No abstract

Thyroid hormones (T3) and their receptors (TR) play a critical role in the function of the pituitary gland, particularly in thyrotropes, where they regulate expression of the alpha- and beta-subunits of TSH. Since the pituitary gland is composed of several cell types, we undertook a characterization of TR subtypes in a murine thyrotropic tumor (TtT-97), an excellent model in which to study thyroid hormone action in thyrotropes. We screened a thyrotrope cDNA library with rat TR alpha 1 and TR beta 1 cDNA probes and isolated cDNAs encoding the mouse TR alpha 1 and TR beta 1 isoforms as well as a partial clone corresponding to the non-T3 binding carboxy-terminal alpha 2 variant. The polymerase chain reaction was used to amplify additional cDNAs for the specific 5' domains of the mouse TR beta 1 and the pituitary-specific TR beta 2 amino-terminal variant. Using hybridization probes that discriminate between the alpha and beta isoforms and their variants, we demonstrated that thyrotropes contain TR alpha 1 and alpha 2 mRNAs as well as transcripts encoding Rev-erbA, which arise by transcription from the opposite strand of the TR alpha gene. In thyrotropes, the ratio of alpha 2 to TR alpha 1 mRNA levels more closely resembled the distribution in mouse brain than that in heart, where the mRNA levels of TR alpha 1 and alpha 2 are comparable. TR beta 1 and TR beta 2 mRNAs were detected in thyrotropes and were of similar size (approximately 6.4 kilobases). Despite the almost complete conservation between the rat and mouse TR beta 1 sequences at the protein level, the mouse and rat TR beta 2-specific N-terminal domains were less conserved, and the mouse protein was shorter by 39 amino acids at the N-terminus. Of the receptor species, only the mRNA encoding the TR beta 2 isoform, which was restricted to thyrotropes, was decreased by T3 treatment, although the mRNA for the alpha 2 variant was also reduced by T3 in thyrotropes and heart tissue. Levels of TR beta 1 mRNA were not changed in liver, but were increased in thyrotropic tumors and also somewhat in brain, an organ that is not responsive to T3 by classical criteria.

We constructed a murine genomic DNA library in lambda EMBL3 and have isolated and determined the nucleotide sequence of the murine thyrotropin beta-subunit (TSH beta) gene. The cloned gene was derived from a thyrotropic tumor and had no detectable rearrangements when compared to the murine TSH beta gene in total genomic DNA. The murine TSH beta gene is 5 kb in size and consists of five exons and four introns. The 5' untranslated region of the mRNA is encoded except for a single nucleotide by exons 1, 2, and 3. The protein-coding regions are encoded by exons 4 and 5 while the 3' untranslated region is entirely contained in exon 5. Primer extension analysis using an exon 1-specific primer was used to map the 5' end of the gene. Two transcriptional start sites are present in the murine TSH beta gene which appear to be positioned by two TATAAA sequences located 40 bp apart. In all, 99% of transcripts initiate at the downstream site. Transcription from both start sites is affected by thyroidal status in both murine pituitaries and in TtT97 thyrotropic tumors. Finally, sequences homologous with putative thyroid-responsive elements and cyclic AMP-responsive elements are present in the 5'-flanking region and may be important in regulating negative and positive effects on TSH beta gene expression.

We have isolated and determined the nucleotide sequence of the mouse alpha-subunit gene of the pituitary glycoprotein hormones. No detectable rearrangements were observed from the cloned gene when compared with total genomic DNA. The gene is approximately 13.5 kb in size and consists of 4 exons and 3 introns. The 5' untranslated region is encoded by exon 1 and 14 bp of exon 2. A large first intron of about 10 kb interrupts the 5' untranslated region. The coding region is present in exons 2, 3, and 4, while the 3' untranslated region is contained entirely within exon 4. A number of mouse B1 and B2 repeats are present in the 5'-flanking region, the first and third introns, as well as in the 3'-flanking region. In contrast to the mouse beta-subunit gene of thyrotropin, primer extension analysis revealed that the alpha-subunit gene has a single transcriptional start site and that the primary transcript does not exhibit alternative exon splicing. The transcriptional start site is identical in both mouse pituitaries and in TtT97 thyrotropic tumors and is responsive to thyroidal status. The 5'-flanking region contains a sequence that is homologous to a single copy of the duplicated 18-bp cyclic AMP-responsive element of the human alpha-subunit gene. Sequences homologous to putative thyroid-responsive and estrogen responsive elements are present in the 5'-flanking region and may be important in the multihormonal regulation of the alpha-subunit gene in throtrophs and gonadotrophs, respectively.

Twenty-four strains of gram-positive facultative cocci, arranged primarily in small clusters, were isolated from the surface of the human tongue. With the exception of 14 catalase-negative isolates, these strains were identical in cultural and biochemical characteristics and in deoxyribonucleic acid base composition. All cultures produced viscous growth in both liquid and agar media. They fermented glucose anaerobically, reduced nitrate beyond nitrite, were benzidine-positive, and failed to grow in the presence of 5% NaCl or at 45 C. In addition, they exhibited guanine plus cytosine (G + C) contents of 55.4 to 58.3%. These isolates differed from strains of pediococci, aerococci, and micrococci which were included for comparison. On the basis of G + C content, these organisms appear to be intermediate between micrococci and staphylococci; however, on the basis of anaerobic glucose fermentation, it is suggested that they be placed in the genus Staphylococcus. It is proposed that they be recognized as S. salivarius. When gram-positive facultative cocci, other than streptococci, have been isolated from the oral cavity, different investigators have assigned them to one or more of the following genera: