Role In Cleavage Research Articles

Early embryonic development of the dipteran insect Heteropeza pygmaea in the presence of cytoskeleton-affecting drugs.

Embryos of the paedogenetically reproducing gall midge Heteropeza pygmaea develop floating in the haemocoel of a so-called mother larva. The egg membranes remain permeable and the embryos increase in size during embryonic development by taking up nutrients from the haemolymph. Such embryos can be cultured in vitro, i.e. in haemolymph drops obtained from mother larvae. We tested the effects of several drugs known to interact with cytoskeletal elements on different stages of embryonic development, including cleavage and gastrulation. The drugs were added to the in vitro cultures and the effects were studied with time-lapse cine-micrography. Colchicine and vinblastine blocked cleaving eggs in metaphase stage and arrested yolk globule oscillation. In spite of such a block blastoderms once formed continued development through germ band formation and extension and also increased in size. Cytochalasin B did not affect the stage of cleavage; however, it inhibited gastrulation and subsequent morphogenetic processes and also prevented size increase. We conclude that (1) the functioning of microtubules is needed for yolk globule oscillation during cleavage interphases but not for the gastrulation processes subsequent to blastoderm formation and (2) microfilaments do not play an important role in cleavage, at least not for the orderly succession of the cleavage divisions, but are essential for the morphogenetic movements associated with gastrulation. We suggest that during cleavage a limited stock of microtubules and their precursors is responsible for both transport of chromosomes during mitoses and translocation of organelles during interphase. Yolk oscillation seems to be a secondary effect and of minor or no importance for the normal course of embryonic development.

Sequence requirements for splicing of higher eukaryotic nuclear pre-mRNA

We determined the effect on splicing of 24 point mutations in the 5′ and 3′ splice region of the large rabbit β-globin intron. In vitro, 3′ AG mutations drastically reduce 5′ cleavage and abolish splicing. In vivo, the same mutations elicit efficient splicing at a cryptic, rather than the correct, 3′ splice site. In vitro, mutations at all but 2 positions of the consensus 5′ splice region impair correct splicing and promote joining of exon 1 to exon 3. In vivo, the same mutations show no effect, except for those converting 5′ GT to AT or GA, which cause accumulation of lariat intermediate in vitro and in vivo. We conclude that the 5′ GT need not be conserved for 5′ cleavage and that it plays an important role in cleavage and exon joining at the 3′ splice site.

Glycyl-glycine hydrolase of rat brain: Distribution and role in cleavage of glycine-rich oligopeptides

STERN, F. AND N. MARKS. Glycyl-glycine hydrolase of rat brain: Distribution and role in cleavage of glycine-rich oligopeptides. BRAIN RES. BULL. 4(1) 49–55, 1979.—The specificity and distribution of glycyl-glycine hydrolase (E.C.3.4.13.1) were studied in rat brain by several analytical procedures. Compared to other dipeptides Gly-Gly was unique in terms of its activation by Co 2+, which had no effect on other substrates or inhibited their breakdown. Gly-Gly was cleaved at a rate of 180 μmoles per g wet wt. per hr in the absence of Co 2+. It rose 4-fold on addition of Co 2+, and 2-fold with Mn 2+; it is thus comparable in rate to many other exopeptidases. Its pH optimum was 7.9 with an apparent K m of 4.3 in the absence of metal ion, and 2.7 in its presence. For glycine-rich tri- and oligopeptides, activation by metal ion occurred only with substrates yielding Gly-Gly as an intermediate (Gly-Gly-Gly, Leu-Gly-Gly, enkephalin, delta sleep inducing peptide). Activity was highest in striatum and cerebellum and lowest in spinal cord and pituitary. In subcellular fractions over 40% of the activity was present in the cytosol, with lesser amounts in the debris, nuclear and mitochondria! fractions. Activity increased during development, reaching maximum values during myelination. The enzyme was partially purified by CM-cellulose chromatography or isoelectric focusing; it was stable when stored at −70°C. Crude and partially purified enzymes were inhibited by Zn 2+, Ni 2+, Fe 2+, a number of sulfhydryl reagents, and EDTA. Enzyme activity in vitro greatly exceeded that required to degrade Gly-Gly generated as a result of normal protein turnover and may account for the absence of this dipeptide in soluble extracts.

The role of cleavage in the localization of developmental potential in the Ctenophore Mnemiopsis leidyi

This study defines the time period during which the cellular components that specify comb plates and photocytes become localized in different parts of blastomeres prior to their segregation to separate daughter cells. At the two-cell stage the factors which specify comb plates are localized at the aboral pole of the blastomeres. There is not a significant localization of the factors which specify comb plates and photocytes along the tentacular axis of the embryo. At the four-cell stage, the factors which specify comb plates become localized at one end of the tentacular axis of the blastomeres; however, the factors which specify photocytes have not yet become localized. At the eight-cell stage, the factors which specify these two cell types are segregated to different blastomeres. The role of cleavage in setting up these localized regions of developmental potential has been studied by reversibly inhibiting selected cleavages. After the first division, the pattern of cleavage that follows a period of cleavage inhibition corresponds to the pattern occurring in untreated embryos that began development at the same time. This situation is similar to the “clock” system, which controls many aspects of the pattern of cleavage in sea urchin embryos. The extent to which the factors that specify comb plates and photocytes become localized in a given region of a blastomere is correlated with the kind of cleavage which occurs after a block. Most of the activity involved in localizing developmental potential takes place during cleavage.