Domain assembly of NAADP-gated two-pore channels.
Domain assembly of NAADP-gated two-pore channels.
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DOI：
10.1042/bj20111617
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发表时间：
2012-01-01
期刊：
影响因子：
--
通讯作者：
Patel S
中科院分区：
文献类型：
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作者：
Churamani D;Hooper R;Brailoiu E;Patel S
TPC（双孔通道）最近已被确定为Ca 2+动员信使NAADP（烟酸腺嘌呤二核苷酸磷酸）的靶点。TPC具有独特的结构，由胞质末端、两个疏水结构域（I和II）（每个疏水结构域包含六个跨膜区和一个孔）和连接胞质环组成;然而，关于这些通道如何组装知之甚少。在本论文中，我们报告说，域I和II的人TPC能够独立插入膜，而连接域的环未能插入。TPC 1的结构域I内的跨膜区对也能够插入，这与疏水区的顺序翻译整合一致。然而，前两个跨膜区的插入是低效的，这表明在翻译过程中跨膜区之间可能存在相互作用。这两个结构域，和每对跨膜区域内的结构域I，能够形成寡聚体，突出了显着的冗余驱动寡聚体形成的分子决定簇。每个疏水结构域在交联时形成二聚体。TPC 1的前四个跨膜区也形成二聚体，而跨膜区5和6，包括孔环，形成二聚体和四聚体。因此，TPC可能通过跨膜区域之间的差异相互作用组装成二聚体。本研究为TPC的膜插入和寡聚化提供了新的分子视角。
TPCs (two-pore channels) have recently been identified as targets for the Ca2+-mobilizing messenger NAADP (nicotinic acid–adenine dinucleotide phosphate). TPCs have a unique structure consisting of cytosolic termini, two hydrophobic domains (I and II) each comprising six transmembrane regions and a pore, and a connecting cytosolic loop; however, little is known concerning how these channels are assembled. In the present paper, we report that both domain I and II of human TPCs are capable of independent insertion into membranes, whereas the loop linking the domains fails to insert. Pairs of transmembrane regions within domain I of TPC1 are also capable of insertion, consistent with sequential translational integration of hydrophobic regions. Insertion of the first two transmembrane regions, however, was inefficient, indicating possible interaction between transmembrane regions during translation. Both domains, and each pair of transmembrane regions within domain I, were capable of forming oligomers, highlighting marked redundancy in the molecular determinants driving oligomer formation. Each hydrophobic domain formed dimers upon cross-linking. The first four transmembrane regions of TPC1 also formed dimers, whereas transmembrane regions 5 and 6, encompassing the pore loop, formed both dimers and tetramers. TPCs thus probably assemble as dimers through differential interactions between transmembrane regions. The present study provides new molecular insight into the membrane insertion and oligomerization of TPCs.