Isolation, characterization, and expression of the gene encoding the late histone subtype H1-gamma of the sea urchin Strongylocentrotus purpuratus.

We cloned and characterized the gene encoding H1-gamma, a late histone subtype of the sea urchin species Strongylocentrotus purpuratus. The predicted primary sequence of H1-gamma is 216 amino acids in length and has a net charge of +70, which is high for a somatic H1 histone. The H1-gamma gene appears to be a unique sequence gene that is not tightly linked to the core histone genes. The 770-base-pair transcribed region of the H1-gamma gene is bordered on the 5' side by two previously described H1-specific sequence elements and on the 3' side by a hairpin loop structure and CAGA box sequences. We detected 3,900 stored maternal H1-gamma mRNA transcripts per egg. The number of H1-gamma transcripts per embryo rises by 9.5 h postfertilization, but the maximum rate of accumulation (4,300 molecules per min per embryo) occurs in the late-blastula-stage embryo between 14 and 21 h after fertilization. The number of H1-gamma mRNA molecules peaks 21 h after fertilization when there are 2.0 X 10(6) molecules per embryo (a 500-fold increase) and then decreases over the next 3.25 h to 1.3 million molecules per embryo. Between 24 and 82 h after fertilization the number of H1-gamma transcripts declines steadily (210 molecules per min per embryo) to reach approximately 5.4 X 10(5) H1-gamma mRNAs by 82 h postfertilization. Surprisingly, the number of late H1 mRNA molecules per embryo is greater than the number of late H2B mRNA molecules beginning at the early gastrula stage of development.

Comparison of the late H1 histone genes of the sea urchins Lytechinus pictus and Strongelocentrotus purpuratus.

We have isolated and sequenced a gene encoding a late H1 histone subtype from the sea urchin species L. pictus. The primary structure of the late H1 subtype encoded by this gene is 209 amino acids in length, and has a net positive charge of 67. This gene is present in a single copy per haploid genome and encodes an mRNA of 752 nucleotides. Late H1 transcripts are detected in the unfertilized egg and are most prevalent in gastrulating embryos. Comparison of 375 bp of 5' flanking sequences of the L. pictus late H1 gene and the H1-gamma gene of a distantly related sea urchin species, S. purpuratus, reveals large blocks of sequences that are identical between the two genes. To determine if these conserved 5' sequences are present in other members of the sea urchin H1 gene family, the analogous region of S. purpuratus H1-alpha, an early H1 gene, was sequenced. The homology between the flanking sequences of the early and late families was limited to consensus sequences which are found upstream of all H1 genes. The possible regulatory implications of these findings are discussed.

Temporal expression of late histone messenger RNA in the sea urchin Lytechinus pictus.

Sea urchin histones are encoded by several multigene families. The temporal expression of one of these families, the late histones, has been studied during the early development of Lytechinus pictus. Using a nuclease S1 assay, we detected about 10,000 transcripts encoding both late H3 and H4 proteins in the unfertilized egg. This suggests that the late genes were active at some point during oogenesis. The number of late gene transcripts begins to increase 6.5 hr after fertilization (64-cell stage), indicating that these genes probably become reactivated 4.5-6.5 hr after fertilization. The maximum rate of accumulation of transcripts (4600 molecules per min per embryo) occurs 9-14 hr after fertilization (from blastula stage to hatching). The number of transcripts peaks 21 hr after fertilization (onset of gastrulation) when the embryo has accumulated 1.8 X 10(6) copies of each late mRNA (a 164-fold increase). A 5.5-fold increase in the relative rate of transcription, between 7 and 15 hr after fertilization, is partly responsible for the accumulation of these gene products. The relative synthesis of early histone message, which is encoded by a different family, decreases 18-fold during this time. Synthesis of the late transcripts continues at the higher rate after accumulation has ceased (24 hr after fertilization). The number of late transcripts begins to decrease 48 hr after fertilization, reaching about 10,000 copies at 72 hr.

Transcriptional control of the isoleucine-valine messenger RNA's in E. coli K-12.

Hybridization of messenger ribonucleic acid (mRNA) isolated from Escherichia Coli K-12 to deoxyribonucleic acid (DNA) from lambdaCI857st68h80dilv was used to detect isoleucine-valine (ilv) specific mRNA. A number of strains partially constitutive for the isoleucine-valine enzymes had levels of ilv mRNA 2 to 3-fold higher than the parent strain. Starvation for any of the branched-chain amino acids resulted in a 20 to 23-fold increase in ilv mRNA as compared to repressed levels. These differences were not due to altered growth rates or to changes in the stability of ilv mRNA. These data indicate that regulation of the isoleucine-valine enzymes by multivalent repression occurs mainly at the level of transcription. Kinetics of elongation of ilv mRNA after repression are consistent with the assumption that the mechanism of multivalent repression involves the prevention of further initiations by RNA polymerase.