fertilization is the process in which 2 different gamete cells , a sperm and an oocyte , unite to produce a zygote . for fertilization to be successful for example , after sperm has differentiated completely , various extracellular factors such as epididymosomes and albumin alter the structure and function of the plasma membrane of the sperm . in addition , in terminally differentiated gamete cells , various sterols , sphingolipids , glycolipids , and glycosylphosphatidylinositol- ( gpi- ) anchored proteins are localized on cell membrane microdomains that are called lipid rafts . lipid raft components are often examined by using detergent - resistant membrane domains ( drms ) , which enrich these components so that their distributions and functions can be visualized on the cell surface by using putative raft markers [ 1 , 2 ] . since lipid rafts in gametes contain signaling proteins that regulate intracellular functions and cell signaling , these domains are important for sperm maturation , fertilization , and early embryogenesis [ 3 , 4 ] . in this paper sperm are highly differentiated haploid cells with a head and a tail ( flagellum ) . the head consists of a nucleus , an acrosome , and a small amount of cytoplasm , while the tail consists of a motility apparatus , mitochondria , an axoneme , and cytoskeletal structures . although these structures are necessary for sperm to swim and fertilize oocytes , these structures are not functional after spermatogenesis until the plasma membrane is modified during epididymal transit ( figure 1(a ) ) . in mammals , the sperm mature in the epididymis ; however , in other animals , sperms mature in the spermiduct . previous studies have demonstrated that the modifications of the sperm plasma membrane that occur during epididymal transit include changes in its lipid and protein composition , modifications of surface proteins , and increased total negative charge of the extracellular surface [ 8 , 9 ] . electron microscopy studies showed that the epididymal lumen contained membranous vesicles that which called epididymosomes [ 1012 ] . these vesicles , which are particularly rich in sphingomyelin ( sm ) and arachidonic acids , are secreted via apocrine secretion . in addition , two - dimensional gel electrophoresis and liquid chromatography - quadrupole time - of - flight ( lc - qtof ) analyses show that epididymosomes contain endoplasmin ( heat shock protein 90 1 ; hsp901 ) , 70  kda heat shock protein 5 , chaperones , and other exosomes [ 1316 ] . in addition , integral membrane proteins , such as gpi - anchored proteins , also are associated with epididymosomes . several gpi - anchored proteins in epididymosomes have been found in the mature sperm of various animals . for example , he5 ( cd52 ) is found in human sperm , spam 1 and hyaluronidase are present in mouse sperm [ 1821 ] , and p26h   and p25b   are found in hamsters and bulls , respectively . in addition , epididymosomes localize to the sperm membrane during epididymal transit and contribute to the formation of various membrane structures such as lipid rafts in sperm . furthermore , in sperm membrane lipids , the percentage composition of both sm and various polyunsaturated fatty acids , which are mainly arachidonic , docosapentaenoic , and docosahexaenoic acids , increases throughout the epididymal tract . six genes , known as he1he6 , are expressed specifically in the human epididymis [ 2628 ] . mutations in he1 cause niemann - pick type c2 ( npc2 ) disease , a fatal neurovisceral disorder that is characterized by the accumulation of cholesterol in lysosomes . he1 is a small , soluble glycoprotein with 132 amino acids that binds to cholesterol , but not to cholesterol derivatives that have hydrophilic substitutions on their isooctyl side chains [ 2932 ] . xu et al .   determined the x - ray crystallographic structure of bovine npc2 protein complexed with cholesterol sulfate . together , these studies showed that he1 binds to cholesterol in vitro and may regulate the cholesterol content in sperm throughout the epididymal tract . however , other studies have demonstrated that the ratio of total phospholipid to total cholesterol does not change during epididymal transit [ 12 , 34 ] . previously , we used filipin as a cytochemical probe for membrane cholesterol in the sperm plasma membrane during epididymal maturation ( figure 2 ) . our results show that the filipin signal decreased at the post - acrosomal region during epididymal transit previous studies have shown that the rate of pregnancy in humans decreases as the amount of time between a vasectomy and its reversal increases [ 3638 ] . in addition , sperm from men who have undergone vasectomy reversal have higher levels of he1 , cholesterol , and ganglioside gm1 compared with sperm from fertile men who have not had vasectomy . these findings suggest that he1 regulates the amount or localization of sperm lipid rafts in the epididymis , which produces mature sperm . several studies have shown that modification of sm , polyunsaturated fatty acids , cholesterol , and gm1 changes the fluidity of sperm membranes and the composition of lipid rafts . for example , lipid diffusion in the plasma membrane of mouse sperm increased significantly during their transition from the caput epididymis to the cauda epididymis . reported that pheochromocytoma pc12 cells that overexpress gm1 did not exhibit any neurite formation , even after stimulation with nerve growth factor ( ngf ) . furthermore , increased expression of gm1 reduces membrane fluidity , disorders the lipid raft , and changes the intracellular localization of ngf receptors and related signaling molecules . further studies are needed to elucidate the mechanisms and molecules that promote sperm maturation in the epididymis . after maturing in the epididymis , sperm are able to swim and fertilize an oocyte ( figure 1(b ) ) . in invertebrates that reproduce by external fertilization , in contrast , in mammals that reproduce by internal fertilization , sperm are unable to fertilize oocytes immediately after they can swim . instead , sperm acquire this ability after moving to the uterus and remaining for an appropriate period of time [ 42 , 43 ] . the biochemical process that confers this ability to sperm is called  capacitation .  during sperm capacitation , protein tyrosine phosphorylation occurs along with hyperactivated flagellar beating ; further , the acrosome reaction is induced , and the sperm penetrates the zona pellucida ( zp ) and finally binds and fuses with the oocyte . in addition , the organization of membrane proteins and lipids changes significantly [ 4448 ] . in vitro , capacitation requires a high concentration of albumin in culture medium to decrease the sperm cholesterol / phospholipid ratio [ 49 , 50 ] . fluorescent and electron microscopic studies have shown that a combination of bicarbonate and albumin promotes membrane distribution and cholesterol efflux [ 35 , 51 , 52 ] . albumin also decreases the amount of sialic acid , ganglioside , and triglyceride in the sperm plasma membrane . in addition , methyl--cyclodextrin ( mbcd ) promotes sperm capacitation in vitro , by decreasing the amount of cholesterol in the plasma membrane and disrupting lipid rafts ; however , the critical concentration of mbcd should be considered [ 5356 ] . the strong mbcd treatment also reveals some proteins that are involved in capacitation - dependent processes and zp binding . the physiological relevance of raft reordering in the sperm surface is to create protein complexes involved in zp binding [ 58 , 59 ] . cholera toxin subunit b ( ctb ) , which binds to ganglioside gm1 with a high affinity , has been widely used as a reporter of the distribution of lipid rafts and to study the ctb - binding pattern of sperm in both in vitro and in vivo conditions . we , along with several other research groups , also have developed methods to investigate the distribution of gm1 in fixed sperm . these studies have revealed that gm1 expression is significantly altered during sperm capacitation and the acrosome reaction [ 6062 ] . in contrast , the ctb - binding pattern of sperm is variable and depends on their fixation conditions . in living sperm , selvaraj et al .   demonstrated that the cyclodextrin treatment does not change the distribution of gm1 in mouse or bovine sperm . however , the use of specific fixation conditions induced stimulus - specific patterns of gm1 distribution . specifically , in mouse sperm , gm1 was broadly localized from the postacrosomal region to throughout the sperm head . .   showed that the distribution of gm1 in boar sperm changes sequentially , from the tail to the head , during mbcd - mediated capacitation . one of our previous studies also demonstrated the ctb- binding pattern in murine sperm in physiological conditions . briefly , ejaculated sperm were collected from the female mice when the sperm were first detected in their oviducts ( approximately 3 hours after copulation ) . their reproductive tracts were divided into 4 parts , namely , the oviduct , uterine region near the oviduct , uterine region near the cervix , and vagina . while the sperm migrated from the uterus to the oviduct , ctb fluorescence was lost from the postacrosomal region . in addition , gm1 interacted with seminal vesicle secretion 2 ( svs2 ) , which is secreted from seminal vesicles and inhibits sperm capacitation . since svs2 is a highly basic protein , increasing evidence suggests that sperm capacitation is regulated by the distribution of gm1 or its charge on the sperm plasma membrane . however , staining pattern by ctb is not consistent with that by other probes , such as lysenin and antibody against thy-1.2 [ 65 , 66 ] . lysenin and the antibody have high affinities for sphingomyelin and gpi - anchored protein enriched in the drm fractions , respectively . these invaginations are formed from lipid rafts by polymerization of caveolins , which are palmitoylated integral membrane proteins that bind to cholesterol with high affinity [ 67 , 68 ] . previous studies have demonstrated that caveolin-1 and -2 are enriched in the triton x-100-insoluble membrane fraction of mature sperm and localize to the acrosomal membrane [ 69 , 70 ] . since these caveolins disappear after the completion of the acrosome reaction , they are thought to regulate the acrosome reaction . however , caveolin-1-and -2-null mice are fertile , and the distribution of gm1 in caveolin-1-deficient mouse sperm is comparable to that in wild - type mouse sperm [ 63 , 71 , 72 ] . these findings suggest that caveolae are not required for the presence of lipid rafts in the acrosomal membrane and fertilization of mouse sperm . however , several other studies indicate that lipid rafts in the apical ridge head area of sperm have affinity for the zp [ 57 , 59 ] . since the zp is involved in the acrosome reaction , the function of lipid rafts in this reaction is still controversial . in general , sex hormones stimulate the continuation of the first meiotic division of oocytes . however , in some animals , this occurs after the oocyte is released from an inhibitory environment . in both vertebrates and invertebrates , the first meiotic division of oocytes is asymmetric , which results in the formation of a relatively large oocyte and small polar bodies . during meiotic arrest , oocytes are fertilized by sperm , and then the second meiotic division is completed . the activation of oocytes upon fertilization is a ca - dependent process in all animals [ 73 , 74 ] . furthermore , intracellular ca ( [ ca]i ) is a key regulator of many cellular functions . in oocytes , increased [ ca]i stimulates the continuation of the second meiotic division and formation of the second polar body , followed by formation of male and female pronuclei [ 47 , 75 ] . the signaling pathway that regulates this increase in [ ca]i is highly conserved among species . in this pathway , phospholipase c-(plc- ) dependent production of inositol 1,4,5-triphosphate ( ip3 ) triggers the increase in [ ca]i , which propagates from the endoplasmic reticulum . in contrast , the binding of sperm to the oocyte plasma membrane is species specific . currently , there are 2 hypotheses about how a sperm binds and activates an oocyte , namely , a transmembrane receptor mechanism that involves g proteins and a soluble sperm factor mechanism . xenopus oocytes have a g protein - coupled sperm receptor that activates plc on the plasma membrane ( figure 1(c ) ) . in addition , lipid - raft - associated proteins , such as uroplakin iii ( xupiii ) and its tetraspanin - binding partner uroplakin ib ( upib ) , are involved in the sperm - oocyte membrane interaction and subsequent oocyte activation . specifically , xupiii is cleaved by sperm protease and promotes oocyte activation via src tyrosine kinase and plc signaling [ 76 , 78 , 79 ] . xupiii and upib form a complex on the oocyte plasma membrane and colocalize with gm1 . in addition , cd9 , a tetraspanin that is involved in the sperm - oocyte fusion in mice , is found in the drm fraction of xenopus oocytes ; however , cd9 does not interact with xupiii or upib . these findings suggest that the mechanism of sperm - oocyte fusion in xenopus is different from that in mouse , although lipid rafts are involved in this process in both species . furthermore , 2 plasma membrane proteins , cd9 and cd81 , are important molecules in murine sperm - oocyte fusion [ 81 , 82 ] . cd9 and cd81 belong to the tetraspanin superfamily and form a complex with integrin 31 ; moreover , cd63 that forms a complex with cd9 and cd81 localize- along with phosphatidylinositol 4-kinase within lipid raft - like microdomains in a431 and ht1080 cell lines . female cd9 knockout mice are infertile ; although they produce oocytes that mature normally , these oocytes can not fuse with sperm . cd81 , which has a similar structure and function as cd9 , also is involved in gamete fusion . however , the effects of deleting cd81 are less dramatic than those of deleting cd9 [ 82 , 84 ] . the expression level of cd9 is not affected by a deficiency of cd81 in murine oocytes and vice versa . both cd9 and cd81 are localized on the surface of murine oocytes ( figure 3(a ) ) . however , their distributions were completely different . as previously described , cd9 was expressed on the oocyte microvilli . whereas cd81 was distributed in microdomain - like structures between microvilli that expressed cd9 . however , both cd9 and cd81 were concentrated at the sperm attachment site ( figure 3(b ) ) . cyclodextrin , a lipid - raft disruptor , inhibited sperm - oocyte fusion and decreased the percentage of two - cell formation in a dose - dependent manner ( figure 3(c ) ) . similarly , filipin had the same effect on the oocyte plasma membrane . as a result , it is likely that cd9 and cd81 are important for coordinating the sperm - oocyte fusion process in mice . after fertilization , the zygote divides into a blastocyst that can be implanted into the uterus in mammals . at the 8-cell stage , a murine embryo undergoes compaction to form polarized morulae ( figure 1(d ) ) . filia - mater complexes localize asymmetrically in the apical cytocortex of 2-cell embryos due to their absence near cell - cell contact . although this asymmetry is reversible when the blastomeres of 2- and 4-cell embryos are separated , filia - mater complexes are detected at the apical subcortex of these findings indicate that the plasticity of the localization of filia - mater complexes may reflect the cell fate determination of preimplantation mouse embryos . the outer cells of the morulae become the mural trophectoderm while the inner cells form the inner cell mass ( icm ) of the blastocyst . originally , embryonic stem ( es ) cells were isolated from the icm of the blastocyst and were shown to be pluripotent and self - renewing . the studies about filia - mater complexes and es cells suggest that cell - cell adhesion that results from compaction may be responsible for pluripotency . many cell - surface antigens that are markers of pluripotency have been identified in the icm , es cells , and embryonic carcinoma cells . the most common marker of murine es cells is stage - specific embryonic antigen ( ssea)-1 . specifically , ssea-1 is highly expressed in the morula stage , suppressed after compaction , and then expressed only in the icm of the blastocyst . the lewis epitope of ssea-1 ( gal1  4(fuc1  3)-glcnac1  3gal ) also is found on glycosphingolipids and glycoproteins [ 8991 ] . mouse blastomeres and embryonic carcinoma cells identify each other and aggregate by recognizing this epitope [ 92 , 93 ] . similarly , ssea-3 and -4 are common markers of human pluripotent stem cells and are highly expressed before the morula stage but decline afterwards . the ssea-4 epitope is found in the lipid rafts of achn , a human renal cancer cell line . in viable murine embryos , an anti - ssea-4 antibody detects ssea-4 over the entire membrane surface , with some accumulation at the interface between blastomeres . a similar pattern of localization also was also observed with ctb staining . because other sseas are homologous to glycosphingolipids and share similar chemical properties , they are also likely to be enriched in lipid - rafts . ssea-1 is colocalized with adhesion - related proteins , such as cd9 , icam-1 , and pecam-1 , in the contact regions of murine embryos and es cells . these proteins also are lipid raft - associated proteins [ 102104 ] that are distributed in a way similar to ssea-4 . collectively , these findings suggest that ssea-1 , -3 , -4 are localized in lipid rafts , are involved in cell - cell adhesion , and may contribute to the pluripotency of mouse es cells . in addition to cell adhesion , lipid rafts play an important role in cytokinesis , which is a complex process that involves dynamic cortical rearrangement . surprisingly , clathrin mutations that affect endocytosis cause defects in cytokinesis in many organisms [ 106 , 107 ] . .   showed that clathrin and caveolae are localized at the cleavage furrow in zebrafish blastomeres . in embryos ganglioside gm1 , cholesterol , and tyrosine - phosphorylated proteins also have been found in the cleavage furrow and plane of sea urchin embryos . in these embryos , drms contain src and plc , which are tyrosine phosphorylated at the site of cytokinesis . furthermore , activation of these enzymes is required for furrow progression . these studies suggest that caveolae and lipid rafts contribute to cytokinesis in early developmental embryos .
cell membranes are composed of many different lipids and protein receptors , which are important for regulating intracellular functions and cell signaling . to orchestrate these activities , 
 the cell membrane is compartmentalized into microdomains that are stably or transiently formed . 
 these compartments are called  lipid rafts  . in gamete cells that lack gene transcription , distribution of lipids and proteins on these lipid rafts is focused during changes in their structure and functions such as starting flagella movement and membrane fusion . in this paper , we describe the role of lipid rafts in gamete maturation , fertilization , and early embryogenesis .