Muscle-specific F-Box only proteins facilitate BK channel β1 subunit downregulation in vascular smooth muscle cells of diabetes mellitus

Dai Min Zhang, Tongrong He, Zvonimir S. Katusic, Hon Chi Lee, Tong Lu

Research output: Contribution to journalArticlepeer-review

39 Scopus citations

Abstract

Rationale: Activity of the large conductance Ca-activated K (BK) channels is profoundly modulated by its β1 subunit (BK-β1). However, BK-β1 expression is downregulated in diabetic vessels. The ubiquitin-proteasome system (UPS) is a major mechanism of intracellular protein degradation. Whether UPS participates in BK-β1 downregulation in diabetic vessels is unknown. Objective: We hypothesize that UPS facilitates vascular BK-β1 degradation in diabetes. Methods and results: Using patch clamp and molecular biological approaches, we found that BK-β1-mediated channel activation and BK-β1 protein expression were reduced in aortas of streptozotocin-induced diabetic rats and in human coronary arterial smooth muscle cells (CASMCs) cultured in high glucose. This was accompanied by upregulation of F-box only protein (FBXO)-9 and FBXO-32 (atrogin-1), the key components of the Skp1-Cullin-F-box (SCF) type ubiquitin ligase complex. BK-β1 expression was suppressed by the FBXO activator doxorubicin but enhanced by FBXO-9 small interfering RNA or by the proteasome inhibitor MG-132. Cotransfection of atrogin-1 in HEK293 cells significantly reduced Flag-hSlo-β1 expression by 2.16-fold, compared with expression of Flag-hSlo-β1V146A (a mutant without the PDZ-binding motif). After cotransfection with atrogin-1, the ubiquitination of Flag-hSlo- β1 was increased by 1.91-fold, compared with that of hSlo-β1V146A, whereas cotransfection with atrogin-1ΔF (a nonfunctional mutant without the F-box motif) had no effect. Moreover, inhibition of Akt signaling attenuated the phosphorylation of forkhead box O transcription factor (FOXO)-3a and enhanced atrogin-1 expression, which in turn suppressed BK-β1 protein levels in human CASMCs. Conclusions: Downregulation of vascular BK-β1 expression in diabetes and in high-glucose culture conditions was associated with FOXO-3a/FBXO-dependent increase in BK-β1 degradation.

Original languageEnglish (US)
Pages (from-to)1454-1459
Number of pages6
JournalCirculation research
Volume107
Issue number12
DOIs
StatePublished - Dec 10 2010

Keywords

  • BK channelβ subunit
  • diabetes mellitus
  • protein degradation
  • ubiquitin-proteasome system

ASJC Scopus subject areas

  • Physiology
  • Cardiology and Cardiovascular Medicine

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