a, Northern blot analysis oflamp-2 expression. Total RNA was hybridized using a lamp-2cDNA2 probe and a murine glyceraldehyde-3-phosphate dehydrogenase probe.b, Western blot analysis of LAMP-2 expression. In control liver and kidney extracts the glycosylated LAMP-2 molecules were detected. In LAMP-2-/- tissues no LAMP-2 product was found.c, Increased mortality in LAMP-2deficient outbred (C57B6/Jx129SV) mice. Forty-two control (closed circles) andLAMP-2-deficient mice (open circles) were monitored over 300 days.d, Loss of weight in early dying (n = 7 males) and surviving LAMP-2-deficient mice (n = 11 males) in comparison with control mice ( n= 12 males) monitored for 60 days after birth.a, Ultrastructure of autophagic vacuoles (arrowheads) in pancreatic acinar gland cells from a 6.5-month-oldLAMP-2-deficient mouse.b, Autophagic vacuoles containing glycogen (g), cytosol and cellular constituents in a hepatocyte of an 8-day-oldLAMP-2-/- mouse. m, mitochrondria; bc, bile canaliculus.c, Accumulation of numerous early autophagic vacuoles in cultured LAMP-2-/- hepatocytes. Arrow, large late autophagic vacuole containing partially degraded material. Insert, magnification of the region indicated by an asterisk with typical earlyautophagic vacuoles containing undigested cellular constituents. Scale bars: a, 3.5 µm; b, 0.2 µm; c, 1.3 µm; insert, 0.9 µm.
a, Vacuole in a fibre of the soleus muscle. b, Ultrastructure of a vacuole in a cardiomyocyte. Scale bars: a, 1 µm; b, 1.5 µm.
c, Typical original twitch recordings of developed force of heart muscles from control (thin line) and LAMP-2-deficient (thick line) mice. d, Average values of developed force. Preparations of LAMP-2 deficient mice had a significantly lower baseline developed force at a stimulation frequency of 4 Hz and 10 Hz; P 0.01.
a, Volume density of early (Avi) and late (Avd) autophagic vacuoles from four independent cultures. Examples of Avi and Avd are shown in b and c, respectively. Bars represent 0.4 µm.
d, Labelling density of cathepsin-D as estimated in two independent experiments. e, Labelling density of LAMP-1.
f, Degradation of long-lived proteins followed over 24 h in amino acid and fetal-calf-serum-containing medium. g, Effect of amino-acid (AA) deprivation and 3-methyladenine (MA) on protein degradation during 4 h of chase. The error bars in a, d, e and g represent s.e.m.