Interaction with 14-3-3 proteins promotes functional expression of the potassium channels TASK-1 and TASK-3
Interaction with 14-3-3 proteins promotes functional expression of the potassium channels TASK-1 and TASK-3
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DOI：
10.1113/jphysiol.2002.027052
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发表时间：
2002-11-15
影响因子：
5.5
通讯作者：
Daut, J
中科院分区：
文献类型：
--
作者：
Rajan, S;Preisig-Müller, R;Daut, J
两个孔域钾离子通道TASK-1、TASK-3和TASK-5具有5个氨基酸的保守c端基序。截断TASK-1的c端，大大降低了在爪蟾卵母细胞或HEK293细胞中异种表达后测量到的电流，并降低了gfp标记的通道蛋白的表面膜表达。双杂交分析表明，TASK-1、TASK-3和TASK-5的c端结构域与适配器蛋白14-3-3的异构体相互作用，而TASK-4不与之相互作用。在TASK-1的末端c端，一个五肽基序RRx(S/T)x被发现足以与14-3-3进行弱但显著的相互作用，而TASK-1的最后40个氨基酸则需要与14-3-3进行强结合。如果在TASK-1或TASK-3的c端缺失一个氨基酸，就会破坏14-3-3的结合，并大大降低非洲爪蟾卵母细胞中观察到的宏观电流。不能与14-3-3异构体（V411*, S410A, S410D）相互作用的TASK-1突变体也只产生非常弱的宏观电流。相比之下，与14-3-3样野生型通道相互作用的突变体TASK-1 S409A显示出正常的宏观电流。联合注射14-3-3 cRNA可使爪蟾卵母细胞的TASK-1电流增加约70%。在HEK293细胞中共转染后，TASK-1和14-3-3（但不包括TASK-1DeltaC5和14-3-3）可以共免疫沉淀。此外，TASK-1和14-3-3可以在突触膜提取物和突触后密度膜中共免疫沉淀。我们的研究结果表明，14-3-3与TASK-1或TASK-3的相互作用可能促进通道向表面膜的运输。
The two-pore-domain potassium channels TASK-1, TASK-3 and TASK-5 possess a conserved C-terminal motif of five amino acids. Truncation of the C-terminus of TASK-1 strongly reduced the currents measured after heterologous expression in Xenopus oocytes or HEK293 cells and decreased surface membrane expression of GFP-tagged channel proteins. Two-hybrid analysis showed that the C-terminal domain of TASK-1, TASK-3 and TASK-5, but not TASK-4, interacts with isoforms of the adapter protein 14-3-3. A pentapeptide motif at the extreme C-terminus of TASK-1, RRx(S/T)x, was found to be sufficient for weak but significant interaction with 14-3-3, whereas the last 40 amino acids of TASK-1 were required for strong binding. Deletion of a single amino acid at the C-terminal end of TASK-1 or TASK-3 abolished binding of 14-3-3 and strongly reduced the macroscopic currents observed in Xenopus oocytes. TASK-1 mutants that failed to interact with 14-3-3 isoforms (V411*, S410A, S410D) also produced only very weak macroscopic currents. In contrast, the mutant TASK-1 S409A, which interacts with 14-3-3-like wild-type channels, displayed normal macroscopic currents. Co-injection of 14-3-3 cRNA increased TASK-1 current in Xenopus oocytes by about 70%. After co-transfection in HEK293 cells, TASK-1 and 14-3-3 (but not TASK-1DeltaC5 and 14-3-3) could be co-immunoprecipitated. Furthermore, TASK-1 and 14-3-3 could be co-immunoprecipitated in synaptic membrane extracts and postsynaptic density membranes. Our findings suggest that interaction of 14-3-3 with TASK-1 or TASK-3 may promote the trafficking of the channels to the surface membrane.