Differential translatability in vitro of multiple messenger RNAs encoding the beta-subunit of mouse thyrotropin.
The mouse TSH beta gene contains two start sites of transcription and exhibits alternative splicing among its first three exons, which encode 5'-untranslated mRNA sequences. Expression of the mouse TSH beta gene, therefore, gives rise to multiple mRNAs, each with a unique 5'-untranslated region. We have determined the relative translational efficiencies of these mRNAs in vitro, and we demonstrate that one of them directs the synthesis of a novel TSH beta presubunit. The four TSH beta mRNAs that are expressed from the down-stream transcription start site (TSS2) and the major mRNA derived from the up-stream start site (TSS1) were transcribed in vitro and translated in reticulocyte lysates and wheat germ extracts. The mRNA from TSS1 gave a novel TSH beta presubunit due to initiation of translation at an up-stream AUG unique to this mRNA. The novel presubunit contained a 17-amino acid NH2-terminal extension sequence, compared to the normal TSH beta presubunit, which is encoded by each of the mRNAs from TSS2. Despite the fact that the NH2-terminal extension sequence appeared to lack the characteristics of a signal peptide, the novel TSH beta presubunit was processed about 50% as efficiently by microsomal membranes as the normal presubunit, with glycosylation and cleavage by signal peptidase. There was an approximately 2-fold range in relative translatability among the different TSH beta mRNAs, and the mRNA encoding the novel TSH beta presubunit had the highest translational efficiency. Our data, therefore, suggest that the longer presubunit may be synthesized in vivo in significant amounts and give rise to a novel mature TSH beta subunit.
Department of Biochemistry, Temple University School of Medicine, Philadelphia, Pennsylvania 19140.
M M
MM
Januszeski
J A
JA
Gurr
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002874:Chromosome Mapping; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D015870:Gene Expression; D066298:In Vitro Techniques; D015636:Magnesium Chloride / Q000378:metabolism; D051379:Mice; D008815:Mice, Inbred Strains; D008861:Microsomes / Q000378:metabolism; D008969:Molecular Sequence Data; D010902:Pituitary Gland / Q000378:metabolism; D010957:Plasmids; D016133:Polymerase Chain Reaction; D011189:Potassium Chloride / Q000494:pharmacology; D014176:Protein Biosynthesis / Q000187:drug effects; D012333:RNA, Messenger / Q000235:genetics*; D012156:Reticulocytes / Q000378:metabolism; D013972:Thyrotropin / Q000096:biosynthesis*; D013997:Time Factors; D014158:Transcription, Genetic
D012333:RNA, Messenger; D015636:Magnesium Chloride; D011189:Potassium Chloride; D013972:Thyrotropin