Exonic splicing enhancer-dependent splicing of the gonadotropin-releasing hormone premessenger ribonucleic acid is mediated by Tra2α, a 40-kilodalton serine/arginine-rich protein

Jae Young Seong, Jin Han, Sungjin Park, Wolfgang Wuttke, Hubertus Jarry, Kyungjin Kim

Research output: Contribution to journalArticle

18 Citations (Scopus)

Abstract

In an earlier study, we found that excision of the first intron (intron A) from the rat GnRH primary transcript is attenuated in non-GnRH-producing cells. This attenuation can be partially relieved by exonic splicing enhancers (ESEs) located in GnRH exons 3 and 4. In the present study, we confirmed that intron A of the mouse GnRH pre-mRNA was not excised in a HeLa nuclear extract (NE) in vitro or in COS-7 cells in vivo. Intron A could, however, be partially removed when exon 3 and/or 4 were linked to exon 2. In the presence of an ESE in exon 4 (ESE4), an addition of GT1 NE further increased the excision rate of intron A, whereas the addition of KK1 (a non-GnRH-producing cell) NE decreased it. To define the GnRH neuron-specific splicing activity, GT1 NE was fractionated by ultracentrifugation and ammonium sulfate precipitation. A 50-90% ammonium sulfate pellet (ASP50-90) fraction was further precipitated with 20 mM MgCl2 to isolate a serine/arginine-rich (SR) protein fraction. Among the ASP fractions, ASP40-50 significantly increased the excision rate of intron A in the presence of HeLa NE or SR protein-rich fraction. However, the ASP40-50 fraction alone could not remove intron A. This result suggests the presence of a cofactor protein(s) in the ASP40-50 fraction that may mediate the interaction between a 3′ spliceosome complex and the ESE4-SR protein complex. UV cross-linking and gel mobility shift analysis revealed that Tra2α but not other SR proteins tested, specifically binds to ESE4. Moreover, Tra2α stimulated intron A excision in a dose-dependent manner. These results imply that Tra2α and a cofactor protein in the ASP40-50 fraction are involved in mediating the GnRH neuron-specific excision of intron A from the GnRH primary transcript.

Original languageEnglish
Pages (from-to)2426-2438
Number of pages13
JournalMolecular Endocrinology
Volume16
Issue number11
DOIs
Publication statusPublished - 2002 Nov 1
Externally publishedYes

ASJC Scopus subject areas

  • Molecular Biology
  • Endocrinology, Diabetes and Metabolism

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