A subfragment of the beta tropomyosin gene is alternatively spliced when transfected into differentiating muscle cells.
A subgenomic fragment of the chicken beta tropomyosin gene which contains two alternative exons flanked by common exons was isolated and placed under the control of the SV 40 early promoter. This construction was subsequently used to transfect quail myoblasts together with a Neomycin resistance gene, and to isolate stable transfectants. mRNAs were isolated before and after differentiation and analyzed using a modification of the primer extension method. We show that myoblasts accumulate transcripts which contain the non muscle specific exon joined to the common exons while myotubes accumulate transcripts containing the muscle specific exon. These results, therefore demonstrate that such a subgenomic fragment contains all the necessary information to direct a correct developmentally regulated mutually exclusive splicing. They also strongly suggest that trans acting factors must be involved in the switch of the splicing pattern which takes place during the transition from myoblasts to myotubes. The same regulation cannot be faithfully reproduced during transient expression, since no difference in the use of exons 6A/6B is observed during differentiation and two aberrant minor splicing products are obtained which contain or lack both exons. We suggest that failure of exon 6A to splice to exon 6B is due to the existence of some structural constraints which lower the efficiency with which the intron between them is excised.
Libri
D
D
Molecular Biology Department, Pasteur Institute, Paris, France.
Marie
J
J
Brody
E
E
Fiszman
M Y
MY
D000818:Animals; D002645:Chickens; D004625:Embryo, Nonmammalian; D005091:Exons; D005796:Genes; D009132:Muscles / Q000378:metabolism; D009154:Mutation; D010957:Plasmids; D011401:Promoter Regions, Genetic; D011784:Quail; D012326:RNA Splicing; D012333:RNA, Messenger / Q000235:genetics*; D015183:Restriction Mapping; D013539:Simian virus 40 / Q000235:genetics; D014158:Transcription, Genetic; D014162:Transfection; D014335:Tropomyosin / Q000235:genetics*
D012333:RNA, Messenger; D014335:Tropomyosin