6803 lacking the genes for phytoene desaturase or ζ-carotene desaturase, there was a complete loss of PSII assembly, while functional PSI complexes were assembled, albeit with slightly altered electron transfer kinetics with respect to the wild-type complex (Bautista et al., 2005). In agreement with the higher sensitivity of PSII assembly to Car availability, Trebst and Depka (1997) reported a specific effect on the synthesis of the D1 subunit of PSII RC upon treatment with phytoene desaturase inhibitors. On the other hand, it has recently been reported that in lycopene-β-cyclase mutants of Arabidopsis (Arabidopsis thaliana) that have a decreased amount of β-carotene (bound to the RC) with respect to most of the xanthophyll pool pigments (bound to the LHCs), the level of accumulation of PSI complexes, particularly that of the LHC I complement, was more affected that that of PSII, probably also because of an increased sensitivity to photodamage of mutated PSI RC (Cazzaniga et al., 2012; Fiore et al., 2012).
In this investigation, we have studied the accumulation and functionality of the major chromophore-binding complexes of the photosynthetic apparatus, PSI, PSII, and Cyt b6f, in a Car-less mutant of the green alga Chlamydomonas reinhardtii (FN68) that is blocked at the first committed step of Car biosynthesis, namely, phytoene synthesis (McCarthy et al., 2004). Although the mutant is incapable of growing under phototrophic or photomixotrophic conditions, it can grow in complete darkness on a medium supplemented with a carbon source. Here, we show that the PSII core and antenna complexes fail to accumulate in the mutant and that the Cyt b6f complex accumulates to approximately one-tenth of the wild-type level. On the other hand, the PSI reaction center accumulates in FN68 and possesses electron transfer properties that are remarkably similar to those of wild-type PSI. Interestingly, we find that the level of PSI accumulation differs in other phytoene synthase null mutants, suggesting that additional mutations in one or other of these strains affect PSI stability. Nevertheless, our findings demonstrate that Cars are not required for either the assembly or the functionality of PSI in vivo.
The Car Deficiency and Light Sensitivity of FN68 Are Due to a Lesion in the Phytoene Synthase Gene
The FN68 mutant of C. reinhardtii was isolated in the 1970s as an extremely light-sensitive strain blocked in the biosynthesis of phytoene, the first product of the Car biosynthetic pathway (Foster et al., 1984). FN68 has been described as a "colorless" or "white" mutant (Foster et al., 1984; McCarthy et al., 2004), although the strain is in fact pale green due to the presence of Chl a (Fig. 1). Our pigment analysis has confirmed that FN68 lacks any detectable phytoene or other Cars and has only trace (less than 2%) amounts of Chl b, but it accumulates approximately 20% of wild-type Chl a levels, (Supplemental Fig. S1). This value is comparable with the approximately 16% Chl a reported by McCarthy et al. (2004), although the precise amount of Chl a in FN68 appears to be dependent on cell age (data not shown). A molecular analysis of FN68 and other Car-less mutants of C. reinhardtii by McCarthy et al. (2004) revealed that all these mutants carry point mutations in the nuclear gene (PSY) encoding phytoene synthase. In the case of FN68, the lesion is a frame-shift mutation in exon 2, which also results in the loss of an SphI restriction site (Supplemental Fig. S2). In order to confirm that this mutation was exclusively responsible for the observed phenotype, we transformed FN68 with a 4.4-kb PCR-amplified genomic fragment containing the wild-type PSY. Transformants were selected based on their ability to form colonies in the light. As shown in Figure 1, the FN68::PSY transformants have restored pigmentation, are insensitive to light, and are capable of phototrophic growth (although phototrophic growth overall is slower than in the wild type, probably as a result of unrelated secondary mutations acquired by FN68 over 40 years of maintenance of the strain in the dark). The presence of the introduced wild-type PSY allele, along with the original mutant allele, in each transformant line was confirmed by molecular analysis using the SphI polymorphism as a marker (Supplemental Fig. S2).
Cars Are Required for the Accumulation of PSII and LHCs But Not PSI
Figure 2 shows the absorption spectra of whole cells of C. reinhardtii recorded at room temperature (Fig. 2A) and the fluorescence emission spectra recorded at 77 K (Fig. 2B). Spectra from FN68 are compared with those of the wild type and also the mutant Fud7-P71, which lacks PSII and most of the LHC complement but harbors wild-type amounts of PSI (Byrdin et al., 2006). The absorption spectrum of the wild-type strain displays a pronounced shoulder at 645 nm, arising from absorption of Chl b present in the LHCs, whereas this feature is absent in the Fud7-P71 and FN68 spectra. The contribution of carotene, and of Chl b, to the wild-type spectrum is seen as a strong absorption between 450 and 500 nm. Again, this feature is absent in the FN68 spectrum and is reduced in the Fud7-P71 mutant, where only PSI Cars are present. The fluorescence emission spectrum from the wild type shows the characteristic peaks, at 685, 695, and 715 nm, that are assigned to PSII (685/695 nm) and PSI (715 nm); a shoulder is also observed at 682 nm and reflects the preequilibrated emission from LHC II (Butler, 1978; Rijgersberg and Amesz, 1980). FN68 lacks the PSII signals but shows a principal broad band at 715 nm arising from functional PSI, indicating that, at least at low temperature, energy transfer within the Chl a bound to the PSI core antenna complexes is not perturbed to any significant extent. The Fud7-P71 spectrum is very similar to that of FN68 at wavelengths longer than 690 nm, but the former shows a clear peak at 682 nm, which arises from the residual fraction of antenna, most likely disconnected from the reaction centers, present in this strain.
Figure 2C shows the separation of photosynthetic complexes on a nondenaturing gel following solubilization of thylakoid membranes with the mild detergent β-n-dodecyl maltoside. In the case of wild-type membranes, three main bands can be resolved on the gel, and these are assigned as nondissociated PSI/PSII supercomplexes, the PSI-LHC I supercomplex, and the monomeric LHC complexes (Supplemental Fig. S3). In the FN68 mutant, only a single band is observed on the nondenaturing gel, which migrates slightly below the PSI-LHC I supercomplex and which is assigned to the PSI core complex. This indicates that both the PSII core and the LHC I and LHC II complexes accumulate to a level that is below the detection sensitivity of the technique (Fig. 2C). Moreover, the lack of PSII, as well as the LHC complement in FN68, is confirmed by western-blot analysis, using antibodies directed against the PSII core protein D1 and outer antenna complexes of PSII and PSI, respectively (Fig. 2D). In contrast, the accumulation of two of the core PSI subunits (PsaD and PsaF) is seen in FN68, indicating PSI accumulation in the mutant (Fig. 2D). This level of PSI was judged to be approximately 25% of the wild type, as shown in Supplemental Figure S4, although the actual level appears to vary depending on cell age (data not shown). The absence of Cars, therefore, appears not to prevent the stable accumulation of PSI, a complex that contains approximately 22 β-carotene molecules (Jordan et al., 2001; Ben-Shem et al., 2003). Their absence, however, does result in a failure to accumulate the PsaN subunit. This peripheral subunit is bound by weak electrostatic interactions to the lumenal face of PSI, and PsaN does not bind any of the β-carotenes itself (Ihalainen et al., 2002). It is most likely that either structural distortions at the lumenal face are responsible for a failure of PsaN binding, leading to the degradation of the unassembled subunit, or the nucleus-encoded subunit is not imported correctly into the thylakoid membranes, and therefore is degraded, since this process is ΔpH (for change in pH) dependent (Nielsen et al., 1994) and photosynthetic electron transfer (ET) is severely impaired in FN68. In both cases, the steady-state level of PsaN will fall below the level of detection.
In contrast to PSI, the stable accumulation of the PSII complex appears to be dependent on one or more of the 12 β-carotene molecules present in the core of PSII (Guskov et al., 2009; Umena et al., 2011). The failure of PSII accumulation in Car-deficient mutants, or in plant seedlings treated with the Car inhibitor norflurazon, has been noted previously (Trebst and Depka, 1997; Humbeck et al., 1989; Bolychevtseva et al., 1995; Bautista et al., 2005; Inwood et al., 2008). Similarly, the absence of carotenes severely impairs the accumulation of both LHC I and LHC II, which substantiates the results obtained in vitro and points toward a crucial structural role of xanthophylls in promoting and/or stabilizing the folding of these complexes (Plumley and Schmidt, 1987; Sandonà et al., 1998).
Rescue of FN68 by transformation with wild-type PSY results in the restoration of wild-type levels of both PsaN and D1 as well as the LHC complement for both PSII and PSI, confirming that it is the absence of Cars that is the cause of the observed destabilization effects and the resulting accelerated degradation of the unassembled subunits (Supplemental Fig. S5).
Stable Accumulation of the Cytb6f Complex Is Affected by Loss of the Single β-Carotene Molecule
The Cyt b6f complex contains a single Car molecule of unknown function (Kurisu et al., 2003; Stroebel et al., 2003). As shown in Figure 2D, the loss of this Car has a significant effect on the steady-state level of the complex, as judged by the accumulation of the Cyt f subunit in FN68. The level of Cyt b6f was investigated further by comparing the accumulation of other subunits in the mutant with the wild type: a Cyt b6f mutant lacking subunit IV (ΔpetD) and the Fud7-P71 strain using antibodies directed against Cyt f, Cyt b6, and subunit IV (Fig. 3A). In addition, parallel gels were stained with the heme stain 3,3′,5,5′-tetramethylbenzidine to reveal the Cyt-binding subunits of the complex (Fig. 3B). As observed for the Cyt f subunit, the other subunits show only a low level of accumulation (below 15%), although the actual amount was found to be variable between different cell preparations (e.g. compare FN68 Cyt f levels in Fig. 3 with those in Supplemental Fig. S5). It is likely that this reflects differences in the growth phase of the FN68 cultures, since it has been shown that Cyt b6f levels decrease significantly in other mutants affected in the complex once cells start to enter stationary phase (Choquet and Vallon, 2000).
The Car-Minus Cytb6f Complex in FN68 Is Functional
We investigated whether the loss of the Car molecule affected the functionality of the remaining Cyt b6f by monitoring the turnover of the complex in response to a single, saturating, turnover flash. Cyt f is mainly reduced under dark, microaerobic conditions. Consequently, the kinetics of Cyt f oxidation by plastocyanin can be monitored spectroscopically following actinic laser-flash excitation of PSI. FN68 was compared with the Fud7-P71 strain, which contains a wild-type amount of both PSI and Cyt b6f complexes. Measurements were performed in this strain, because the absence of the PSII core subunit and the low antenna contents greatly reduce background absorption, resulting in a large increase in the signal-to-noise ratio, and also allows measurements in the near UV using whole cells (Guergova-Kuras et al., 2001; Byrdin et al., 2006). Moreover, apart from the accumulation of Cyt b6f and the residual presence of a small fraction of LHCs, its thylakoid composition closely matches that observed in FN68.
In Figure 4, A and B, the kinetics recorded at several wavelengths in the region of maximal absorption of the more diagnostic Cyt α and the β-bands are shown. Optical transients were analyzed by a global fitting routine (400–600 nm) to extract the decay time constants and decay-associated spectra (DAS), which are shown in Figure 4, C and D. The DAS are internally normalized by the maximal bleaching of the difference spectrum associated with the metastable electron donor P700 (P700+-P700), which peaks at 430 nm.
In the control and the Car-deficient strains, three kinetic components were required to describe the kinetics following a single turnover flash. Two of these have virtually identical DAS and reflect the heterogeneous kinetics of P700+ reduction by plastocyanin (Fig. 4, C and D). Their associated lifetimes are 6.5 and 67 μs, in agreement with previous investigations of C. reinhardtii (Santabarbara et al., 2009; for review, see Haehnel, 1984). The slower component has a 354-μs lifetime, and its DAS is fully consistent with the oxidation of Cyt f (for review, see Hope 2000); it displays a maximum at approximately 554 nm, which is a characteristic feature of Cyt f (Fig. 4, C and D, insets), and a major positive peak at 425 nm in the Cyt γ band (Fig. 4, C and D). A nondecaying (i.e. lifetime [τ] > 1 ms) component with large amplitude is also observed, which reflects, principally, the reduction of oxidized Cyt f and the oxidation of Cyt b. Although the lifetime of all three components developing in the hundreds of microseconds time scale is virtually the same in the FN68 and the Fud7-P71 strains, the DAS spectrum attributed to oxidation of Cyt f has an approximately 10-fold smaller amplitude in FN68, after normalization to P700+ bleaching. Thus, this characterization of Cyt b6f catalytic activity following a single turnover flash indicates that the residual fraction of the complex in the FN68 strain is functional.
Electron Transfer Reactions in the PSI RC of FN68
In order to understand the role of bound Cars in the electron transfer properties of the PSI RC, we have measured the kinetics of the oxidation of the secondary electron acceptor, phylloquinone A1, in whole cells and at room temperature. Laser-induced optical transients were acquired in the 360- to 570-nm interval (selected kinetics are shown in Supplemental Fig. S6) and globally fitted to extract time constants (Table I) and DAS (Fig. 5). The kinetic traces recorded in the FN68 and Fud7-P71 strains are globally described by three lifetime components. Two of these decays fall in the nanosecond time scale and have values of 24 and 258 ns in the Fud7-P71 strain and 22 and 316 ns in the FN68 mutant. Thus, while the apparent rate of the fast component of A1− oxidation, which is associated with the PsaB-bound phylloquinone (A1B; Guergova-Kuras et al., 2001; Ali et al., 2006; Byrdin et al., 2006), has almost the same value in the FN68 and the control strain, the slow phase, which principally arises from the oxidation of the PsaA-bound A1A quinone (Guergova-Kuras et al., 2001; Byrdin et al., 2006; Santabarbara et al., 2008, 2010c), is affected, although not in a dramatic fashion, by the absence of Cars in the complex.
The DAS of the two nanosecond components assigned to A1− oxidation show clear positive peaks at approximately 480 and 510 nm and negative peaks at approximately 500 nm in the control strain, which are attributed to electrochromism (ECS) of RC pigments in response to the field generated by the electron transfer reactions. In contrast, a single broad and asymmetric feature peaking at approximately 482 nm is observed in FN68. The absence of fine spectral structure at wavelengths longer than 465 nm in the DAS of both the 24- and 316-ns components is simply the result of the absence of Cars in the PSI RC of the FN68 mutant. At the same time, it highlights the fact that the electrochromic signal, in this wavelength interval, has an important, and at several wavelengths, dominant, contribution from Chl. An intense signal peaking at approximately 450 nm, which is observed in both the 24- and 316-ns DAS of FN68, can be attributed primarily to the electrochromic response of Chl(s) near the phylloquinone cofactors. This feature is more resolved in FN68 than in the control strain, probably due to the disappearance of overlapping ECS signals from Cars. Interestingly, as noted previously by Bautista et al. (2005) in Car-deficient mutants of Synechocystis sp. PCC 6803, the maximum bleaching is blue shifted by 5 nm in the 24-ns DAS (450 nm) with respect to the 316-ns DAS (455 nm). This difference can be interpreted in terms of a different local environment of the Chl(s) near A1A and A1B, providing further evidence of the assignment of these kinetic phases.
The DAS of the approximately 6-μs decay components reflects the reduction of P700+ by plastocyanin (PC). The P700+PC-P700PC+ difference spectra recorded in the control and FN68 strains are very similar at wavelengths lower than 460 nm. However, at longer wavelengths, clear differences are seen, which can also be attributed to the absence of Cars in the RC of FN68. The spectrum of P700+ reduction exhibits only a broad feature peaking at about 490 nm in FN68, whereas in Fud7-P71 RC, features originating from electrochromic shifts in Chls near P700 are observed at 485, 505, and 530 nm.
The results obtained in vivo in the FN68 strain of C. reinhardtii are qualitatively in excellent agreement with a previous investigation by Bautista et al. (2005), who studied the effect of mutations at the level of phytoene desaturase and ζ-carotene desaturase on isolated PSI from Synechocystis sp. PCC 6803. Even though these mutations are downstream to phytoene synthase, they still result in strong depletion of Car from isolated PSI particles (Bautista et al., 2005). In PSI containing either only phytoene or ζ-carotene, it was shown that the lifetime of the rapid phase of A1− oxidation was substantially unaffected, but the slow component displayed a moderate lengthening (Bautista et al., 2005), as observed in whole cells of FN68 (Fig. 5). However, while the relative amplitudes of 20- and 250-ns DAS were substantially unaltered in the Car biosynthetic mutants of Synechocystis sp. PCC 6803 compared with the wild type, we observed a slight decrease in the relative contribution of the approximately 20-ns phase with respect to the 250-ns phase in the FN68 strain. Based on the kinetic measured at 390 nm (and the overall DAS in the 360- to 400-nm range), the amplitude ratio is shifted from 1:1.4 (20:250 ns) in the control strain RC to 1:2.3 in the FN68 Car-depleted complex (Table I). As a large perturbation of extinction coefficients associated with the absorption of either the semiquinone or quinone form of A1A and A1B appears unlikely (note that the total absorption difference with respect to P700+ is substantially unmodified in FN68 with respect to the control strain), the simpler rationalization for the small decrease in amplitude of the approximately 20-ns phase with respect to the approximately 250-ns phase is a subtle redistribution of the relative amount of electrons (statistically) transferred by the ETA and ETB redox chains in favor of the former. That such redistribution can indeed occur has been demonstrated in site-directed mutants affecting either the coordination of ET cofactors located upstream of A1 (Byrdin et al., 2006; Li et al., 2006; Santabarbara et al., 2008) or altering the strength of hydrogen bonding to A1 (Santabarbara et al., 2010c; Srinivasan et al., 2011).
Pulsed Electron Paramagnetic Resonance Analysis of the PSI RC in FN68 at 265 and 100 K
The kinetics of electron transfer in the PSI reaction center lacking carotenes was also investigated by time-resolved pulsed electron paramagnetic resonance (EPR), a technique complementary to transient absorption, which has also been extensively used to characterize the secondary/ternary ET reactions in PSI (Santabarbara et al., 2005a; Srinivasan and Golbeck 2009). When applied to laser-flash excitation, pulsed EPR has the advantage of being selective for spin-correlated radical pairs, such as [P700+A1−], which is by far the dominating signal detected in thylakoid membranes (Santabarbara et al., 2005b, 2006, 2010b). Therefore, experiments can be performed in the wild-type background without loss of sensitivity. A summary of the comparative analysis of the kinetics of ET in PSI monitored by time-resolved electron spin echo (ESE) spectroscopy in the wild type and FN68 is presented in Table II (a more detailed discussion and the experimental data are presented in Supplemental Fig. S7).
The decay of the ESE monitored at 265 K (close to room temperature) provides information on [P700+A1A−] only, since [P700+A1B−] is not detected due to the temporal resolution of the spectrometer (approximately 50 ns). The analysis of time-resolved ESE experiments is in qualitative agreement with the results obtained by optical spectroscopy, indicating a slower oxidation of A1A− in FN68 compared with the wild type. However, the effect on the kinetics is more pronounced when monitored at 265 K than at room temperature, suggesting that the absence of carotene in PSI might influence the activation barrier of this reaction (see the discussion of Supplemental Fig. S6 for further details).
Moreover, we have studied the decay of ESE signal at 100 K. At this temperature, the ESE signal provides information concerning charge recombination reactions rather than forward ET (Schlodder et al., 1998; Agalarov and Brettel 2003; Santabarbara et al., 2005a, 2010b). Both in the wild type and in FN68, under conditions in which the successive electron acceptor iron-sulfur cluster (FX) is initially oxidized, the signal is dominated by an approximately 20-μs lifetime that arises from [P700+A1A−] (Santabarbara et al., 2005b, 2006). Upon FX prereduction, the decay becomes biphasic, and it is characterized by lifetimes of approximately 2 and 20 μs, both in the wild type and FN68. The approximately 2-μs phase was shown to arise from the [P700+A1B−] radical pair (Santabarbara et al., 2005b, 2006, 2010b). The need to reduce FX to observe [P700+A1B−] was interpreted in terms of a difference in the driving forces for A1A− and A1B− oxidation, being thermodynamically unfavorable (i.e. the standard free energy difference > 0) for the former and favorable (standard free energy difference < 0) for the latter (Santabarbara et al., 2005a, 2010a). Such "functional asymmetry" in A1A/B energetic properties appears to be qualitatively conserved in FN68. This is also confirmed by the observation that PSI assembled in FN68 is capable of reducing the terminal acceptor iron-sulfur clusters FA/B at low temperature (Supplemental Fig. S8). Finally, the relative amplitudes of the approximately 2- and 20-μs phases (Table II) provide a measure of the statistical utilization of ETB and ETA (Santabarbara et al., 2005b, 2006, 2010b). The analysis of charge recombination reaction monitored by the ESE decay also shows a redistribution in favor of ETA, although it is less pronounced than that observed at room temperature.
Stability and Functionality of the Electron Transfer Components in the Absence of Carotenes
As discussed above, the Cyt b6f complex binds one β-carotene and one Chl a molecule as prosthetic groups in addition to cofactors active in electron transfer reactions (Kurisu et al., 2003; Stroebel et al., 2003). Spectroscopic investigations (Peterman et al., 1998; Dashdorj et al., 2005) of the bound chromophores indicate that the β-carotene molecule does not directly interact with the Chl a. Thus, the β-carotene bound to Cyt b6f should be incapable of directly quenching the Chl triplet state. The function of these two pigment molecules within the complex are at present unknown, although it has been suggested that β-carotene might play a photoprotective role by directly scavenging singlet oxygen (Dashdorj et al., 2005). It has also been proposed that the β-carotene might play a structural role, acting as a lipid analog and stabilizing the interaction between transmembrane α-helices of subunit IV, Cyt b6, and the minor subunits encoded by the petL, petM, and petN genes (Smith et al., 2004).
The impairment of the accumulation and function of Cyt b6f in the FN68 mutant provides support for the latter suggestion. As the strain was grown in absolute darkness, it is extremely unlikely that the defect in complex accumulation is the result of photooxidative stress. In fact, the growth of FN68 is inhibited even upon illumination at very low photon flux densities (10 μE m−2 s−1) in a nitrogen atmosphere supplemented with CO2 but deprived of oxygen (Supplemental Fig. S9). However, given the very similar structure of the mitochondrial Cyt bc1 complex (Crofts, 2004), the effect of the absence of the single β-carotene in Cyt b6f appears to be quite dramatic, and it could be an indirect result of a more complex interplay between physiological processes affected by the absence of Cars in the mutant. In fact, the residual amount of complex accumulated appears to be, kinetically, as efficient as the wild type. As the assembly of Cyt b6f in vivo is assisted by a complex folding machinery (Nakamoto et al., 2000), it is possible that β-carotene, rather than affecting the stability of the complex per se, is required at some stage during the assembly process.
The requirement of carotenes for the stabilization of folding is probably the reason for the lack of any significant accumulation of either the LHC complexes or the RC subunits of PSII. In these cases, not only are Cars much more abundant cofactors (three to four molecules per protein monomer) compared with the single molecule bound to Cyt b6f, but they have been shown to be fundamental for in vitro refolding of LHCs (Plumley and Schmidt 1987; Paulsen et al., 1993; Sandonà et al., 1998).
The only exception to the strict requirement for Cars to obtain a detectable level of accumulation of functional photosynthetic complexes is the RC of PSI, which is not only assembled but appears to operate with similar efficiency to the wild-type RC in the FN68 strain. These results are in agreement with the previous study of Bautista et al. (2005), who investigated Synechocystis sp. PCC 6803 mutants that contained only short-chain Cars. On the other hand, an increased sensitivity of PSI to photooxidative damage and a lower level of PSI accumulation have been reported in mutants of Arabidopsis with a lower β-carotene content with respect to xanthophylls compared with the wild type (Cazzaniga et al., 2012). However, the mutation analyzed by Cazzaniga et al. (2012) is far less dramatic than those investigated by Bautista et al. (2005) and in this study, where strains were grown in total darkness so that photoinhibition and photoprotection do not play a role in complex accumulation.
However, the variation in Chl levels observed among different phytoene synthase mutants including FN68 by McCarthy et al. (2004) suggests differences in the levels of accumulated PSI in different mutant backgrounds. This is confirmed by western-blot analysis of four mutants: FN68, w7, lts1-207, and lts-209, in which the PSI levels are estimated to be 25%, 15%, less than 1%, and less than 1%, respectively (Supplemental Fig. S4). Whether this reflects compensatory mutations in FN68 and w7, allowing increased accumulation of a PSI complex lacking carotenes (e.g. a mutation affecting a protease involved in PSI turnover), or additional mutations in the two lts1 strains (or their parent background), leading to increased PSI instability, is not clear and requires further genetic analysis. Nonetheless, the ability of FN68 to assemble a functional PSI RC despite the fact that the PsaA/PsaB heterodimer alone binds 22 β-carotene molecules, making it the second most abundant cofactor after Chl a (approximately 80 molecules per PsaA/PsaB heterodimer), is an intriguing observation. This suggests that the folding of PSI RC subunits in the thylakoid membrane is either substantially different from that of all other photosynthetic complexes or the folding of PSI RC polypeptides is sufficiently stable and the assembly of PSI can take place, to some extent, even in the absence of large amounts of cofactors. In this respect, it is interesting that the electron transfer chain of PSI maintains its functionality even in complexes in which the majority of cofactors have been extracted by treatment with organic solvent (Itoh et al., 2001). This points toward a particular thermodynamic stability of the secondary and ternary structures of the main constituents of the PSI RC compared with PSII and Cyt b6f. Comparison of the crystallographic models of the PSI and PSII core complexes reveals a very similar arrangement of their transmembrane α-helices, which is the most abundant secondary motif. However, while the majority of pigments are bound to PsaA and PsaB in the PSI core, PSII chromophores are bound to four major subunits: D1, D2, CP43, and CP47 (Jordan et al., 2001; Ben-Shem et al., 2003; Guskov et al., 2009). This difference in the arrangement of RC subunits may be the basis for the superior stability of PSI RC, as all the transmembrane secondary structure elements are linked via the lumen and stroma-exposed loops of the PsaA and PsaB polypeptides, while weaker hydrophobic interactions play a larger role in the stabilization of the D1∙D2∙CP43∙CP47 multimer that represents the bulk of the internal antenna and the catalytic core of PSII.
Strains, Media, and Growth Conditions
The wild-type strain CC-1021 (mt+) and the Car-less strains FN68 (mt–) CC-2682, w-7 CC-2843, lts1-207 (mt+) CC-4113, and lts1-209 (mt+) CC-4115 were obtained from the Chlamydomonas Center (www.chlamy.org). The ΔpetD (mt+) deletion strain and Fud7-P71, a double mutant with a ΔpsbA deletion and reduced light-harvesting antenna (Byrdin et al., 2006), were from the Paris Culture Collection. All strains were maintained on Tris-acetate-phosphate medium under continuous illumination at a photon flux density of approximately 10 μE m−2 s−1 or, for the Car-less strains, in complete darkness. Cells were harvested by centrifugation at 1,500g for 5 min and immediately suspended in the appropriate medium for analysis.
Nuclear Transformation of FN68
The mutant was transformed using the glass beads method as described by Purton and Rochaix (1995) using a 4.4-kb PCR product containing the entire PSY gene amplified from wild-type genomic DNA using primers PSYg5′ and PSYg3′ (Supplemental Table S1). Transformant colonies were selected on Tris-acetate-phosphate plates under light of approximately 40 µE m−2 s−1 and scored for the presence of the wild-type PSY allele by PCR analysis using primers PSY5′ and PSY3′ (Supplemental Table S1) followed by digestion of the PCR product with SphI.
Preparative and Analytical Techniques
Thylakoid Purification
Cells were harvested by centrifugation during the logarithmic growth phase and suspended in minimal growth medium to a concentration equivalent to 1 optical density at 680 nm. The thylakoids were purified by Yeda press (100 kg cm−2 fractionation as described previously; Santabarbara et al., 2007) with the following modifications: the harvesting buffer (0.6 m Suc, 100 mm Tricine-NaOH, pH 7.8, 20 mm NaCl, 10 mm MgCl2, and 2 mm CaCl2) also contained DNase I (100 μg mL−1), phenylmethylsulfonyl fluoride (30 μg mL−1), and bovine serum albumin (1%, w/v). The lysate was centrifuged at 250g for 5 min to remove unbroken cells and starch (all operations were performed either at 4°C or on ice and under darkness or very dim green light). The supernatant was then subjected to fractional centrifugation at 3,500g (10 min), 15,000g (10 min), and 35,000g (15 min). All pellets were washed in a buffer containing 0.1 m Suc, 20 mm Tricine-NaOH, pH 7.8, 10 mm NaCl, 5 mm MgCl2, and 2 mm CaCl2, centrifuged under the respective precipitation conditions, and finally suspended to a concentration equivalent to 500 μg mL−1 Chl a in 30 mm Tricine, pH 7.8, 10 mm NaCl, and 5 mm MgCl2 for subsequent biochemical analysis. The light 35,000g fractions were used for comparative analysis of the wild type and FN68 (Fig. 2). The 3,500g thylakoid fraction was used for comparison of C. reinhardtii and spinach (Spinacia oleracea; Supplemental Figure S3).
SDS-PAGE and Western-Blot Analysis
Total protein extracts were prepared by growing cells to a density of approximately 4 × 106 mL−1, centrifuging at 6,000g for 5 min, and resuspending the pellet in 0.8 m Tris-HCl, pH 8.3, 0.2 m sorbitol, and 1% β-mercaptoethanol to a final concentration of approximately 8 × 107 mL−1. Aliquots (50 µL) of boiled samples were separated by electrophoresis on a SDS-15% polyacrylamide gel and transferred to nitrocellulose membranes. Membranes were incubated with primary polyclonal antibodies at a dilution of 1:1,000. Membranes were incubated with a goat anti-rabbit secondary antibody at 1:5,000 dilution, conjugated to horseradish peroxidase, and enhanced chemiluminescence was used for autoradiographic detection (GE Healthcare).
In SDS-PAGE experiments aimed at resolving Cyt b6f subunits, the samples were separated on a 12% to 18% gradient polyacrylamide gel containing 8 m urea. The gel was stained with 3,3′,5,5′-tetramethylbenzidine and revealed with hydrogen peroxide (Thomas et al., 1976).
Deriphat-PAGE was carried out essentially as described by Peter et al. (1991) with the following modifications: a thin 4% acrylamide stacking gel was poured upon the 9% acrylamide running gel, and Deriphat concentration in the running buffer was reduced to 0.1% (w/v). For simultaneous analysis of wild-type and FN68 thylakoids, samples at equal total Chl concentration (10 μg) were solubilized using β-n-dodecyl maltoside at a final detergent:Chl weight ratio of 80:1. After 20 min of incubation on ice, samples were spun at 14,000 rpm for 30 min in a benchtop refrigerated centrifuge (4°C) to remove insolubilized material, and the supernatant was immediately applied to gels. Gels were run at a constant voltage of 50 V in the dark at 4°C.
EPR Spectroscopy
Continuous-Wave EPR
Continuous-wave EPR spectra were recorded on a JEOL RE1X spectrometer equipped with a cylindrical resonator that allows illumination of the sample in the spectrometer. The instrument is fitted with a cryostat (Oxford Instruments; ESR9) cooled with liquid helium, and the temperature is controlled with an ITC 5 unit (Oxford Instruments).
Time-Resolved EPR
Pulsed EPR kinetics was measured in a Bruker ESP580 spectrometer, operating at X-band (approximately 9.5 GHz), fitted with a dielectric resonator (Bruker model 1052 DLQ-H 8907) that has been described previously in detail (Santabarbara et al., 2006). The temporal resolution of the spectrometer is approximately 50 ns, limited by laser jitter and cavity ringing.
The samples at a Chl concentration equivalent to 2 mg mL−1 were placed in calibrated quartz tubes of 3 mm i.d. and incubated under an argon atmosphere for 30 min, on ice, prior to any chemical addiction. Photochemical reduction of FA/B and FX was obtained as described previously (Santabarbara et al., 2005b, 2006).
Optical Spectroscopy
Fluorescence Emission Spectra
Fluorescence emission spectra were measured in a home-built apparatus: the excitation source consisted of a diode laser emitting at 465 nm, and the emission was collected by a CCD detector after dispersion through a spectrograph (Ocean Optics; HR4000) with a resolution of 1 nm. The fluorescence was excited, and the emission collected was from the surface of the sample through a fiber-optic system. The temperature was set by directly immersing the sample, at a concentration equivalent to 1 absorbance unit cm−1 at 680 nm and contained in a metal holder, into liquid nitrogen. The spectra were recorded after 10 min of incubation in the liquid nitrogen bath.
Absorption Spectra
Absorption spectra were measured in a home-built single-beam spectrometer, in the white light from a halogen lamp (Eurosep) shone on the sample, placed in a 1-cm path-length quartz cuvette, via a fiber-optic bundle, and the transmission was collected by a CCD detector (Ocean Optics; HR4000). The absorption spectrum was calculated from the recordings of the transmission. The optical resolution of the apparatus is 1 nm.
Time-Resolved Optical Spectroscopy
Time-resolved different absorption measurements in the submicrosecond time scale were performed employing a home-built pump-probe spectrophotometer that allows measurements to be performed in scattering biological material; the setup has been discussed previously in detail (Beal et al., 1999). In brief, the sample was excited at 700 nm, with a 7-ns pulse (full width at half maximum), obtained by pumping a dye laser (LDS 698) with a frequency doubled neodymium-doped yttrium aluminum garnet laser (Brilliant; Quantell). The intensity of the actinic pulse was set to a level ensuring single-turnover excitation of approximately 70% of the RCs. The kinetics were probed at different wavelength by 5-ns pulses (full width at half maximum) obtained by pumping an Optical Parametric Oscillator (Panther OPO; Continuum) with a frequency tripled neodymium-doped yttrium aluminum garnet laser (Surelite I; Continuum). The temporal resolution of the spectrometer is approximately 5 ns before instrument function deconvolution.
The cells were suspended at concentration equivalent to approximately 2 optical density cm−1 at 678 nm in a solution containing HEPES-NaOH buffer at pH 7 and 20% (w/v) Ficoll to avoid sedimentation of the sample during the measurements. The uncoupler carbonylcyanide p-trifluoromethoxy phenylhydrazone (10 μm) was added to the sample to avoid the establishment of long-lived ion fields across the thylakoid membrane.
Data Analysis
Global fittings of single-turnover decays associated with PSI and Cyt b6f turnover were performed using a home-written routine operating in MatLab (Cambridge Soft) that minimizes the sum of squared residues by a combination of Simplex and Levenberg-Marquardt algorithms. The time decay of the ESE has been fitted with a sum of exponential functions as described previously (Santabarbara et al., 2005b).
Supplemental Data
The following materials are available in the online version of this article.
Supplemental Figure S1. HPLC analysis of the wild type and the FN68 mutant of C. reinhardtii.
Supplemental Figure S2. Restriction and PCR analysis of the FN68 mutation.
Supplemental Figure S3. Deriphat-PAGE and immunodecoration of native gels analysis.
Supplemental Figure S4. Quantification of PSI levels in FN68 by immunoblot analysis.
Supplemental Figure S5. Immunoblot analysis of FN68 transformed with the wild-type PSY gene.
Supplemental Figure S6. Kinetics of light-induced electron transfer reaction in PSI.
Supplemental Figure S7. Kinetics of light-induced electron transfer reaction at 265 and 100 K monitored by pulsed EPR spectroscopy.
Supplemental Figure S8. Continuous-wave EPR light-minus-dark difference spectra.
Supplemental Figure S9. Comparison of growth under different light intensities and atmosphere composition.
Supplemental Table S1. Oligonucleotide sequences from PSY used for PCR amplification.
We acknowledge the early contribution made to this work by the late Mike Evans (University College London). We are grateful to Paul Fraser (Royal Holloway, University of London) for help with the HPLC analysis of FN68. We thank Yves Pierre (Institut de Biologie Physicho Chimique) for his technical advice and Giuseppe Zucchelli (Istituto di Biofisica, Consiglio Nazionale delle Ricerche) for stimulating discussions. We thank Jean-David Rochaix (University of Geneva) for the PsaA, p11, and p15 antibodies and Peter Nixon (Imperial College London) for the D1 antibodies.
The authors responsible for distribution of materials integral to the findings presented in this article in accordance with the policy described in the Instructions for Authors (www.plantphysiol.org) are: Stefano Santabarbara (stefano.santabarbara{at}cnr.it) and Saul Purton (s.purton{at}ucl.ac.uk).
1 This work was supported by the Biotechnology and Biological Sciences Research Council (grant no. CO0350 and a studentship to K.A.), by a Royal Thai Government Scholarship (to T.W.), and by the U.S. Department of Energy (grant no. DE–FG02–08ER15989 to K.E.R.).
[C] Some figures in this article are displayed in color online but in black and white in the print edition.
[OA] Open Access articles can be viewed online without a subscription.
[W] The online version of this article contains Web-only data.
- Car
- carotenoid
- light-harvesting complex
- Cyt
- cytochrome
- Chl
- chlorophyll
- decay-associated spectra
- electron paramagnetic resonance
- electron spin echo
- τ
- lifetime
- Received August 9, 2012.
- Accepted November 14, 2012.
- Published November 16, 2012.


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We're celebrating being named 10th out of 100 in the Sunday Times Best Not-for-Profit Organisations to work for list.
As well as being in the top 10 we've also received Three Star accreditation which is the highest standard of workplace engagement and reflects 'extraordinary' levels of workplace engagement.
The Sunday Times Best Companies survey is the UK's benchmark for rating and celebrating the very best employers to work for.
The award is based on the responses from our team to a confidential questionnaire focusing on eight areas, including how good our organisation's leadership is, what our team thinks of our organisation, opportunities for personal growth, pay and benefits, relationships and general well-being. As the questionnaire is completely secure and anonymous, it gave our team the opportunity to express how they really felt.
Alex Heritage, Chief Executive, said: "To have been recognised as one of the UK's Best Not-For-Profit Organisations to work for is a real achievement. We recognise that when our staff feel positive about their job and the TU, they are much more likely to go out of their way to deliver a great service to our clients, which will ultimately benefit patients. The health, wellbeing and development of our staff is very important to us and I'm delighted that our work on these issues has been recognised."
"We try to ensure staff develop their talents and that there are lots of opportunities for them to learn and engage in our wider work. It's clear to us that delivering job satisfaction is not simply a question of wage, it's a matter of professional development, engagement and recognition of the value of our staff.
"One of the areas we specialise in is Organisational Development, and as well as developing our own staff, we're committed to growing a thriving and sustainable workforce of transformation professionals in the NHS for the NHS."
Audrey Abbott, Graduate Trainee Project Manager, said: "As a graduate it is really exciting to work for one of the UK's Best Not-For-Profit Organisations. The focus on learning and development means there are endless opportunities at the TU.
"One of my favourite aspects of working here is the importance and value placed on culture. All of our work (internal and external) centres around our 4 values (people focused, integrity, driven and empower), and these are seen every day in how we work together and with our clients."
Jonathan Austin, Founder and CEO of Best Companies, said: "I would like to congratulate the NHS Transformation Unit on their achievement of a Three Star Accreditation status and being named as one of the UK's Best Companies to Work for. Best Companies have been producing the Accreditation standard since 2006 to acknowledge excellence in the workplace. We are proud that organisations like the TU focus on their employees and have been rewarded with an engaged workforce who tell us that they're a best company to work for."

subset: commoncrawl


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Officially, summer ended at the beginning of March.
But this week's cold and rainy weather really heralded the beginning of Autumn. Did you have to dig out the wet weather gear? Maybe even a sweatshirt?
If you're anything like us, you spent the week indoors and noticed, ever so slightly, your summer glow beginning to disappear. Without daily doses of beautiful sunshine, your skin will begin to lose that golden tan and leave behind traces of the damage a summer in the sun can inflict.
Sunspots, freckles, tan lines, scars or fine wrinkles – all indications of what the harsh Australian sun can do to your skin whilst frolicking in the waves, or soaking in the sun at the park.
If you have had too much sun during summer, this is the perfect time to address premature aging from sun damage with an intensive treatment routine. Sound scary? Not even in the slightest! It's a great opportunity – whilst the weather begins to change – to kickstart a new skincare routine for the cooler months.
Here's how to restore your skin for Autumn in a simple, two step routine.
Step One: Brighten
Start with our Brighten Vitamin C Serum to even skin tone and pigmentation, whilst protecting your skin with antioxidants.
Our Brighten Vitamin C Serum:
- Contains a concentration of actives to lighten and brighten the appearance of the skin, and evens out the appearance of age spots and dark pigmentation.
- Contains Vitamin C enriched Kakadu Plum extract and Bergamot energise and revitalise the complexion, leaving it visibly softer, more supple and luminous.
- Uses antioxidants to protect against signs of oxidative stress and make skin feel younger and smoother.
- May result in effective wrinkle reduction and significant freckle and skin lightening in Asian, Hispanic, African American and Caucasian skin types according to human clinic trials.
To use: simply apply onto clean skin. Allow a few minutes for it to soak in, and then apply your daily/nightly moisturiser over the top.
Step Two: Replenish
Your next secret weapon will be our famous Replenish Elixir. Use this as a night-time treatment to boost the elastin and collagen that is broken down by too much exposure to the sun.
Our Replenish Elixir:
- Significantly nurtures collagen formation to visibly improve the appearance of fine lines and wrinkles.
- Contains naturally occurring omega 3, 6, 7 and 9 fatty acids and antioxidants to nourish and energise skin's appearance.
- An exceptional synergy of precious oils imparts a soft, velvety emollience, restoring balance, hydration and natural radiance.
To use: After allowing the Brighten Serum to sink into your skin, rub a few drops of Elixir in between clean fingers, and press gently into the skin on your face, paying special attention to the skin around your eyes, nose, and mouth.
As the weather cools down, you are going to also want to start paying special attention to dry skin. Make sure you work in regular exfoliation, and nourishing face masks, to keep your skin supple and healthy.
And don't forget – just because summer is over, doesn't mean protecting your skin needs to be too!
Make sure you always apply SPF+ moisturiser, and wear a hat if you are planning on being outdoors enjoying the beautiful autumn days.
Until next time,


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♥ Measurements approx. 22cm crown to edge, 11cm turn
Pink Mink Piece Pom Pom Hat
This special pink and brown, wool and mink beanie hat will make you bang up to trend this season! Made of 100% natural wool with a large turnover and chunky knit style, this hat will keep your head super cosy. Also includes a piece pom pom- not a solid ball so it is lighter to wear. Knitted in a simple rib pattern, large to fit a man or woman, it's the perfect gift!
♥ Measurements approx. 22cm crown to edge, 11cm turn
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1 in stock


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Peoples Party of Canada leader Maxime Bernier has hired a fossil fuel divestor as an Electoral District Association (EDA) coordinator in his national party office.
Daniel Tyrie authored this opinion piece for The Waterloo Region Record — an opinion piece that shares many talking points with the eco-radicals you see on campuses these days — demanding the University of Waterloo pull more than $68 million out of "polluting companies" like Shell, British Petroleum, and Exxon Mobil.
At a time when oil companies are fleeing Alberta, like the US-based Devon Energy, companies like BP own three oil sands assets in the Athabasca basin – Maxime Bernier has a staffer working for him that wanted companies operating in the oil sands to tank.
Alberta has a tough time being represented in Ottawa. Confederation has systematically oppressed the West, from equalization to the number of seats they have in Parliament – they are the country's underdog. It's part of the reason Alberta has been so excited about Maxime – he won the CPC leadership vote in Alberta by a landslide, he swept Calgary, most of Edmonton, and split the rural vote with Scheer.
But now we are seeing something different: Maxime Bernier —the "Albertan from Quebec" — hired an anti-oil sands activist, Tyrie worked closely with Fossil Free UW, a campus club hell-bent on stopping pipelines and suffocating Alberta. Bernier's staff now features someone who believed — just eleven months ago — that because the University of Waterloo has money in energy investments, that it is "responsible for devastating our climate and our future."
I contacted Daniel Tyrie about this, and he told me that he has since changed and that he was caught up in a sort of undergraduate climate hysteria. Tyrie said, "[He] now views fossil fuels and pipelines in particular as necessary for both economic stability and national unity in Canada."
The PPC told The Rebel that they were not aware of the article written by Tyrie at the time he was hired. They added that Tyrie was with Maxime at the recent United We Roll convoy in Ottawa, and:
"There is no reason to doubt his beliefs, which in any case cannot have any impact on PPC policies as he is only involved on the organizational side and not on policies."


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True North Troubadours perform in Abbotsford
BC Local News, May. 11, 2018 12:00 a.m.
Concert on Saturday, May 12 at House of James
The True North Troubadours will perform at the House of James coffeehouse in Abbotsford on Saturday, May 12.
The group consists of Joel Dietrich and Wayne and Diane Rempel. Rich vocal harmonies with acoustic guitar interplay define the essence of this engaging group.
Paying tribute to the folk music of the '60s, the True North Troubadours currently perform popular songs by artists such as Peter, Paul and Mary; Ian and Sylvia Tyson; Bob Dylan; Gordon Lightfoot and others.
The group is also supported by acoustic bassist Steve Labish, who lays down the groove with authority and passion.
"I've been a huge fan of Peter, Paul and Mary and it's such a delight to have True North Troubadours keep their music alive in such an enjoyable way," said Lando Klassen, House of James
The concert starts at 7 p.m. Tickets are $15 at the door. More information is available online at houseofjames.com
Source: BC Local News. The Abbotsford News


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For accuracy, fewer recalls, and clarity in dense breast tissue, 3D mammography is the top of the line. However, those who are looking for top-tier technology like this won't find just one option when they take to the market to find their next system. A number of offerings from Hologic, GE, Siemens, Fuji, and others are available either new or on the secondary market.
To help buyers looking for 3D mammo narrow down their options, we'll compare two options they're likely to come across in their search: the Dimensions 3D from Hologic and the Pristina from GE.
Hologic Dimensions 3D vs. GE Pristina
Image Quality
The image quality of the Dimensions and the Pristina are comparable. Both systems have a standard detector resolution of 100 microns. Hologic systems with their new Clarity HD detector appear to be pulling ahead at 70 microns, but these are not commonly found on the secondary market yet. Both detectors also offer the same 24 x 29 cm imaging area.1
Tomo-Guided Breast Biopsy
Both systems offer tomosynthesis breast biopsy guidance capabilities. The tomo guidance feature makes biopsies easier for physicians to perform and increases their ability to get to masses positioned in the posterior of the breast by eliminating the table thickness of a traditional prone biopsy system.
Tomo-guided biopsies are also more comfortable for patients. If a patient is disabled, injured, or otherwise limited in movement, the flexible positioning of an upright system can accommodate them.
While GE has grown their market share in mammography over the last few years, the behemoth is still Hologic. Hologic has been steadily increasing its 3D install base since 2011, and it has paid off in market penetration. Because of this, Hologic's engineering bandwidth and responsiveness for 3D mammo is unmatched. We aren't saying that GE service is bad, but any other 3D mammo manufacturer is hard-pressed to keep up with Hologic.
Neither of these systems are available used or refurbished at a level we would call common. No Pristinas have come into our purview. They are still rather new, and their install base is relatively small. Dimensions 3D units are more available, but at what we would call a trickle. Still, those in search of either system on the secondary market are far more likely to find a Dimensions 3D at this point.
Current Price Averages for Popular Digital Mammo Systems
A Note on Self Compression
With the Pristina, GE offers a feature package called "Self Compression". As the name suggests, the package puts the patient in control of the level of compression on their breast. The idea behind the feature is that the patient will be more comfortable and experience less pain during the procedure if they are "in the driver's seat".
Self Compression has been met with mixed reviews from physicians, however, as some patient's pain tolerance may not measure up to the level of compression that is needed to get an image of sufficient quality. In these cases the results of a screening would be less reliable, if not unusable.
The Takeaway
While the Dimensions 3D and the Pristina are very comparable in image quality and both systems offer 3D tomo-guided biopsy, the sheer lack of availability for the Pristina will preclude most secondary market buyers from owning one in the near future. In the meantime, the Dimensions 3D can be found at a strong value and with a likelihood of more responsive service. Those who have the latitude in their budget to purchase a new system will be well-served by either clinically, but should still mind our note on service responsiveness.
If 3D mammography is on your wishlist, our team would love to talk with you about the Dimensions 3D or any other solution that has caught your eye. To learn more, click the button below tell us how we can help and get pricing information on digital mammo equipment.
1 https://www.itnonline.com/chart/breast-tomosynthesis-systems
2 https://www.accessdata.fda.gov/cdrh_docs/pdf18/K182951.pdf


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Kilim are manufactured by tightly interweaving the warp and weft strands of the weave to produce a flat surface with no pile. Kilim are made by Indian villagers.
They are involved in weaving from generations. And they are well expert in to create any designs. Kilim weaving is not only a source for income, also this is the way of describe their happiness, thought. This is a art of peace loving Indian people.
Color of item can very by 5-10% depend upon display setting of picture and mobile. Size : 5 x 8 Feet, 150 x 240 Cm, 60 x 96 Inches. The item "Ikat Attractive Jute Wool Area Rug Home Decorative 5x8 Feet Rag/Carpet DN-874" is in sale since Wednesday, April 3, 2019. This item is in the category "Home & Garden\Rugs & Carpets\Area Rugs". The seller is "indianrugs" and is located in DAUSA, RAJASTHAN.
This item can be shipped worldwide.


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Consuming cannabis is becoming more mainstream. However, Elon Musk is facing backlash for a recent video of him smoking a blunt on Joe Rogan's podcast.
A doctor's case for medical marijuana
Doctor David Casarett asks: What if mainstream health care operated more like a medical marijuana dispensary? Physician David Casarett was tired of hearing hype and half-truths around medical marijuana, so he put on his skeptic's hat and investigated on his own. He comes back with a fascinating report on what we know and what we
Patient Story – Francie Floyd
Francie Floyd has a service dog with her at all times, since she was an infant, she's been having seizures. Patient Story- Francie Floyd – A high-grade fever when she was less than two months old triggered the first of many. During her early childhood, her seizures were under control with the use of medication.
Dr. Eric Baron Discussing History Of Medical Marijuana Migraine Pain Treatment
Dr. Eric P. Baron explains how medical cannabis works with migraine sufferers and pain. Dr. Baron discusses the fascinating historical and scientific facts surrounding medicinal cannabis use, and how marijuana can have a positive impact on migraine and chronic pain disorders. YouTube Description: There are currently 24 states (Ohio is on the verge of becoming
Marijuana Can Help Ease Fibromyalgia Symptoms
Researchers are now pushing the benefits of medical marijuana as the leading treatment for fibromyalgia relief. Fibromyalgia is the chronic pain and sleep disturbances can now be helped by using medical marijuana. Patients have been struggling to find a cure or at least a pain relief. Most pharmaceutical prescriptions have bad side effects or negative
"Jaxs' policy" – School District in Colorado approves Medical Marijuana Use
A school district board in El Paso County Colorado approved "Jaxs' policy" which allows therapeutic marijuana products at its schools. "Jaxs' policy," was approved as part of a scheduled monthly meeting and is the first of its kind in the state, according to the district. The District 49 Board of Education, in Peyton, unanimously, in
Parkinson's denied as a qualifying condition for medical marijuana in Arizona
Medical Marijuana research for Parkinson's is positive with many legal patients finding relief. However, Parkinson's was denied as a condition for medical marijuana in Arizona.
Marijuana Minute April 6 2016: DEA Weighing Reclassification of Cannabis
The DEA considers reclassifying cannabis The Drug Enforcement Administration is once again considering rescheduling marijuana. It's currently classified as Schedule 1 along with methamphetamines and heroin. However, twice in the recent past the DEA has decided against rescheduling. What are Schedule 1 Drugs? The DEA defines Schedule 1 drugs as: Schedule I drugs, substances, or
Marijuana Mania Episode 1
What does it take to create new marijuana strains? In the premiere episode of Marijuana Mania, Berner takes you on an exclusive trip to the Cookie Fam's farm to talk breeding and then to the 3rd Generation Family farm during harvest season to get an inside look at what it actually takes to create some


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Welcome to Project Life – Week 45! I mentioned yesterday on an Instagram photo about how I looked at this photograph of Lance on the left hand side of this spread and recognized that my "little" man is not so little anymore. There have been a few photographs over the years that take me by surprise in this way, I can still envision a couple of them in my mind. I love that these photographs, this documenting through Project Life, slows me down, makes me stop in my tracks, encourages me to appreciate those moments. I am so very grateful for them.
The full 2 page spread. You can click on the photograph for a clearer image.
Surprise! Surprise! (kidding!) The kit I used this week is the Sunshine Edition. The core products I use in my Project Life albums can be found here. Project Life was created by Becky Higgins. You can find out all about this awesome approach to memory keeping here.


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- Las Positas College
- Assessment Center
- Chemistry Study Guides
Assessment Center
International Baccalaureate (IB)
Please bring an official or unofficial copy of your International Baccalaureate (IB)
exam results to the Assessment Center, room 1642 for a prerequisite clearance petiltion
English 4 or 7
Credit may be earned for International Baccalaureate (IB) exams with scores of 4, 5, 6, or 7 as listed in the college catalog. Students who have earned credit from an IB exam should not take a comparable college course because transfer credit will not be granted for both.
Official or unofficial documents must be printed and contain the following information:
1. Your full name (First and Last name).
2. International Baccalaureate (IB) exam score. Please note that credit will be granted for exam scores only, high school course completion alone is not elligible for college course credit or placement.
Unofficial copies of IB scores may be used for placement purposes only. Official documents are required for degree, certificate and transfer evaluation.
For more information please contact:
Building 1600, Room 1642
Assessment Center - Testing Hours


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My Lucario is the ultimate boss of my team---unless he comes up against a ground type. I know he's also weak to fighting and fire, so how do I fix this so ground types don't destroy him?
His moves are:
Close combat
Aura sphere
Dragon pulse
Dark pulse
I'm thinking replacing dragon pulse would work best but would that leave me open to getting owned by dragons too?
*Dont say to remove aura sphere because I won't. I like aura sphere
You should replace Dragon Pulse with Ice Punch. It still covers dragon types and your also covering grass types along with ground types.


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Corkor Vegan Cork Bifold Slim Wallet for Men
A PETA-approved men's bifold slim wallet handcrafted entirely out of highest quality cork leather by Portuguese artisans.
- 6 card slots.
- 2 hidden pockets behind the card slots.
- 1 compartment for notes and bills.
- Bifold slim design.
- Easy access to all cards and notes.
- Stain and water resistant.
- External size: 10.5 × 8.5 × 1 cm / 4.1 × 3.3 × 0.4 in
- Fits maximum banknote size: 19 × 7 cm (flat)
- Weight: 45 g
How durable is cork?
You're probably a bit skeptic. A wallet made of cork? How does that work? Will it last?
Well, it works beautifully
It won't crack or crumble, in fact, the quality and suppleness of cork fabric products can be compared to that of leather goods. Unlike leather, it's natural, water repellent, scratch and stain resistant. It is also very easy to maintain and clean: water and soap will do the trick! It does age, but will do so nicely. It will last you for years to come, with added character.
Handmade with love in Portugal
Local artisans produce perfectly finished pieces in Corkor's workshop. Traditional craftsmanship combines with small batches, to ensure you get premium quality, without the premium price.
|FSC certified cork leather
|Black / Brown
|10.5 × 8.5 × 1 cm
|5600868466656 / 5600868466885
|Packaging dimension
|13 × 9.5 × 2.5 cm
|Packaging weight
|75 g
|Package contents
|1 Corkor vegan cork men's bifold slim wallet (CK153)


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The ECHO Product range is especially suited for rural and WISP (Wireless Internet Service Provider) type applications where cost is most important. The ECHO-ST will save money, while servicing the need to construct a solid and reliable wireless system replacing the requirement for expensive leased lines.
The SIL-ECHO-ST (CPE - Client Premises Equipment) is a low cost driven product, that can be used in Point to Point (2 x ECHO-ST radios in a link) or Point to Multipoint (using ECHO-BS as multipoint collector + Multiple ECHO-ST) configurations.
The ECHO Range is a budget wireless outdoor bridge that has been optimised for long transmission distances and designed to be extremely compact. Couple that with the FREE SilverView network monitoring software and you have an oustanding feature packed solution.
ECHO-ST Product Page
ECHO-BS Product Page
For installation and setup Training Videos please see our Youtube channel:
ECHO Youtube Playlist
The perfect piece of equipment for site survey spectral analysis
Another successful Install Of SilverNet Equipment
Ideal for mobile data applications
Centrally controlled system, Ideal for town centre Wi-Fi
Ideal for those working on a budget
Finalists 4th year running for communication product of the year
SilverNet look forward to a successful partnership with NBM Distribution
Another success for Videcom and SilverNet partnership
Another successful Installation of SilverNet equipment
Another Successful Install
SilverNet to Exhibit at IFSEC 2015 London ExCel
Wireless Network Solution Provides the link for North Lincolnshire Council
Collaboration delivers cost savings and convenience
Distrimedia Appointed as French distributor
SilverNet nomme un distributeur en France
Meyertech and SilverNet Technical Partnership
SilverNet Introduce Shorthaul Wireless Transmission Link
MESH WiFi access and Security working well together
provides police with secure connection to town centre video surveillance
Pro-Vision achieve Master distributor status with SilverNet
SilverNet Opens International Doors
Security and traffic flow management in a new state-of-the-art CCTV and public safety control room
SilverNet Has Moved to Larger Premises
SilverNet Appoints Keith Newton as Business Development Manager
How do you design a wireless system that can transmit multiple cameras back to a control room with PTZ but also have WIFI for roaming security guards to see alarms and video streams on their handheld devices? Just ask SilverNet!
How to install a new security system with a flexible and reliable wireless infrastructure which covered multiple areas to improve security for the Bourneville site.
Dumbarton were desperate to replace poor 3G security systems with a flexible and reliable wireless infrastructure to cover multiple areas, because the town centre had just had new paving stones placed along the High Street so digging up the street/pavement was not an option.
How do you cover multiple tenement blocks with access control and cctv, with added resilience, cost effectively?
How do you improve a site security system with a flexible and reliable wireless infrastructure to cover a whole brewery site and multiple remote locations?
Part of the regeneration of Ballymun was to upgrade the failing cabled and partial old technology wireless infrastructure for access control and security systems with a flexible and reliable wireless infrastructure
Sensors used in the U.S. that alert police to the exact location of gunshots have been installed for the first time in the UK. West Midlands Police has installed dozens of the ShotSpotter sensors in Birmingham in a bid to combat gun crime in the city
Andy Proudfoot of MOCAM 'we were amazed at the stability of the camera movement and quality of the video streamed over the wireless link, it is truly seamless, well done SilverNet'


text: White Gold Diamond Solitaire Engagement Ring Legacy Solitaire - Heirloom rings start with intentional design. Our Legacy rings are built with a durable but comfortable band, classic lines, and secure settings. Twenty five years creating and servicing jewelry has given us the edge of experience in designing a piece that will last long enough to pass down to a generation. This Legacy ring is cast in 14k white gold and features a 0.46ct diamond round. Compass point alignment of the four prongs keep the ring just a bit different, while also protecting the stone.


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Necessity is the mother of invention. And I do believe the sister of invention is creativity. The two are very closely related anyhow. Well, those ladies, along with four hungry children were all in my kitchen several months back. I needed dinner, fast, and the meal I planned had fallen through. The wheels turned, the digging in the refrigerator began, and a recipe was born. To this day, my kids all squeal in delight when I tell them Cottage Mac and Cheese is what's for dinner.
Cottage Mac and Cheese
by Blessed Roots Farm
This recipe feeds at least four hungry kids and one mom. Adjust amounts to make it feed your family.
4 cups dried whole wheat macaroni noodles (or equivalent amount of any pasta)
1 32 oz. container of whole milk cottage cheese
1/4 c. organic butter
1/4 – 1/2 c. whole milk
3 cups fresh spinach, chopped
4 large carrots, peeled and grated
(Adding any leftover meat is completely optional, and quite yummy!)
Freshly ground black pepper and freshly grated parmesan or romano cheese are delicious garnishes.
Fill a large stock pot about 2/3 full with water, and bring to a boil. Add the macaroni noodles (and optional leftover meat if it is frozen) and set the timer so you don't overcook them :). While the macaroni is cooking, shred the carrots. Toss the shredded carrots in the macaroni water. While the noodles and carrots cook, chop the spinach. Add the chopped spinach (and any leftover meat if it is refrigerated) and stir. By this time, your noodle timer should be just about to beep. When your noodles are cooked, drain the noodles and veggies in a colander with fine holes, so you don't lose half of your veggies. Pour the noodles and veggies back into your stock pot and place over low heat. Add butter and stir until melted. Then, add the cottage cheese and the milk. Add enough milk to get the pasta to the consistency you prefer. Top with fresh ground pepper and freshly grated parmesan or romano cheese.
Sounds so weird, I know. But they LOVE IT. And it doesn't come out of a box!
Join me at This Chick Cooks, Real Food Wednesday, and Full Plate Thursday to see more yummy recipes.


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pacific coast highway 1904
Ocean Street, now known as Pacific Coast Highway, and Main Street in Downtown Huntington Beach in 1904
Credit: Chris Epting
Traveling to Huntington Beach? Be prepared to travel responsibly and bring a face mask. While here, please be mindful of your surroundings and thoughtful of others: social distancing, wearing face coverings, and washing your hands. Huntington Beach is doing everything it can to be clean and safe so you can stay healthy and enjoy beautiful Surf City USA.


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WoW powerleveling boost is something that needs to be done for every character you play, but most people hate the boring grind that comes with WoW leveling boost. Thats why we offer to you cheap and efficient service for increasing your character levels by our boosters and saving you some time you would otherwise spend on powerleveling boost WoW.
Many people ask, is it safe to purchase WoW leveling boost?
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Our company uses the payment gateway for gamers – G2A Pay, which has established itself as one of the most convenient payment gateways with the ability to use more than 200+ methods for payment. Thanks to G2A Pay, you will receive the highest level of online payment security, comprehensive encryption solutions and the protection of your personal data. To ensure maximum security of your payments and transactions, confidential information is always encrypted and your credit card information is never stored on our servers.
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Our boosters use VPN every time they play on your account. This is very important and thanks to the use of VPN, you do not have to worry about the security of your account during WoW leveling boost. We choose a VPN server based on your country and city, which allows us to ensure maximum security of your account data while performing WoW powerleveling boost.
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So in conclusion if you want to get WoW powerleveling you can safely buy leveling boost from us – we guarantee that we will use all our security measures and your account will stay safe!


text: Oxford House Medallion in Jackson, Mississippi, United States Free Addiction Resource Category: Free Addiction Treatment Resources and Oxford House Addiction Treatment [ Map Address Oxford House Total Occupancy Phone Reviews No Records FoundSorry, no records were found. Please adjust your search criteria and try again. Google Map Not LoadedSorry, unable to load Google Maps API. Driving Walking Bicycling Public Transport Miles Kilometers 505 S 28th Ave Jackson Mississippi 39402 United StatesHousing for MenTotal Occupancy 7(769) 572-4663 Leave a Review Cancel replyYour email address will not be published. Required fields are marked *Review text Select a rating Name * Email * Website Save my name, email, and website in this browser for the next time I comment. Previous Next


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Well you don't get 3g out of those engines though.. in my current tests i calculated with 2 x 6GW Reactors. At 1500s ISP they produce a thrust of ~1631 kN, so a bit over 0.5g for a 300ton ship. They burn, or rather heat 110kg of propellant per second. Thats almost 8 tons of propellant for just accelerating to 350 m/s and you need some additional energy for a climb to 8-10km altitude to avoid too high dynamic pressures. Also you loose some energy to drag. So i guess we're more looking at 10-15 tons propellant used to get to 350 m/s.djolds1 wrote:Its just those first few hundred meters per second dv before the ram-air effect is possible. Twelve seconds at 3gees to boost upto 350m/s. Early designs such as the Saenger Silverbird and Soviet Keldysh used sleds, but the efficiency of QED/ARC makes a sled superfluous. Twelve seconds of low altitude QED/ARC burn per flight doesn't seem very significant. And using one type of engine (QED/ARC-RBCC) eliminates additional complexity.
To compare that to ducted fans, i get about the same thrust with just 300-400MW shaft power with a total fan area of 16 m^2. Also they would probably not weigh so much more than the propellant saved by using them.
Also those fans would be more like axial compressors with a pressure ratio of about 1:4. They would sit in the airflow path to the QED engine and would benefit the pressure ratio of the engine up to about M2.8.
Heres a little pic i made for roughly illustrating the concept:
Its very basic but shows the general idea.
Yep i thought something like that too.. just use the standard QED engine and feed it with air from a seperate inlet channel.. (see pic above)Oh, IMO forget "Rasterscan the underbody." Instead, scan the REB inside a "standard" (sc)ramjet engine tube. The forebody/diffuser locks forward for "pure" ARC mode, but retracts to allow airflow. Place the REB generator in the engine tube like the rocket engine in a "standard" RBCC.


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The average American holds 10 jobs before reaching the age of 40. If this tells us one thing, it's that we don't know what we want from our job. But what if we could try our jobs before committing to the position? With Betagig you can do just that. The online platform lets you shadow your job before accepting the offer to insure job compatibility. Betagig gives you the ability to make a more informed decision on how you use a third of your day. If job satisfaction is a value of yours, we suggest using Betagig to insure the best results.
LA TechWatch spoke with cofounder and CEO Nicki Klein about Betagig and how they secured their second round of funding.
Who were your investors and how much did you raise? We just secured $1.6M in Seed funding from Altpoint Ventures.
Tell us about your product or service. Betagig is an online platform that allows users to job shadow future careers, creating the world's first "try before you buy" workplace experience. A common trend among students includes reluctantly choosing a major and career path, resulting in their apathy and disillusionment in the workforce. As a result of this flawed career-choosing pattern, millennials have adopted a proclivity to "job-hop" around the market, causing increased turnover within companies, which can easily be prevented with job shadowing. The program is accessible to adults regardless of their prior work experience, but is especially aimed toward college students, recent graduates, and those looking to change careers.
What inspired you to start the company?
Melissa Hargis and I were both career changers multiple times. I changed my college major 8 times and my career 4 times. So the idea of finding something we loved to do was obviously important to us and we found out that this was an issue so many people today struggle with. One day we were walking through Golden Gate Park in San Francisco, and Melissa said "wouldn't it be great if you could change lives with someone for a day" and I said, "No it would be cool if you could swap careers with someone for a day" and the idea flourished from that conversation.
How is it different?
Betagig is the world's first job shadowing platform. Job shadowing has always existed and it happens everyday, but it has always been a chore to seek out opportunities. This platform streamlines the process and it really is the first of its kind.
What market you are targeting and how big is it?
We are targeting the recruitment industry, which is a $124B market here in the US and $440 billion worldwide.
What's your business model?
Our business model is two fold. The first is the free service. For companies, the service is free to use and participate in and they only pay if they hire a candidate. The second is a SaaS model.
What jobs are people seeking to shadow the most?
We have seen the most requests in the entertainment, health and tech industries.
What was the funding process like?
For me, being a female, a first time founder and a founder with a moonshot idea, it was long and difficult .
What are the biggest challenges that you faced while raising capital?
Again, being a female and a first time founder. It's essentially a numbers game.
What factors about your business led your investors to write the check?
One of the main reasons was how the state of the current job market is today and how technology can have a great impact on that. There are not a lot of startups in the HR space right now. In addition to that, it was a big ticket in the presidential election so their is clear need for change and improvement in this space. Nicki also took feedback from every investor on her pitch and vastly improved it over time with each new investor she spoke with.
What are the milestones you plan to achieve in the next six months?
We will be piloting with a large company in Los Angeles soon, as well as launching with a local community college that we are in contract with. Furthermore, we are completing the hiring of our team.
What advice can you offer companies in Los Angeles that do not have a fresh injection of capital in the bank?
Fake it til you make it and keep the passion alive.
Where do you see the company going now over the near term?
We will be growing and scaling very rapidly.
What is your favorite restaurant in LA?
The Misfit in Santa Monica


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What's the Right Moving Truck Size for Your Move?
At a glance
We get it: it's easy to overpay for a too-large truck for fear of running out of room.
If you're on the fence between two sizes, play it safe and go for the larger size. It's better to have unused space you can cushion than to have to leave your prized telescope behind—after all, parting with your things will cost you time, money, and (ahem) emotional stress.
If the moving truck rental company in your area doesn't have your preferred size, you can always mix and match sizes (like pair the largest and smallest sizes together if you're worried about room) or hitch a trailer to the back of your rental.
Lastly, don't wing it! Do some research, talk to a rental rep, and learn from others who have been there and done that—Google reviews and ConsumerAffairs.com are great places to start.
If you decide to size up to a larger truck, make sure you have moving blankets to wedge between items to prevent shifting. If you go down a size, consider adding a trailer to your rental.
The whole picture
If you've moved only a handful of times—or if this is your first rodeo—you may not know which truck size is right for your move. It's even more complicated if the company you're renting from advertises only small, medium, and large sizes (we're looking at you, Budget) that don't match your exact load size.
So how do you pick the right truck size for your move? Start by figuring out the total number of cubic feet you'll fill and the number of rooms you're packing up.
An online moving calculator can tell you how much cubic space (a.k.a. volume) you'll need based on how many beds, boxes, or, I don't know, unicycles you're moving. Plus, most rental truck companies advertise how many rooms each truck size can hold.
It's also helpful to ask yourself the right questions:
- How many rooms are you moving? How many of these are bedrooms with mattresses?
- How many boxes are you moving? (Check out Home Depot's box calculator to get an accurate estimate.)
- How big is your furniture? Do you have any oversized, bulky, or fragile items that may need special treatment?
- How far are you moving?
- Which truck sizes fall in line with your budget?
Book a truck size that has 10–15% more cubic space than your estimated load to ensure you have enough room. (Don't worry, every good rental truck company will spell out truck volume for you).
Once you've nailed down how much stuff you're moving, there are two truck types we'd recommend:
- Pickup trucks
- Box trucks
Of course, there are other options out there, like cargo vans and moving trailers you can hitch to your car or truck. But we'll focus on pickups and box trucks since those are the best for most DIY moves.
Box trucks
Box trucks are the most common moving trucks—you know, the ones you often see parked in front of your apartment building. They're more expensive than some other truck types, but they're still far cheaper than doing a container-style move or going full service.
If you're moving more than what fits in a 1-bedroom apartment, you'll probably want a box truck. In our opinion, a box truck isn't worth the price for smaller loads—chances are you'll be able to find a smaller cargo van better suited for your move.
Rental truck companies usually offer at least two box truck sizes (based on the length of the box itself). Here's the breakdown of which truck sizes are generally considered small, medium, and large.
Small box truck sizes
Do you struggle to find space for your massive book collection in your studio apartment? We feel you. Your Dickens collection deserves more breathing space.
If you're moving a 1-bedroom apartment or smaller, the 10- and 12-foot sizes are where you'll want to start. Queen, full, and twin mattresses fit in these trucks, but most king-size mattresses, sadly, are a no-go.
Small box truck sizes are where most customers overestimate their needs. Most of the time, if you think you can make it work with one size, you'll be able to.
Basically, if you pack up your truck the right way, you'll most likely be able to make do with the size your instincts told you to go with.
To do so, start with the big stuff. Load your couches, beds, and other large pieces of furniture into the truck first—then pile in smaller pieces of furniture along with your boxes.
Small box truck size comparison
Medium box truck sizes
Mid-size box trucks (and larger) are best for long-distance moves and—you guessed it—larger loads. These trucks are also more durable and better suited to long road trips, and there's a whole lot more space for your things.
Medium box trucks are best for two-bedroom apartments, condos, and one- to two-bedroom small homes. A key difference between small and medium box trucks is that king mattresses fit inside the latter.
If you rent a medium-size truck from U-Haul (15 ft. or 17 ft.), it'll come with a mom's attic, which provides extra storage space above the driving cab for mattresses or boxes.
Unless you're towing your vehicle behind your rental truck or toting a family of five, you'll probably have room in your backseat or trunk to haul things (should you need more space).
Medium moving truck size comparison
Large box truck sizes
Do you own a California king mattress? Look no further than 20-plus-foot moving trucks. These large trucks can generally hold items from two- to three-bedroom apartments all the way up to three- to five-bedroom houses. Plus, these trucks can handle the weight of washers and dryers.
Each large U-Haul truck also comes with a mom's attic, so don't worry: you don't need to throw away your comic book collection. You've got extra space to stash them.
Large moving truck size comparison
Enterprise is another rental truck company worth looking into. The company has 14 vehicle sizes—but it does only local moves. Read our full review for more details on what Enterprise has to offer.
Pickup trucks
If you're moving a single room's worth of stuff—or if you have a few bulky pieces of furniture—a pickup truck is a cheap and easy alternative to a full-service move or a larger truck rental.
It makes sense to use a pickup truck if you're transporting a couple chairs and a table or if you're moving into a dorm room or small studio apartment.
Plus, if you're working on home improvement projects, pickups are great for transporting tools, yard debris, and anything else you don't want dirtying up your backseat.
Pickup truck size comparison
|Truck bed dimensions
|7'9" x 5'2" x 1'9"
|Twin or full mattress, 3–4 furniture items
|Weight limit
|1,980 lbs.
|Miles per gallon
|Companies that offer these sizes*
*Budget offers pickup trucks through its sister car rental company.
The better you organize your things into boxes, the fewer moving boxes you'll need. Luckily, we have a whole review on how to pack efficiently, as well as a how-to on using styrofoam, packing peanuts, newspaper, and bubble wrap.
Still can't find your size?
If you're between truck sizes, consider going with the smaller option and adding a trailer to your rental for more space—especially if you're moving awkward or bulky items. For example, lawnmowers aren't allowed inside a rental truck anyway.
(Check out our Moving Companies That Offer Trailer Rentals piece for more deets.)
If you still need a bigger rental truck, consider doing a full-service move or a container-style move. Rental trucks are especially good for smaller, local moves (and let's be real, for cheaper rates), but if you're moving a mansion or from one coast to another, you'll likely need something bigger.
Rental Truck Companies We Like
We dug deep into rental truck options in our Best Moving Truck Rental Companies list, but if you're looking for something specific, here are a few of our favorites:
- U-Haul Review: Best nationwide availability
- Penske Truck Rental Review: Best for guaranteed reservations
- Budget Truck Rental Review: Best budget-friendly prices
- Enterprise Truck Rental Review: Best variety of rental trucks