Molecular identification and functional characterization of a mitochondrial sulfonylurea receptor 2 splice variant generated by intraexonic splicing

Bin Ye, Stacie L. Kroboth, Jie Lin Pu, Jason J. Sims, Nitin T. Aggarwal, Elizabeth M. McNally, Jonathan C. Makielski, Nian Qing Shi*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

46 Scopus citations

Abstract

RATIONALE:: Cardioprotective pathways may involve a mitochondrial ATP-sensitive potassium (mitoKATP) channel but its composition is not fully understood. OBJECTIVE:: We hypothesized that the mitoKATP channel contains a sulfonylurea receptor (SUR)2 regulatory subunit and aimed to identify the molecular structure. METHODS AND RESULTS:: Western blot analysis in cardiac mitochondria detected a 55-kDa mitochondrial SUR2 (mitoSUR2) short form, 2 additional short forms (28 and 68 kDa), and a 130-kDa long form. RACE (Rapid Amplification of cDNA Ends) identified a 1.5-Kb transcript, which was generated by a nonconventional intraexonic splicing (IES) event within the 4th and 29th exons of the SUR2 mRNA. The translated product matched the predicted size of the 55-kDa short form. In a knockout mouse (SUR2KO), in which the SUR2 gene was disrupted, the 130-kDa mitoSUR2 was absent, but the short forms remained expressed. Diazoxide failed to induce increased fluorescence of flavoprotein oxidation in SUR2KO cells, indicating that the diazoxide-sensitive mitoKATP channel activity was associated with 130-kDa-based channels. However, SUR2KO mice displayed similar infarct sizes to preconditioned wild type, suggesting a protective role for the remaining short form-based channels. Heterologous coexpression of the SUR2 IES variant and Kir6.2 in a K transport mutant Escherichia coli strain permitted improved cell growth under acidic pH conditions. The SUR2 IES variant was localized to mitochondria, and removal of a predicted mitochondrial targeting sequence allowed surface expression and detection of an ATP-sensitive current when coexpressed with Kir6.2. CONCLUSIONS:: We identify a novel SUR2 IES variant in cardiac mitochondria and provide evidence that the variant-based channel can form an ATP-sensitive conductance and may contribute to cardioprotection.

Original languageEnglish (US)
Pages (from-to)1083-1093
Number of pages11
JournalCirculation research
Volume105
Issue number11
DOIs
StatePublished - Nov 2009

Keywords

  • Intraexonic splicing
  • Ischemia
  • K channel
  • Mitochondria
  • SUR2

ASJC Scopus subject areas

  • Physiology
  • Cardiology and Cardiovascular Medicine

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