MAZ Elements Alter Transcription Elongation and Silencing of the Fibroblast Growth Factor Receptor 2 Exon IIIb

Abstract

The fibroblast growth factor receptor 2 (FGFR2) gene exons IIIb and IIIc are alternatively spliced in a mutually exclusive and cell type-specific manner. FGFR2 exon choice depends on both activation and silencing. Exon IIIb silencing requires cis-acting elements upstream and downstream of the exon. To examine the influence of transcription on exon IIIb silencing, the putative RNA polymerase II (RNAPII)-pausing MAZ4 element was inserted at different positions within the FGFR2 minigene construct. MAZ4 insertions 5′ to the upstream silencing elements or between exon IIIb and downstream silencing elements result in decreased silencing. An insertion 3′ of the downstream silencing elements, however, has no effect on splicing. An RT-PCR elongation assay shows that the MAZ4 site in these constructs is likely to be a RNAPII pause site. Insertion of another RNAPII pause site into the minigene has a similar effect on exon IIIb silencing. Transfection of in vitro transcribed RNA demonstrates that the cell type specificity of FGFR2 alternative splicing requires co-transcriptional splicing. Additionally, changing the promoter alters both FGFR2 minigene splicing and the MAZ4 effect. We propose that RNAPII pauses at the MAZ4 elements resulting in a change in the transcription elongation complex that influences alternative splicing decisions downstream. The fibroblast growth factor receptor 2 (FGFR2) gene exons IIIb and IIIc are alternatively spliced in a mutually exclusive and cell type-specific manner. FGFR2 exon choice depends on both activation and silencing. Exon IIIb silencing requires cis-acting elements upstream and downstream of the exon. To examine the influence of transcription on exon IIIb silencing, the putative RNA polymerase II (RNAPII)-pausing MAZ4 element was inserted at different positions within the FGFR2 minigene construct. MAZ4 insertions 5′ to the upstream silencing elements or between exon IIIb and downstream silencing elements result in decreased silencing. An insertion 3′ of the downstream silencing elements, however, has no effect on splicing. An RT-PCR elongation assay shows that the MAZ4 site in these constructs is likely to be a RNAPII pause site. Insertion of another RNAPII pause site into the minigene has a similar effect on exon IIIb silencing. Transfection of in vitro transcribed RNA demonstrates that the cell type specificity of FGFR2 alternative splicing requires co-transcriptional splicing. Additionally, changing the promoter alters both FGFR2 minigene splicing and the MAZ4 effect. We propose that RNAPII pauses at the MAZ4 elements resulting in a change in the transcription elongation complex that influences alternative splicing decisions downstream. The processes of transcription and splicing are spatially and temporally linked within the cell (1Goldstrohm A.C. Greenleaf A.L. Garcia-Blanco M.A. Gene (Amst.). 2001; 277: 31-47Crossref PubMed Scopus (143) Google Scholar, 2Maniatis T. Reed R. Nature. 2002; 416: 499-506Crossref PubMed Scopus (925) Google Scholar), and these links may be critical for both constitutive splicing and for the regulation of alternative splicing. Recruitment of several different transcriptional activators to a promoter controls the splicing efficiency of a constitutively spliced Drosophila doublesex intron; strong activators induce higher levels of splicing than weak activators (3Rosonina E. Bakowski M.A. McCracken S. Blencowe B.J. J. Biol. Chem. 2003; 278: 43034-43040Abstract Full Text Full Text PDF PubMed Scopus (56) Google Scholar). This result is not merely an indirect effect of the relative concentration of splicing precursors since the efficiency of splicing does not correlate with the level of transcripts (3Rosonina E. Bakowski M.A. McCracken S. Blencowe B.J. J. Biol. Chem. 2003; 278: 43034-43040Abstract Full Text Full Text PDF PubMed Scopus (56) Google Scholar). Similar promoter-dependent effects on splicing have been demonstrated for the alternatively spliced ED1 exon of the fibronectin gene (4Cramer P. Pesce C.G. Baralle F.E. Kornblihtt A.R. Proc. Natl. Acad. Sci. U. S. A. 1997; 94: 11456-11460Crossref PubMed Scopus (270) Google Scholar, 5Cramer P. Caceres J.F. Cazalla D. Kadener S. Muro A.F. Baralle F.E. Kornblihtt A.R. Mol. Cell. 1999; 4: 251-258Abstract Full Text Full Text PDF PubMed Scopus (257) Google Scholar, 6Kadener S. Cramer P. Nogues G. Cazalla D. de la Mata M. Fededa J.P. Werbajh S.E. Srebrow A. Kornblihtt A.R. EMBO J. 2001; 20: 5759-5768Crossref PubMed Scopus (110) Google Scholar, 7Nogues G. Kadener S. Cramer P. Bentley D. Kornblihtt A.R. J. Biol. Chem. 2002; 277: 43110-43114Abstract Full Text Full Text PDF PubMed Scopus (144) Google Scholar, 8Kadener S. Fededa J.P. Rosbash M. Kornblihtt A.R. Proc. Natl. Acad. Sci. U. S. A. 2002; 99: 8185-8190Crossref PubMed Scopus (97) Google Scholar). Transcription factors that alter the rate of transcript initiation have no effect on splicing, but transcription factors that alter elongation rate affect ED1 inclusion (6Kadener S. Cramer P. Nogues G. Cazalla D. de la Mata M. Fededa J.P. Werbajh S.E. Srebrow A. Kornblihtt A.R. EMBO J. 2001; 20: 5759-5768Crossref PubMed Scopus (110) Google Scholar, 7Nogues G. Kadener S. Cramer P. Bentley D. Kornblihtt A.R. J. Biol. Chem. 2002; 277: 43110-43114Abstract Full Text Full Text PDF PubMed Scopus (144) Google Scholar). Conversely, splicing influence the efficiency of gene transcription in an effect the splicing factor A. A. 2002; PubMed Scopus Google Scholar). Additionally, transcript initiation and elongation both be splicing factors in vitro 2001; PubMed Scopus Google Scholar, Nature. 2001; PubMed Scopus Google Scholar, S. A. A. Biol. 2002; Google Scholar). is a influence between transcription and splicing, is of the for the effect of different transcriptional activators on splicing is that transcription factors may splicing factors to the transcription complex splicing several splicing factors are with transcription factors with transcriptional activators with and transcriptional with A.C. Garcia-Blanco M.A. Mol. Cell. Biol. 2001; PubMed Scopus Google Scholar, 2002; PubMed Scopus Google Scholar). different transcriptional activators may alter transcript elongation alternative splicing in elongation rate have been to splicing in both M. 2003; PubMed Scopus Google and la Mata M. Kadener S. Fededa J.P. M. Cramer P. Bentley D. Kornblihtt A.R. Mol. Cell. 2003; Full Text Full Text PDF PubMed Scopus Google Scholar). both in RNA polymerase II RNA polymerase fibroblast growth factor receptor downstream fibroblast growth splicing upstream splicing downstream splicing splicing and that elongation rate alter alternative splicing in transcription elongation rate may alternative splicing of the gene PubMed Scopus Google Scholar). PubMed Scopus Google demonstrated that insertion of a MAZ4 element between the alternatively spliced exon and downstream element in exon The MAZ4 element of the that is of a RNAPII pause element downstream of the gene R. P. EMBO J. PubMed Scopus (56) Google Scholar). The MAZ4 has been to induce RNAPII in is no that the MAZ4 element is of RNAPII in R. P. EMBO J. PubMed Scopus (56) Google Scholar, M. Mol. Cell. 1999; Full Text Full Text PDF PubMed Scopus Google Scholar, M. EMBO J. PubMed Scopus Google Scholar). these PubMed Scopus Google a in a of of the the MAZ4 site in for exon exon The effect of transcription on both silencing and activation of alternatively spliced exons be the fibroblast growth factor receptor 2 (FGFR2) gene exons are alternatively spliced in a mutually exclusive and cell type-specific manner. Exon is in and exon is in T. Proc. Natl. Acad. Sci. U. S. A. PubMed Scopus Google Scholar, E. P. R. Mol. Cell. Biol. PubMed Scopus Google Scholar). of exon IIIb or IIIc the specificity of the receptor T. Proc. Natl. Acad. Sci. U. S. A. PubMed Scopus Google Scholar, M. D. M. Proc. Natl. Acad. Sci. U. S. A. 2003; PubMed Scopus Google Scholar). The IIIb to and the IIIc to T. Proc. Natl. Acad. Sci. U. S. A. PubMed Scopus Google Scholar). of of FGFR2 to in and in exon IIIc have been with several G. G. S. Mol. Cell. Biol. PubMed Scopus Google Scholar, Garcia-Blanco M.A. 1997; PubMed Scopus Google Scholar, J. M. PubMed Google Scholar, S. Proc. Natl. Acad. Sci. U. S. A. 2001; PubMed Scopus Google Scholar, J. 2002; PubMed Scopus Google Scholar). Additionally, in exon inclusion have been with to in both and G. G. S. Mol. Cell. Biol. PubMed Scopus Google Scholar, Garcia-Blanco M.A. 1997; PubMed Scopus Google Scholar). the are are and are are and G. G. S. Mol. Cell. Biol. PubMed Scopus Google Scholar). The alternative splicing of FGFR2 exons IIIb and IIIc is both and splicing elements of exon IIIb in is weak an splicing upstream splicing and and the downstream element downstream splicing and the 2 E. P. R. Mol. Cell. Biol. PubMed Scopus Google Scholar, R. Mol. Cell. Biol. PubMed Scopus Google Scholar, Garcia-Blanco M.A. Mol. Cell. Biol. 20: PubMed Scopus Google Scholar, Garcia-Blanco M.A. Mol. Cell. 2002; Full Text Full Text PDF PubMed Scopus Google Scholar). of the upstream or downstream in exon IIIb inclusion Garcia-Blanco M.A. Mol. Cell. Biol. 20: PubMed Scopus Google Scholar, Garcia-Blanco M.A. Mol. Cell. 2002; Full Text Full Text PDF PubMed Scopus Google Scholar). activation of exon IIIb is to the silencing of exon. Exon IIIb is and IIIc is and the splicing and A. R. Mol. Cell. Biol. 1997; PubMed Scopus Google Scholar, Garcia-Blanco M.A. Mol. Cell. Biol. PubMed Scopus Google Scholar, Garcia-Blanco M.A. 2003; PubMed Scopus Google Scholar). of or in the of exon IIIb silencing, in that exon A. R. Mol. Cell. Biol. 1997; PubMed Scopus Google Scholar, Garcia-Blanco M.A. Mol. Cell. Biol. PubMed Scopus Google Scholar). examine the between transcription and splicing in the silencing of FGFR2 exon The MAZ4 element and to in at different positions within the FGFR2 minigene construct. the no effect on splicing, the MAZ4 insertions upstream of the exon IIIb silencing elements and within the downstream silencing in an in exon IIIb MAZ4 insertions downstream of the silencing elements no effect on exon IIIb the effect on exon IIIb inclusion was not cell the effect was in the into with minigene transcribed in vitro and into was no effect on splicing. of splicing was that FGFR2 alternative splicing requires co-transcriptional splicing. The MAZ4 effect was of RNAPII pause that the effect be to pause FGFR2 minigene splicing and the MAZ4 effect be changing the promoter the that transcription is to the regulation of FGFR2 alternative splicing. a for the MAZ4 effect on alternative in the MAZ4 element alters splicing decisions a change in the transcription elongation FGFR2 minigene and have been and Garcia-Blanco M.A. Mol. Cell. Biol. PubMed Scopus Google Scholar). into the constructs at positions upstream of exon IIIb downstream of exon IIIb downstream of exon IIIb and downstream of exon IIIb into site. The MAZ4 and pause site and PubMed Scopus Google Scholar). The pause element was S. Mol. Cell. Biol. 2002; PubMed Scopus Google Scholar). The pause site was A. J. D. 1997; PubMed Scopus Google Scholar). for assay was the an site in the exon to an site in the into and into and insertion constructs and and and the promoter of with the S. Garcia-Blanco M.A. PubMed Scopus Google Scholar). of are and in with at and with the 2 and in a the of was with of in of and at of was and the was to the for and was with growth was to for cell or RNA was of RNA was the To in in a in and for on in the for 2 at R. in and Scholar). in and RNA was with and with RNA polymerase and of RNA and in of and to for and at in and for at and RNA was with of and S. 2001; PubMed Scopus Google Scholar). on with a and The of a the for the that the of in that The of was the of a the of the and are in with RNA RNA vitro transcribed minigene with polymerase FGFR2 minigene with that the site is 5′ to the not of and of R. in and Scholar). 2 at of and for RNA was and in a was polymerase the of RNA was into at of the or at 2 with the PubMed Scopus Google Scholar, Proc. Natl. Acad. Sci. U. S. A. PubMed Scopus Google Scholar). of RNA with 2 of and in a of 2 The RNA was to and was for growth RNA was RT-PCR RT-PCR assay to examine elongation of RNAPII has been Garcia-Blanco M.A. Mol. Cell. Biol. 1999; PubMed Scopus Google Scholar). transcription was to and site. of was to 2 of RNA cell in a of to for and at for of of and of was and at for and 2 of and to a and of of with an of and on and a and of to and to the of the in or is the of the RT-PCR the or the for transcription of constructs and for transcription of constructs and and of for the regulation of alternative splicing the rate and of both constitutive splicing and co-transcriptional splicing to the of the splicing is the rate of of splicing factors relative to the rate of transcript be the of transcription factors and splicing factors may influence alternative splicing To the of co-transcriptional splicing on the regulation of FGFR2 alternative splicing, that transcription elongation affect the alternative splicing of these the putative RNAPII pause site MAZ4 was inserted at positions within the FGFR2 minigene Garcia-Blanco M.A. Mol. Cell. Biol. PubMed Scopus Google Scholar). The FGFR2 exons IIIb and IIIc within the of an the This minigene the splicing of the gene in both and Garcia-Blanco M.A. Mol. Cell. Biol. PubMed Scopus Google Scholar). The MAZ4 element in these of the element within a of PubMed Scopus Google Scholar). To for effects and the inserted at the the MAZ4 The of in the insertions has been demonstrated to have no effect on the splicing of G. EMBO J. PubMed Scopus Google Scholar). The MAZ4 element was inserted upstream of An RNAPII pause at the exon and elements have been be no effect on the relative of of the exon and elements is no in the rate of elongation the MAZ4 the insertion was not to alter splicing The was to the of the and was similar to constructs PubMed Scopus Google Scholar). MAZ4 insertion the of downstream silencing elements, the insertion be to exon IIIb silencing. The the of the of the and the downstream silencing element This insertion was to have of an effect than the of has effect on exon IIIb silencing, and M. A. however, the of a to critical elements not the MAZ4 downstream of was not to have effect on silencing of exon IIIb since the RNAPII pause site was downstream of for silencing. the between Exon IIIb and the was into RNA was and the spliced transcripts assay The was the of the of spliced RNA with for of the and and The not be since not to the levels of and these levels not change with of the not the FGFR2 minigene no the of splicing 2 and and than of the of the and than of the Insertion of a was for the no effect on the splicing at of the and and The insertion of the or controls into of the no effect on the of splicing and and and This result was for the and since not alter the of the exon relative to and than downstream of the these elements to exon IIIb This result is different that in the the of insertion between the exon and downstream element an effect on exon inclusion PubMed Scopus Google Scholar). MAZ4 of Exon to the insertion of insertion of the MAZ4 element at silencing of exon in silencing of exon IIIb in an of the the MAZ4 insertion downstream of no effect on the splicing MAZ4 at and in a or in the with the and The in the was a in the was no change in the RNA levels for these insertion of the MAZ4 element upstream of in a in the and and This result a effect on RNAPII that influences downstream splicing The MAZ4 exon IIIb both activation of exon IIIb and silencing of exon the MAZ4 no effect on activation of exon IIIb or silencing of exon IIIc not This result the that the effect in was to a cell type-specific To insertion constructs into a that a of Garcia-Blanco M.A. Mol. Cell. Biol. PubMed Scopus Google Scholar). the of exon IIIb is in Garcia-Blanco M.A. Mol. Cell. Biol. PubMed Scopus Google Scholar). The insertion constructs into and the spliced transcripts with in the in was however, a higher level of exon IIIb for construct. and of was no change in the with the or the at site and the level of for the was higher was than that in was a in the MAZ4 element was inserted upstream of between IIIb and and between and and in was no effect was downstream of similar effect on exon IIIb silencing was with the constructs in a cell that exon IIIb not the MAZ4 effect on exon IIIb silencing was not cell the MAZ4 MAZ4 effect have RNAPII RNAPII a in elongation or a change in the of the RNAPII transcription complex a MAZ4 site. To examine the MAZ4 to or a RT-PCR assay was to the relative level of transcripts Garcia-Blanco M.A. Mol. Cell. Biol. 1999; PubMed Scopus Google Scholar). RNA was cell constructs with MAZ4 or at the and transcription was with to the and site transcription with a of upstream of for the transcription the relative levels of transcripts and site are the in The a are in both transcripts the MAZ4 than the The in the was than the effect on level of in in transcripts the MAZ4 than the at the insertion site. of transcripts MAZ4 than at the site. FGFR2 on was that the effect of the MAZ4 element insertion was not to in but a cis-acting effect of on the The MAZ4 with a site M. J. Biol. Chem. 2001; Full Text Full Text PDF PubMed Scopus Google Scholar). has been to a in the of alternative splicing of to the RNA R. 1999; PubMed Scopus Google Scholar, Mol. Cell. Biol. 1999; PubMed Scopus Google Scholar, J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar, 2001; PubMed Scopus Google Scholar, S. A. J. Biol. Chem. 2001; Full Text Full Text PDF PubMed Scopus (110) Google Scholar, M. EMBO J. 2002; PubMed Scopus Google Scholar). have that in vitro transcribed RNA be into RNA into cell be spliced and M. A. EMBO J. PubMed Scopus Google Scholar, T. Cell. Full Text PDF PubMed Scopus Google Scholar). To the MAZ4 effect a cis-acting effect on the RNA of a minigene transcribed in vitro with polymerase and into The a but not and not the The splicing of these was was no in the splicing of and MAZ4 at the and and of the RNA no effect that result was not to of the with RNA and of the of a to the RNA no effect on the splicing not that the MAZ4 effect was co-transcriptional splicing and was not a cis-acting effect on the an FGFR2 splicing similar to that of both transcription and splicing in Garcia-Blanco M.A. Mol. Cell. 2002; Full Text Full Text PDF PubMed Scopus Google Scholar). the of splicing was in vitro transcribed and was of and that the of alternative splicing was transcription and splicing. The of splicing regulation was in not result was not to silencing of exon the of co-transcriptional splicing in the of alternative splicing of the FGFR2 RNA Exon IIIb the MAZ4 effect was a effect of RNAPII and not a effect of the MAZ4 RNAPII pause into the FGFR2 of the pause downstream of a the pause site was the downstream of the gene and the pause site was the downstream of the gene S. Mol. Cell. Biol. 2002; PubMed Scopus Google Scholar, P. D. EMBO J. PubMed Scopus Google Scholar). of these the of an upstream site in a and have been to induce RNA polymerase R. P. EMBO J. PubMed Scopus (56) Google Scholar, S. Mol. Cell. Biol. 2002; PubMed Scopus Google Scholar, P. D. EMBO J. PubMed Scopus Google Scholar). The pause site was the of the gene D. D. J. Mol. Biol. PubMed Scopus (56) Google Scholar). This pause site RNAPII in in vitro transcription and in A. J. D. 1997; PubMed Scopus Google Scholar, D. D. J. Mol. Biol. PubMed Scopus (56) Google Scholar). of these RNAPII pause to the MAZ4 for these pause into the FGFR2 cell and splicing was no in FGFR2 splicing for the and pause relative to controls at insertion site and the pause site a in the inserted at the insertion site was however, no change in the splicing at the and insertion of the RNAPII pause was to change the FGFR2 splicing in a similar but not to The MAZ4 splicing decisions have been to be transcription promoter (3Rosonina E. Bakowski M.A. McCracken S. Blencowe B.J. J. Biol. Chem. 2003; 278: 43034-43040Abstract Full Text Full Text PDF PubMed Scopus (56) Google Scholar, P. Pesce C.G. Baralle F.E. Kornblihtt A.R. Proc. Natl. Acad. Sci. U. S. A. 1997; 94: 11456-11460Crossref PubMed Scopus (270) Google Scholar, 5Cramer P. Caceres J.F. Cazalla D. Kadener S. Muro A.F. Baralle F.E. Kornblihtt A.R. Mol. Cell. 1999; 4: 251-258Abstract Full Text Full Text PDF PubMed Scopus (257) Google Scholar, 6Kadener S. Cramer P. Nogues G. Cazalla D. de la Mata M. Fededa J.P. Werbajh S.E. Srebrow A. Kornblihtt A.R. EMBO J. 2001; 20: 5759-5768Crossref PubMed Scopus (110) Google Scholar, 7Nogues G. Kadener S. Cramer P. Bentley D. Kornblihtt A.R. J. Biol. Chem. 2002; 277: 43110-43114Abstract Full Text Full Text PDF PubMed Scopus (144) Google Scholar, 8Kadener S. Fededa J.P. Rosbash M. Kornblihtt A.R. Proc. Natl. Acad. Sci. U. S. A. 2002; 99: 8185-8190Crossref PubMed Scopus (97) Google Scholar). To that the MAZ4 effect on splicing was on the FGFR2 minigene and insertion constructs downstream of the the constructs the of transcripts that the constructs that of the and constructs was into cell and splicing was no change in the splicing between the constructs with and the and The constructs the of in relative to the insertions the constructs MAZ4 was at the and insertion the splicing change the transcripts the FGFR2 minigene splicing was the promoter than The relative to the in the and The in was likely to regulation of exon IIIb silencing, since was a in the the effect of MAZ4 insertions was in the with the and was an in the with MAZ4 insertion at and however, the of was and the the MAZ4 insertion at in a in relative to the insertion was than in the the in the at the insertion site than in the constructs to in the to the was no effect of the MAZ4 insertion at the site. The in the MAZ4 was inserted at relative to the insertion was the that FGFR2 alternative splicing regulation is a for the silencing of an alternatively spliced transcription different cis-acting elements for silencing or splicing of an exon the RNAPII complex and are for several factors influence silencing or splicing may The of of to elements The relative of and factor in to the relative of and factor influence with the transcription complex may influence factors The factors may be with the transcription complex and be to the RNA into of or with the transcription complex may or factor 2002; PubMed Scopus Google Scholar, A.L. Sci. Full Text PDF PubMed Scopus Google Scholar). To examine the between transcription and of alternative splicing, PubMed Scopus Google inserted the MAZ4 element between the alternatively spliced exon and a downstream PubMed Scopus Google an in exon inclusion in the of the and that effect was RNAPII at the MAZ4 for exon silencing be The have however, that to be at in the of FGFR2 We propose that in to RNAPII the MAZ4 element a effect on the transcription complex that alters downstream splicing We have that co-transcriptional splicing is for the alternative splicing of the FGFR2 Transfection of in vitro transcribed RNA in of the and in different cell and not was be a of splicing The for mutually exclusive splicing of exon IIIb or IIIc is not on co-transcriptional however, the cell type-specific inclusion of an exon requires level of The silencing of FGFR2 exon IIIb in was the putative RNAPII pause element MAZ4 at positions within the FGFR2 of the and the MAZ4 element in an in the a of exon IIIb silencing are with the for the of alternative splicing PubMed Scopus Google Scholar), the that not that RNAPII at the not alter the between splicing and silencing since the pause the exon and are the effect insertion a effect of the MAZ4 site on the RNAPII elongation This effect be a change in elongation rate of the or a change in the of the transcription vitro transcription a of transcription at the MAZ4 site M. EMBO J. PubMed Scopus Google Scholar). Additionally, RT-PCR assay to examine RNAPII elongation in that is a in the of the MAZ4 site This result be in the MAZ4 site be RNAPII RNAPII or the RNAPII elongation We not that MAZ4 to a in exon silencing. not be in and the in transcript levels be than the in a in the elongation rate of the a in the of transcripts at the pause site be This was the RT-PCR of to the was downstream of the and the was in the between exon IIIc and exon D. was no in the of transcripts the pause site and downstream of the MAZ4 site not the RNAPII elongation rate is not likely to have been the MAZ4 site. that MAZ4 exon IIIb silencing the RNAPII elongation complex and in The splicing effect the MAZ4 insertion was between and is the of or at the MAZ4 site. be with no effect on exon IIIb in or of the polymerase that and not be to have an effect on splicing. a polymerase at MAZ4 site may with the of for of silencing. factors the and the and have been Garcia-Blanco M.A. Mol. Cell. 2002; Full Text Full Text PDF PubMed Scopus Google Scholar, M. R. Mol. Cell. Biol. 1999; PubMed Scopus Google Scholar). within and of in IIIb inclusion Garcia-Blanco M.A. Mol. Cell. 2002; Full Text Full Text PDF PubMed Scopus Google Scholar). pause of the RNAPII at the MAZ4 site the upstream silencing elements on the RNA in to the transcription The of the RNA or transcription complex may or for silencing the insertion downstream no effect on splicing. This site be downstream of the that to are An alternative for the effects of MAZ4 insertion is that the MAZ4 a splicing factor site in the the an M. J. Biol. Chem. 2001; Full Text Full Text PDF PubMed Scopus Google Scholar). has been in of alternative splicing of several is a of splicing to the site in the of splicing element in and splicing of the exon R. 1999; PubMed Scopus Google Scholar, J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar, S. A. J. Biol. Chem. 2001; Full Text Full Text PDF PubMed Scopus (110) Google Scholar). is an of splicing of the exon and of the exon in Mol. Cell. Biol. 1999; PubMed Scopus Google Scholar, M. EMBO J. 2002; PubMed Scopus Google Scholar). We not the MAZ4 effect was to of a cis-acting factor to the since splicing of in vitro transcribed RNA was not the of the MAZ4 however, does not the that a factor be the The of transcriptional is the that another pause is of a similar effect on splicing. with insertion of the pause site at and in the pause site no effect inserted at The for with the MAZ4 result is not however, may be effects of the pause on the RNAPII elongation This is the that pause and no effect on FGFR2 splicing inserted into the for for the MAZ4 effect the PubMed Scopus Google that between exon and downstream element an effect on splicing. This result is with for alternative splicing of the insertions effect on alternative splicing of the FGFR2 minigene This is with a change to the transcription than a between splicing and silencing. is that the be than downstream with no effect on silencing of exon to both upstream and downstream of exon IIIb has been to a of Garcia-Blanco M.A. Mol. Cell. Biol. 2001; PubMed Scopus Google Scholar). that of silencing be the downstream of the exon. An in the between transcription and alternative splicing is that at the promoter may influence splicing decisions downstream. in promoter elements that influence the elongation rate of the polymerase alternative splicing decisions (3Rosonina E. Bakowski M.A. McCracken S. Blencowe B.J. J. Biol. Chem. 2003; 278: 43034-43040Abstract Full Text Full Text PDF PubMed Scopus (56) Google Scholar, P. Pesce C.G. Baralle F.E. Kornblihtt A.R. Proc. Natl. Acad. Sci. U. S. A. 1997; 94: 11456-11460Crossref PubMed Scopus (270) Google Scholar, 6Kadener S. Cramer P. Nogues G. Cazalla D. de la Mata M. Fededa J.P. Werbajh S.E. Srebrow A. Kornblihtt A.R. EMBO J. 2001; 20: 5759-5768Crossref PubMed Scopus (110) Google Scholar, 7Nogues G. Kadener S. Cramer P. Bentley D. Kornblihtt A.R. J. Biol. Chem. 2002; 277: 43110-43114Abstract Full Text Full Text PDF PubMed Scopus (144) Google Scholar, 8Kadener S. Fededa J.P. Rosbash M. Kornblihtt A.R. Proc. Natl. Acad. Sci. U. S. A. 2002; 99: 8185-8190Crossref PubMed Scopus (97) Google Scholar). different splicing factors have been with both transcription factors and with transcriptional that splicing be the that are with the transcription complex at the of transcript initiation 2002; PubMed Scopus Google Scholar). in the a of exons are alternatively spliced to downstream The choice of exon is alternative promoter exon has promoter and the promoter exon is spliced to the exons Cramer P. R. T. Mol. Cell. 2002; Full Text Full Text PDF PubMed Scopus Google Scholar). Similar promoter-dependent may influence FGFR2 alternative splicing. changing the promoter the promoter to the promoter the splicing of both the minigene and the MAZ4 insertion constructs are with a in the MAZ4 element the transcription elongation complex in a to exon IIIb silencing. This that a factor with the elongation complex is for FGFR2 alternative splicing. We and for and for critical of the and of the Garcia-Blanco and for