Communicated by Peter H. Byers
Molecular Basis of Two-Exon Skipping (Exons 12 and 13) by c.1248+5g>a in OXCT1 Gene: Study on Intermediates of OXCT1 Transcripts in Fibroblasts
Article first published online: 22 JAN 2013
© 2012 Wiley Periodicals, Inc.
Volume 34, Issue 3, pages 473–480, March 2013
How to Cite
Hori, T., Fukao, T., Murase, K., Sakaguchi, N., Harding, C. O. and Kondo, N. (2013), Molecular Basis of Two-Exon Skipping (Exons 12 and 13) by c.1248+5g>a in OXCT1 Gene: Study on Intermediates of OXCT1 Transcripts in Fibroblasts. Hum. Mutat., 34: 473–480. doi: 10.1002/humu.22258
Contract grant sponsors: Ministry of Health, Labor and Welfare of Japan; Ministry of Education, Culture, Sports, Science and Technology of Japan.
- Issue published online: 18 FEB 2013
- Article first published online: 22 JAN 2013
- Accepted manuscript online: 21 DEC 2012 12:54PM EST
- Manuscript Accepted: 30 NOV 2012
- Manuscript Received: 2 APR 2012
- Ministry of Health, Labor and Welfare of Japan
- Ministry of Education, Culture, Sports, Science and Technology of Japan
- two-exon skipping;
- splicing order;
- splice site;
- heteronuclear RNA;
- succinyl-CoA:3-ketoacid CoA transferase
The molecular basis of simultaneous two-exon skipping induced by a splice-site mutation has yet to be completely explained. The splice donor site mutation c.1248+5g>a (IVS13) of the OXCT1 gene resulted predominantly in skipping of exons 12 and 13 in fibroblasts from a patient (GS23) with succinyl-CoA:3-ketoacid CoA transferase (SCOT) deficiency. We compared heteronuclear RNA (hnRNA) intermediates between controls’ and GS23's fibroblasts. Our strategy was to use RT-PCR of hnRNA to detect the presence or absence of spliced exon clusters in RNA intermediates (SECRIs) comprising sequential exons. Our initial hypothesis was that a SECRI comprising exons 12 and 13 was formed first followed by skipping of this SECRI in GS23 cells. However, such a pathway was revealed to be not a major one. Hence, we compared the intron removal of SCOT transcript between controls and GS23. In controls, intron 11 was the last intron to be spliced and the removal of intron 12 was also rather slow and occurred after the removal of intron 13 in a major pathway. However, the mutation in GS23 cells resulted in retention of intron 13, thus causing the retention of introns 12 and 11. This “splicing paralysis” may be solved by skipping the whole intron 11–exon 12–intron 12–exon 13–mutated intron 13, resulting in skipping of exons 12 and 13.