A Purine-rich Intronic Element Enhances Alternative Splicing of Thyroid Hormone Receptor mRNA

Document Type

Article

Language

eng

Format of Original

16 p.

Publication Date

2001

Publisher

Cold Spring Harbor Laboratory Press

Source Publication

RNA

Source ISSN

1355-8382

Original Item ID

doi:10.1017/S1355838201002084

Abstract

The mammalian thyroid hormone receptor gene c-erbAα gives rise to two mRNAs that code for distinct isoforms, TRα1 and TRα2, with antagonistic functions. Alternative processing of these mRNAs involves the mutually exclusive use of a TRα1-specific polyadenylation site or TRα2-specific 5' splice site. A previous investigation of TRα minigene expression defined a critical role for the TRα2 5' splice site in directing alternative processing. Mutational analysis reported here shows that purine residues within a highly conserved intronic element, SEα2, enhance splicing of TRα2 in vitro as well as in vivo. Although SEα2 is located within the intron of TRα2 mRNA, it activates splicing of a heterologous dsx pre-mRNA when located in the downstream exon. Competition with wild-type and mutant RNAs indicates that SEα2 functions by binding trans-acting factors in HeLa nuclear extract. Protein–RNA crosslinking identifies several proteins, including SF2/ASF and hnRNP H, that bind specifically to SEα2. SEα2 also includes an element resembling a 5' splice site consensus sequence that is critical for splicing enhancer activity. Mutations within this pseudo-5' splice site sequence have a dramatic effect on splicing and protein binding. Thus SEα2 and its associated factors are required for splicing of TRα2 pre-mRNA.

Comments

Published version. RNA, No. 7 (2001): 859-874. DOI. © 2001 Cold Spring Harbor Laboratory Press. Used with permission.

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