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Role of an inhibitory pyrimidine element and polypyrimidine tract binding protein in repression of a regulated alpha-tropomyosin exon.
RNA. 1998 Jan; 4(1):85-100.RNA

Abstract

Splicing of exons 2 and 3 of a-tropomyosin (TM) involves mutually exclusive selection of either exon 3, which occurs in most cells, or of exon 2 in smooth muscle (SM) cells. The SM-specific selection of exon 2 results from the inhibition of exon 3. At least two essential cis-acting elements are required for exon 3 inhibition, the upstream and downstream regulatory elements (URE and DRE). These elements are essential for repression of TM exon 3 in SM cells, and also mediate a low level of repression of exon 3 in an in vitro 5' splice site competition assay in HeLa extracts. Here, we show that the DRE consists of at least two discrete components, a short region containing a number of UGC motifs, and an essential pyrimidine-rich tract (DY). We show that the specific sequence of the DY element is important and that DY is able to bind to factors in HeLa nuclear extracts that mediate a low background level of exon 3 skipping. Deletion of a sequence within DY identified as an optimal binding site for PTB impairs (1) regulation of splicing in vivo, (2) skipping of exon 3 in an in vitro 5' splice site competition, (3) the ability of DY competitors to affect the 5' splice site competition in vitro, and (4) binding of PTB to DY. Addition of recombinant PTB to in vitro splicing reactions is able to partially reverse the effects of the DY competitor RNA. The data are consistent with a model for regulation of TM splicing that involves the participation of both tissue-specific and general inhibitory factors and in which PTB plays a role in repressing both splice sites of exon 3.

Authors+Show Affiliations

Department of Biochemistry, University of Cambridge, United Kingdom.No affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

9436911

Citation

Gooding, C, et al. "Role of an Inhibitory Pyrimidine Element and Polypyrimidine Tract Binding Protein in Repression of a Regulated Alpha-tropomyosin Exon." RNA (New York, N.Y.), vol. 4, no. 1, 1998, pp. 85-100.
Gooding C, Roberts GC, Smith CW. Role of an inhibitory pyrimidine element and polypyrimidine tract binding protein in repression of a regulated alpha-tropomyosin exon. RNA. 1998;4(1):85-100.
Gooding, C., Roberts, G. C., & Smith, C. W. (1998). Role of an inhibitory pyrimidine element and polypyrimidine tract binding protein in repression of a regulated alpha-tropomyosin exon. RNA (New York, N.Y.), 4(1), 85-100.
Gooding C, Roberts GC, Smith CW. Role of an Inhibitory Pyrimidine Element and Polypyrimidine Tract Binding Protein in Repression of a Regulated Alpha-tropomyosin Exon. RNA. 1998;4(1):85-100. PubMed PMID: 9436911.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Role of an inhibitory pyrimidine element and polypyrimidine tract binding protein in repression of a regulated alpha-tropomyosin exon. AU - Gooding,C, AU - Roberts,G C, AU - Smith,C W, PY - 1998/1/22/pubmed PY - 1998/1/22/medline PY - 1998/1/22/entrez SP - 85 EP - 100 JF - RNA (New York, N.Y.) JO - RNA VL - 4 IS - 1 N2 - Splicing of exons 2 and 3 of a-tropomyosin (TM) involves mutually exclusive selection of either exon 3, which occurs in most cells, or of exon 2 in smooth muscle (SM) cells. The SM-specific selection of exon 2 results from the inhibition of exon 3. At least two essential cis-acting elements are required for exon 3 inhibition, the upstream and downstream regulatory elements (URE and DRE). These elements are essential for repression of TM exon 3 in SM cells, and also mediate a low level of repression of exon 3 in an in vitro 5' splice site competition assay in HeLa extracts. Here, we show that the DRE consists of at least two discrete components, a short region containing a number of UGC motifs, and an essential pyrimidine-rich tract (DY). We show that the specific sequence of the DY element is important and that DY is able to bind to factors in HeLa nuclear extracts that mediate a low background level of exon 3 skipping. Deletion of a sequence within DY identified as an optimal binding site for PTB impairs (1) regulation of splicing in vivo, (2) skipping of exon 3 in an in vitro 5' splice site competition, (3) the ability of DY competitors to affect the 5' splice site competition in vitro, and (4) binding of PTB to DY. Addition of recombinant PTB to in vitro splicing reactions is able to partially reverse the effects of the DY competitor RNA. The data are consistent with a model for regulation of TM splicing that involves the participation of both tissue-specific and general inhibitory factors and in which PTB plays a role in repressing both splice sites of exon 3. SN - 1355-8382 UR - https://www.unboundmedicine.com/medline/citation/9436911/Role_of_an_inhibitory_pyrimidine_element_and_polypyrimidine_tract_binding_protein_in_repression_of_a_regulated_alpha_tropomyosin_exon_ L2 - http://www.rnajournal.org/cgi/pmidlookup?view=long&pmid=9436911 DB - PRIME DP - Unbound Medicine ER -