Determinants of TRPV4 activity following selective activation by small molecule agonist GSK1016790A.
Determinants of TRPV4 activity following selective activation by small molecule agonist GSK1016790A.
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DOI：
10.1371/journal.pone.0016713
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发表时间：
2011-02-14
期刊：
影响因子：
3.7
通讯作者：
O'Neil RG
中科院分区：
文献类型：
--
作者：
Jin M;Wu Z;Chen L;Jaimes J;Collins D;Walters ET;O'Neil RG
TRPV 4（Transient Receptor Potential Vanilloid 4）通道可被多种刺激激活，包括低渗应激、非伤害性热应激、机械应激和一些小分子激动剂（如佛波酯4α-PDD）。GSK 1016790 A（GSK 101）是最近发现的TRPV 4特异性小分子激动剂。在瞬时转染TRPV 4的HeLa细胞（HeLa-TRPV 4）中评价其对TRPV 4活性的物理决定因素的影响。GSK 101（10 nM）在HeLa-TRPV 4细胞中引起TRPV 4特异性Ca 2+内流，但在对照转染细胞中不引起，其可以在活化的早期阶段而不是晚期阶段被钌红和无Ca 2+培养基更显著地抑制，反映了明显的部分脱敏。Western blot分析表明，GSK 101活化并不诱导质膜上TRPV 4表达的增加，但引起HeLa TRPV 4细胞中质膜上TRPV 4的立即和持续下调。膜片钳分析也显示了早期的部分脱敏的通道，这是Ca 2 +-独立的。TRPV 4亚基组装的FRET分析表明，GSK 101诱导的TRPV 4通道激活/脱敏不是由于质膜上同源四聚体通道形成的改变。可以得出结论，GSK 101特异性激活TRPV 4通道，导致质膜上通道表达的快速部分脱敏和下调。TRPV 4亚基组装似乎发生在从ER/高尔基体到质膜的运输过程中，并且不被激动剂刺激改变。
TRPV4 (Transient Receptor Potential Vanilloid 4) channels are activated by a wide range of stimuli, including hypotonic stress, non-noxious heat and mechanical stress and some small molecule agonists (e.g. phorbol ester 4α-PDD). GSK1016790A (GSK101) is a recently discovered specific small molecule agonist of TRPV4. Its effects on physical determinants of TRPV4 activity were evaluated in HeLa cells transiently transfected with TRPV4 (HeLa-TRPV4). GSK101 (10 nM) causes a TRPV4 specific Ca2+ influx in HeLa-TRPV4 cells, but not in control transfected cells, which can be inhibited by ruthenium red and Ca2+-free medium more significantly at the early stage of the activation rather than the late stage, reflecting apparent partial desensitization. Western blot analysis showed that GSK101 activation did not induce an increase in TRPV4 expression at the plasma membrane, but caused an immediate and sustained downregulation of TRPV4 on the plasma membrane in HeLa-TRPV4 cells. Patch clamp analysis also revealed an early partial desensitization of the channel which was Ca2+-independent. FRET analysis of TRPV4 subunit assembly demonstrated that the GSK101-induced TRPV4 channel activation/desensitization was not due to alterations in homotetrameric channel formation on the plasma membrane. It is concluded that GSK101 specifically activates TRPV4 channels, leading to a rapid partial desensitization and downregulation of the channel expression on the plasma membrane. TRPV4 subunit assembly appears to occur during trafficking from the ER/Golgi to the plasma membrane and is not altered by agonist stimulation.
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发表时间：
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影响因子：
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作者：
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