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Exon repression by polypyrimidine tract binding protein.
RNA. 2005 May; 11(5):699-716.RNA

Abstract

Polypyrimidine tract binding protein (PTB) is known to silence the splicing of many alternative exons. However, exons repressed by PTB are affected by other RNA regulatory elements and proteins. This makes it difficult to dissect the structure of the pre-mRNP complexes that silence splicing, and to understand the role of PTB in this process. We determined the minimal requirements for PTB-mediated splicing repression. We find that the minimal sequence for high affinity binding by PTB is relatively large, containing multiple polypyrimidine elements. Analytical ultracentrifugation and proteolysis mapping of RNA cross-links on the PTB protein indicate that most PTB exists as a monomer, and that a polypyrimidine element extends across multiple PTB domains. The high affinity site is bound initially by a PTB monomer and at higher concentrations by additional PTB molecules. Significantly, this site is not sufficient for splicing repression when placed in the 3' splice site of a strong test exon. Efficient repression requires a second binding site within the exon itself or downstream from it. This second site enhances formation of a multimeric PTB complex, even if it does not bind well to PTB on its own. These experiments show that PTB can be sufficient to repress splicing of an otherwise constitutive exon, without binding sites for additional regulatory proteins and without competing with U2AF binding. The minimal complex mediating splicing repression by PTB requires two binding sites bound by an oligomeric PTB complex.

Authors+Show Affiliations

Department of Microbiology, Immunology & Molecular Genetics, University of California Los Angeles, Los Angeles, CA 90095-1662, USA.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

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

Language

eng

PubMed ID

15840818

Citation

Amir-Ahmady, Batoul, et al. "Exon Repression By Polypyrimidine Tract Binding Protein." RNA (New York, N.Y.), vol. 11, no. 5, 2005, pp. 699-716.
Amir-Ahmady B, Boutz PL, Markovtsov V, et al. Exon repression by polypyrimidine tract binding protein. RNA. 2005;11(5):699-716.
Amir-Ahmady, B., Boutz, P. L., Markovtsov, V., Phillips, M. L., & Black, D. L. (2005). Exon repression by polypyrimidine tract binding protein. RNA (New York, N.Y.), 11(5), 699-716.
Amir-Ahmady B, et al. Exon Repression By Polypyrimidine Tract Binding Protein. RNA. 2005;11(5):699-716. PubMed PMID: 15840818.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Exon repression by polypyrimidine tract binding protein. AU - Amir-Ahmady,Batoul, AU - Boutz,Paul L, AU - Markovtsov,Vadim, AU - Phillips,Martin L, AU - Black,Douglas L, PY - 2005/4/21/pubmed PY - 2005/5/25/medline PY - 2005/4/21/entrez SP - 699 EP - 716 JF - RNA (New York, N.Y.) JO - RNA VL - 11 IS - 5 N2 - Polypyrimidine tract binding protein (PTB) is known to silence the splicing of many alternative exons. However, exons repressed by PTB are affected by other RNA regulatory elements and proteins. This makes it difficult to dissect the structure of the pre-mRNP complexes that silence splicing, and to understand the role of PTB in this process. We determined the minimal requirements for PTB-mediated splicing repression. We find that the minimal sequence for high affinity binding by PTB is relatively large, containing multiple polypyrimidine elements. Analytical ultracentrifugation and proteolysis mapping of RNA cross-links on the PTB protein indicate that most PTB exists as a monomer, and that a polypyrimidine element extends across multiple PTB domains. The high affinity site is bound initially by a PTB monomer and at higher concentrations by additional PTB molecules. Significantly, this site is not sufficient for splicing repression when placed in the 3' splice site of a strong test exon. Efficient repression requires a second binding site within the exon itself or downstream from it. This second site enhances formation of a multimeric PTB complex, even if it does not bind well to PTB on its own. These experiments show that PTB can be sufficient to repress splicing of an otherwise constitutive exon, without binding sites for additional regulatory proteins and without competing with U2AF binding. The minimal complex mediating splicing repression by PTB requires two binding sites bound by an oligomeric PTB complex. SN - 1355-8382 UR - https://www.unboundmedicine.com/medline/citation/15840818/Exon_repression_by_polypyrimidine_tract_binding_protein_ DB - PRIME DP - Unbound Medicine ER -