INVESTIGADORES
MUÑOZ Manuel Javier
artículos
Título:
Influence of Polymerase II Processivity on Alternative Splicing Depends on Splice Site Strength
Autor/es:
GUADALUPE NOGUÉS, MANUEL J. MUÑOZ, AND ALBERTO R. KORNBLIHTT
Revista:
JOURNAL OF BIOLOGICAL CHEMISTRY
Referencias:
Año: 2003 vol. 278 p. 52166 - 52171
ISSN:
0021-9258
Resumen:
Transcription and pre-mRNA splicing are coordinated temporally
and spatially, and both processes can influence each other. In particular,
control of transcriptional elongation by RNA polymerase II has proved to be
important for alternative splicing regulation. In this report we demonstrate
that the efficiency of exon recognition by the splicing machinery is crucial for
the elongation control. Alternative splicing of the fibronectin extra domain I
(EDI) is because the polypyrimidine tract of its 3'-splice site occurs
suboptimal. By mutating the polypyrimidine tract of EDI in two different
positions, individually or in combination, and by disrupting its exonic splicing
silencer, we managed to generate minigenes with increasing degrees of exon
recognition. Improvement of exon recognition is evidenced by independence from
the splicing regulator SF2/ASF for inclusion. The mutated minigenes were used to
transfect human cells in culture and study the responsiveness of EDI alternative
splicing to activation or inhibition of pol II elongation. Our results revealed
that responsiveness of exon skipping to elongation is inversely proportional to
3'-splice site strength, which means that the better the alternative exon is
recognized by the splicing machinery, the less its degree of inclusion is
affected by transcriptional elongation.