Poly(ADP‐ribosylation) of atypical CS histone variants is required for the progression of S phase in early embryos of sea urchins

Abstract

The patterns of poly(ADP-ribosylation) in vivo of CS (cleavage stage) histone variants were compared in sea urchin zygotes at the entrance and the exit of S1 and S2 in the initial developmental cell cycles. This post-translational modification was detected by Western immunoblots with rabbit sera anti-poly(ADP-ribose) that was principally reactive against ADP-ribose polymers and slightly against ADP-ribose oligomers. The effect of 3 aminobenzamide (3-ABA), an inhibitor of the poly(ADP-ribose) synthetase, on S phase progression was determined in vivo by measuring the incorporation of 3H thymidine into DNA. The results obtained indicate that the CS histone variants are poly(ADP-ribosylated) in a cell cycle dependent manner. A significantly positive reaction of several CS variants with sera anti-poly(ADP-ribose) was found at the entrance into S phase, which decreases after its completion. The incubation of zygotes in 3-ABA inhibited the poly(ADP-ribosylation) of CS variants and prevented both the progression of the first S phase and the first cleavage division. These observations suggest that the poly(ADP-ribosylation) of atypical CS histone variants is relevant for initiation of sea urchin development and is required for embryonic DNA replication.

Decreased heterogeneity of CS histone variants after hydrolysis of the ADP-ribose moiety

“…The evidence presented in this report confirms our previous suggestions that CS histone variants are extensively poly(ADP-ribosylated) in zygotes of sea urchins as well as CS variants present in unfertilized eggs (Imschenetzky et al, 1991a;1992). Our conclusion is based on three major lines of evidence: (1) the incorporation of 3H-adenosine in vivo into poly(ADPribosylated) CS histone variants during the initial cell cycles of sea urchin zygotes; (2) the immunodetection of the poly(ADP-ribose) chains with polyclonal antibodies elicited against polymers of ADP-ribose; and (3) the detection of the ribose residues present in the poly(ADP-ribose) moiety by its reaction with dansyl hydrazine.…”

“…The main considerations to homologue the function of CS histone variants with histones can be summarized as follows: CS variants pack the DNA into small nucleo-protein particles which in turn are further organized into higher ordered structures [Imschenetzky et al, 1989al. CS variants are extensively poly(ADP-ribosylated) in response to embryonic demands for DNA replication and/or DNA repair [Imschenetzky et al, 1991a;1992. The synthesis de novo of the CS variants is confined to the S phase of the cell cycle in early zygotes and the newly synthesized CS variants are immediately targeted to nuclei and bound to DNA [Imschenetzky et al, 1995b1, as well as histones from most cells [Stein et al, 19941.…”

“…Based on immunobiochemical detection of poly(ADP-ribosylated) CS histone variants we have previously suggested that this heterogeneity may be due to the extensive poly-(ADP-ribosylation) of these proteins. We have also shown that this post-translational modification is a pre-requisite for S phase in zygotes [Imschenetzky et al, 1991a;1992;19931. It is well established that this post-translational modification induces a great degree of electrophoretic heterogeneity. This is reflected by a family of subbands which migrate depending on the length of ADP-ribose moiety bound to the protein [Huletsky et al, 1985;Althaus and Richter, 1987;Boulikas, 1990;Boulikas et al, 19903.…”

Identification of a cysteine protease responsible for degradation of sperm histones during male pronucleus remodeling in sea urchins

“…The substrate selectivity shown by the cysteineprotease described in this report strongly suggests its role in the degradation of SpH, which in turn determines male pronucleus chromatin remodeling. This selectivity may be explained by the extensive poly(ADP-ribosylation) of CS histone variants in unfertilized eggs and zygotes (Imschenetzky et al, 1991b(Imschenetzky et al, , 1993(Imschenetzky et al, , 1996b. We speculate that the high concentration of negative charges provided by the ADP-ribose moieties bound to the CS histone variants may protect these proteins from proteolysis by this enzyme.…”

Chromatin remodeling during sea urchin early development: molecular determinants for pronuclei formation and transcriptional activation

“…In contrast, the CS histone variants either in unfertilized eggs or during the initial cleavage cycles are extensively poly(ADP-ribosylated). This modification is a requirement for a correct initiation of the cleavage divisions and varies at different stages of the initial cell cycle (Imschenetzky et al, 1991b(Imschenetzky et al, , 1993(Imschenetzky et al, , 1996a. Poly(ADP-ribosylation) of chromosomal proteins is catalyzed by the poly(ADP-ribose) polymerase (PARP), a nuclear enzyme found in most eukaryotic cells.…”