Alternative splicing and translation of glutamine synthetase gene = 글루타민 합성효소 유전자의 선택적 스플라이싱과 번역

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The expression of glutamine synthetase (GS), catalyzing the ATP-dependent conversion of glutamate and ammonia into glutamine, is transcriptionally and post-transcriptionally regulated. The genomic structure of canine GS shown here is basically similar to that of other mammals in that it is composed of seven exons and six introns. Using 5’cRACE and RT-PCR, we identified an additional exon (120 bp) in the first intron, designated here as exon 1’. By means of alternative splicing, the GS gene produces an altered form of GS transcript with 5’-untranslated region (UTR) containing the exon 1’. This alternative transcript is abundantly expressed in brain, while it is found at lower level in other tissues. In the human and mouse GS genes, extra exons are also found at corresponding site of the intron 1 but with different sizes. An exon trapping experiment for the GS gene in COS-7, MDCK, and SK-N-SH cells revealed that the pattern of alternative splicing is variable in different cell types. The propensity of forming a secondary structure is predicted to be considerably higher with the presence of extra 5’-UTR, suggesting a possibility of translational effect. In order to test this, we carried out a reporter assay for fusions with different 5’-UTRs, demonstrating that the long form with extra 5’-UTR was translated 20- and 10-fold less than the short one in SK-N-SH and Neuro-2A cells, respectively. Similarly, translations of human and mouse transcripts with extra 5’-UTRs were less efficient, showing 6-8 fold reductions in SK-N-SH cells. Furthermore, when we mutated an ATG sequence contained in the exon 1’, the suppression of translation was partially relieved, suggesting that the negative regulation by an extra 5’-UTR is, to some extent, due to an abortive translation from the upstream ATG. This regulation was confirmed by comparing the in vitro translation products of capped in vitro transcripts for the short and long transcripts. From this experiment, we found that two ...
Park, Chan-Kyuresearcher박찬규researcher
한국과학기술원 : 생물과학과,
Issue Date
231025/325007  / 000995213

학위논문(박사) - 한국과학기술원 : 생물과학과, 2003.8, [ viii, 115 p. ]


Alternative splicing; Glutamine synthetase gene; Translational regulation; 번역 조절; 선택적 스플라이싱; 글루타민 합성효소 유전자

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