Inhibition of translation and modification of translation factors during apoptosis induced by the DNA-damaging agent MMS in sea urchin embryos

Translational control was investigated in sea urchin eggs and embryos in response to the DNA-damaging agent methyl methanesulfonate (MMS). We have shown in this report that exposure of sea urchin embryos to MMS induces drastic effects on protein synthesis activity, and on translation factors level, integrity and post-translational modifications. In response to the treatment of embryos by the DNA-damaging agent MMS, protein synthesis is inhibited independently of the translation inhibitor 4E-BP and in correlation with phosphorylation of the translation factor eIF2alpha subunit. Furthermore, a low molecular weight form of translation initiation factor eIF4G is detected correlatively with MMS-induced apoptosis. We propose that modifications of translation factors play an important role in protein synthesis modulation that occurs during DNA-damage induced apoptosis.     

Expression of type IV collagen-degrading activity during early embryonal development in the sea urchin and the arresting effects of collagen synthesis inhibitors on embryogenesis

Type IV collagen-degrading activity was expressed in homogenates of Lytechinus pictus embryos during embryogenesis. Activity was concentrated 1,600-fold by ammonium sulfate fractionation, ion exchange, and gel chromatography and could not be activated further upon trypsin or organomercurial treatment. This enzyme activity could also degrade gelatin but had no affinity for type I, III, and V collagens. Activity was inhibited by addition of excess type IV collagen or gelatin, but was unaffected by addition of excess amounts of non-collagenous proteins of the extracellular matrix. Chelators such as 1,10-phenanthroline or Na₂EDTA reduced activity to control levels. Inhibitors of plasmin and of serine and thiol proteases were without effect. Type IV collagen-degrading activity first became apparent at the stage of early mesenchyme blastula. It then increased by a small increment and remained stable up to the stage of late mesenchyme blastula, coinciding with first detection of collagen synthesis and the appearance of the archenteron. Thereafter, a sharp increase in activity was observed, concurrently with remodelling of the archenteron. Maximum activity was attained at prism stage and was retained throughout to pluteus-larva stage. The specific inhibitors of collagen biosynthesis 8,9-dihydroxy-7-methyl-benzo[b]quinolizinium bromide and tricyclodecane-9-yl xanthate arrested sea urchin embryo development at early blastula, prevented the invagination of the archenteron, and reverted the expression of type IV collagen-degrading activity to non-detectable levels. Removal of the inhibitors allowed embryos to gastrulate and express type IV collagen-degrading activity.     

Microtubule assembly is required for the formation of the pronuclei,nuclear lamin acquisition,and DNA synthesis during mouse,but not sea urchin,fertillization

Microtubule assembly is required for the formation of the male and female pronuclei during mouse, but not sea urchin, fertilization. In mouse oocytes, 50 μM colcemid prevents the decondensation of the maternal meiotic chromosomes and of the incorporated sperm nucleus during in vitro fertilization. Nuclear lamins do not associate with either of the parental chromatin sets although peripherin, the PI nuclear peripheral antigen, appears on both. DN A synthesis docs not occur in these fertilized, colcemid-arrested oocytes. This effect is limited to the first hours after ovulation, since colcemid added 4–6 hours later no longer prevents pronuclear development, lamin acquisition, or DNA synthesis. Neither microtubule stabilization with 10 μM taxol nor microfilament inhibition with 10 μM cytochalasin D or 2.2 μg/ml lalrunculin A prevent these pronuclear events; these drugs will inhibit the apposition of the pronuclei at the egg center. In sea urchin eggs, colcemid or griseofulvin treatment doe? not result in the same effect and the male pronucleus forms with the attendant accumulation of the nuclear lamins. The differences in the requirement for microtubule assembly during pronucleus formation may be related to the cell cycle: In mice the sperm enters a meiotic cytoplasm, whereas in sea urchin eggs it enters an interphase cytoplasm. Refertilization of mitotic sea urchin eggs was performed to test the possibility that this phenomenon is related to whether the sperm enters a meiotic/mitotic cytoplasm or one at interphase; during refertilization at first mitosis, the incorporated sperm nucleus is unable to decondense and acquire lamins. These results indicate a requirement for microtubule assembly for the progression from meiosis to first interphase during mouse fertilization and suggest that the cytoskeleton is required for changes in nuclear architecture necessary during fertilization and the cell cycle.