Analysis of the 5′-Flanking Regions of Rat Inhibin α- and β-B-Subunit Genes Suggests Two Different Regulatory Mechanisms

Abstract

The genes encoding rat inhibin alpha- and beta-B-subunits were isolated and characterized. Both genes contain one intron that interrupts the region coding for the precursor portion of the alpha- and beta-B-subunits. The transcription start sites of alpha- and beta-B-subunit genes were determined by primer extension and nuclease mapping assay using mRNA from rat ovary and testis. Transcription of the alpha-subunit gene initiates predominantly at three adjacent sites with similar intensity. Several potential transcription start sites of beta-B-subunit gene are spread over 150 nucleotides upstream from translation initiation site. Neither of these two genes contains obvious TATA or CCAAT boxes. The alpha-subunit gene contains many GA clusters in the promoter region, while beta-B-subunit gene is highly GC rich. Several GGGCGG repeats and their inverted sequences, which are the potential binding sites for transcription factor Spl, were observed at the 5'-end as well as at the coding region of the beta-B-subunit gene. The potential cAMP-responsive element CTGCGTCAG was identified in alpha-but not beta-B-subunit gene. This sequence is identical to the cAMP- and phorbol ester-inducible DNA fragment found in human preproenkephalin gene. The different structure of the promoter region of rat alpha- and beta-B-subunit genes and the presence of a potential cAMP-inducible DNA sequence in alpha- but not beta-B-subunit gene is consistent with the hypothesis that transcription of alpha- and beta-B-subunit genes in rat is regulated by different mechanisms.

Genomic Cloning and Sequence Analyses of the Bovine α‐, βA‐ and βB‐Inhibin/activin Genes

“…A similar comparison of the bovine &inhibidactivin subunit with those of human (Mason et al, 1989), rat (Esch et al, 1987;Feng, et al, 1989), porcine (Mason et al, 1985) and ovine (Rodgers, 1991) sequences provided similarity scores of 95.1% (91.0%), 92.6% (86.4%), 94.3% (91.4%; partial porcine P,-subunit sequence) and 97.3 o/c (97.4%), respectively. The similarity between the two bovine Binhibid activin subunit amino acid sequences is 35.5% (42.5%), although the mature portions are more similar at 61.7% (62.9%).…”

“…[5'-ATG CCC TTG CTT TGG CTG AGA GGA TTT-3'; sequence identical to nucleotides 1-27 from the translational start site of the human (Mason et al, 1986), rat (Esch et al, 1987) and porcine (Mason et al, 1985) PA-subunit genes], GAC TTC C-3'; sequence based on consensus of nucleotides 193-223 relative to the translational start site of the human PB subunit (Mason et al, 1989) in comparison with the rat (Feng et al, 1989) and porcine &subunit sequences (Mason et al, 1985)] and oPB3 [5'-GAG TAC AAC ATC GTC AAG CGG GAC GTG CCC-3'; sequence identical to nuclrotides 1162-1191 and 1144-1173 relative to the translational start sites of the human (Mason et al, 1989) and rat (Esch et al, 1987;Feng et al, 1989) P,-subunit-coding sequences, respectively, and to a segment near the 3' end of the porcine P,-subunit-coding sequence (Mason et al, 1985)]. …”

“…A comparison of the sequence data obtained for part of the 2.2-kb Sac1 fragment with the P,-inhibidactivin cDNA and genomic sequences indicated that it contained 867 bp of 5'-flanking bovine PB DNA, 451 bp of the structural gene and approximately 0.9 kb intron. The exod intron junction was deduced by comparison of the determined sequence data with that reported for the rat (Feng et al, 1989), sheep (Rodgers, 1991) and human (Mason et al, 1989) P,-inhibidactivin genes, together with the likely exod intron sequence (the 'GT-AG' rule; see Mount, 1982) and the sequences of the human (Mason et al, 1986), rat (Esch et al, 1987) and pig (Mason et al, 1985) P,-subunit cDNAs. The 4.5-kb XhoI fragment overlapped the 2.2-kb SacI fragment and was used to obtain an additional 1052 bp of 5'-flanking sequence.…”

“…The sequences and structural features of the inhibidactivin subunits has led to their inclusion as members of the TGF-p superfamily (for reviews see Kingsley, 1994;MassaguC et al, 1994). Genomic clones of the a-inhibin gene from man (Stewart et al, 1986;Mason et al, 1987), rat (Feng et al, 1989;Albiston et al, 1990;Pei et al, 1991) and mouse ( Su and Hsueh, 1992), the human P,-inhibin/activin gene (Stewart et al, 1986;Mason et al, 1987;Tanimoto et al, 1991) and the pBinhibidactivin gene from man (Forage et al, 1987;Mason et al, 1987;Mason et al, 1989), rat (Feng et al, 1989) and sheep (Rodgers, 1991) have been isolated and sequenced. Comparisons of the inhibidactivin genomic clones with their cDNAs reveal that the a-, PA-and P,-inhibidactivin genes are each comprised of two exons separated by an intron of approximately 1.5-2.1, 9-10 and 2.6-3.0 kb, respectively.…”

GATA Factors and Androgen Receptor Collaborate To Transcriptionally Activate theRhox5Homeobox Gene in Sertoli Cells

“…This postmitotic somatic cell, which is in intimate contact with developing germ cells in seminiferous tubules in the testis, is crucial for all phases of male gametogenesis, including germ cell proliferation, meiosis, and differentiation (15). Genes necessary for the Sertoli cell to support male gametogenesis have been identified, and some progress has been made in identifying regulatory mechanisms that control the transcription of some of these genes, including follicle-stimulating hormone receptor (Fshr), cathepsin L, the inhibin-␤ B subunit, transferrin, Mullerian inhibitory substance, androgen-binding protein, Dnmrt 1, GATA1, GATA4, and tissue plasminogen activator (12,13,15,17,18,21,35,41,43,53). Regulatory regions and specific cis elements that participate in the transcriptional regulation of these genes have been identified in primary Sertoli cells and Sertoli cell lines.…”