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 journalArticlepeer-review

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

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