Ovothiol replaces glutathione peroxidase as a hydrogen peroxide scavenger in sea urchin eggs.

Despite its potential toxicity, H2O2 is used as an extracellular oxidant by Stronglylocentrotus purpuratus eggs to cross-link their fertilization envelopes. These eggs contain 5 mM 1-methyl-N alpha,N alpha-dimethyl-4-mercaptohistidine (ovothiol C), which reacts with H2O2. In consuming H2O2 and being reduced by glutathione, ovothiol acts as a glutathione peroxidase and replaces the function of the enzyme in eggs. The ovothiol system is more effective than egg catalase in destroying H2O2 at concentrations produced during fertilization and constitutes a principal mechanism for preventing oxidative damage at fertilization.

Isolation and characterization of sequences from mouse chromosomal DNA with ARS function in yeasts.

Fragments of chromosomal DNA from a variety of eucaryotes can act as ARSs (autonomously replicating sequence) in yeasts. ARSs enable plasmids to be maintained in extrachromosomal form, presumably because they function as initiation sites for DNA replication. We isolated eight different sequences from mouse chromosomal DNA which function as ARSs in Saccharomyces cerevisiae (bakers' yeast). Although the replication efficiency of the different mouse ARSs in yeasts appears to vary widely, about one-half of them functions as well as the yeast chromosomal sequence ARS1. Moreover, five of the ARSs also promote self replication of plasmids in Schizosaccharomyces pombe (fission yeast). Each of the ARSs was cloned into plasmids suitable for transformation of mouse tissue culture cells. Plasmids were introduced into thymidine kinase (TK)-deficient mouse L cells by the calcium phosphate precipitation technique in the absence of carrier DNA. In some experiments, the ARS plasmid contained the herpes simplex virus type 1 TK gene; in other experiments (cotransformations), the TK gene was carried on a separate plasmid used in the same transformation. In contrast to their behavior in yeasts, none of the ARS plasmids displayed a significant increase in transformation frequency in mouse cells compared with control plasmids. Moreover, only 1 of over 100 cell lines contained the original plasmid in extrachromosomal form. The majority of cell lines produced by transformation with an ARS TK plasmid contained multiple copies of plasmid integrated into chromosomal DNA. In most cases, results with plasmids used in cotransformations were similar to those for plasmids carrying TK. However, cell lines produced by cotransformations with plasmids containing any one of three of the ARSs (m24, m25, or m26) often contained extrachromosomal DNAs.

Glucocorticoid induction of egg white mRNAs in chick oviduct.

Glucocorticoids induce ovalbumin and conalbumin mRNA in oviducts from chicks withdrawn from prior estrogen treatment. The magnitude and the kinetics of the responses obtained, either in vivo or in vitro, are comparable to those obtained with estrogen or progesterone. With cultured oviducts, 1 nM dexamethasone is sufficient for half-maximal accumulation of nuclear receptors and partial induction of both mRNAs, while maximal levels of receptors and both mRNAs are achieved with 30 to 100 nM dexamethasone. Competition experiments show that dexamethasone acts by binding to a class of high affinity receptors distinct from the sex steroid receptors. Dexamethasone acts synergistically with estrogen, but not with progesterone.

A somatomedin-like peptide hormone is required during the estrogen-mediated induction of ovalbumin gene transcription.

Expression of the ovalbumin gene in chicken oviduct explant cultures requires the presence of a somatomedin-like peptide hormone in addition to estrogen. Insulin, proinsulin and multiplication-stimulating activity (MSA) are equally active substitutes for this peptide hormone, and maximal induction requires about 0.5 micrograms/ml; fetal calf serum can partially substitute for these factors. The equipotency of insulin and proinsulin indicates that insulin receptors are not involved, and the activity of MSA suggests that the active receptor is specific for somatomedins. The permissive effect of the peptide factor occurs within 1-2 hr and is required for the initiation of estrogen-mediated transcription on the ovalbumin gene. In contrast, transcription from the conalbumin gene is fully induced by estrogen in the presence or absence of peptide factors or serum, despite the fact that these two egg white genes are both transcribed in the same cells in response to the same steroid hormones. We suggest that the interaction of a somatomedin with its membrane-bound receptor generates an intracellular signal that interacts specifically with the ovalbumin gene.

Ovothiols, a family of redox-active mercaptohistidine compounds from marine invertebrate eggs.

We have previously reported a novel thiol compound, 1-methyl-N alpha,N alpha-dimethyl-4-mercaptohistidine, or ovothiol, present at high concentration in the eggs of the sea urchin Strongylocentrotus purpuratus [Turner, E., Klevit, R., Hopkins, P. B., & Shapiro, B. M. (1986) J. Biol. Chem. 261, 13056-13063]. Here we report two related compounds, 1-methyl-N alpha-methyl-4-mercaptohistidine, or ovothiol B, from the scallop Chlamys hastata, and 1-methyl-4-mercaptohistidine, or ovothiol A, from the starfish Evasterias troschelii. These two compounds, as well as the S. purpuratus compound now designated ovothiol C, were isolated from eggs or ovarian tissue by S-carboxymethylation with [3H]iodoacetic acid, ion-exchange chromatography and ion-pairing high-pressure liquid chromatography. The structures of S-(carboxymethyl)ovothiols A and B were determined by 1H NMR, and that of ovothiol A was confirmed by comparison with authentic methylhistidine samples after desulfuration with Raney nickel. In the ovary of each species, the predominant methylation form of ovothiol accounts for at least 80% of the total 4-mercaptohistidine. The ovothiol concentration of the ovary far exceeds that of the testis or somatic tissues. The ovothiol C content of unfertilized S. purpuratus eggs is 1.14 mumol/10(6) eggs, equivalent to approximately 4.3 mM average concentration; the glutathione (GSH + GSSG) content is 0.9 mumol/10(6) eggs. In this species, high ovothiol levels persisted for the first 2 weeks of embryonic development. Ovothiol and glutathione account for virtually all of the trichloroacetic acid soluble-SH groups in the egg; these results are compared to several previous studies.(ABSTRACT TRUNCATED AT 250 WORDS)