SecA is required for membrane targeting of the cell division protein DivIVA in vivo
SecA is required for membrane targeting of the cell division protein DivIVA in vivo
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DOI：
10.3389/fmicb.2014.00058
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发表时间：
2014-02-14
影响因子：
5.2
通讯作者：
Hamoen, Leendert W.
中科院分区：
文献类型：
--
作者：
Halbedel, Sven;Kawai, Maki;Hamoen, Leendert W.
保守蛋白DivIVA参与革兰氏阳性菌的不同形态发生过程。在枯草芽孢杆菌中，蛋白质定位于细胞分裂位点和细胞极，并作为调节分裂位点选择的蛋白质和孢子形成所需的蛋白质的支架。为了鉴定与DivIVA结合的其他蛋白质，我们进行了体内交联实验。出现的一个可能的候选者是分泌马达ATP酶SecA。SecA突变体已经被描述为抑制孢子形成，并且由于DivIVA是孢子形成所必需的，我们检查了DivIVA在这些突变体中的定位。令人惊讶的是，DivIVA是离域的，这表明SecA是DivIVA靶向所必需的。为了进一步证实这一点，我们进行了SecA耗竭和抑制实验，这进一步表明DivIVA定位依赖于SecA。细胞分级分离实验表明SecA对于DivIVA与细胞膜的结合是重要的。这是出乎意料的，因为DivIVA不含信号序列，并且能够在体外结合人工脂质膜而无需其他蛋白质的支持。SecA是蛋白分泌和膜插入所必需的，因此其在DivIVA定位中的作用可能是间接的。SecA在DivIVA折叠和/或靶向可能的替代作用进行了讨论。
The conserved protein DivIVA is involved in different morphogenetic processes in Gram-positive bacteria. In Bacillus subtilis, the protein localizes to the cell division site and cell poles, and functions as a scaffold for proteins that regulate division site selection, and for proteins that are required for sporulation. To identify other proteins that bind to DivIVA, we performed an in vivo cross-linking experiment. A possible candidate that emerged was the secretion motor ATPase SecA. SecA mutants have been described that inhibit sporulation, and since DivIVA is necessary for sporulation, we examined the localization of DivIVA in these mutants. Surprisingly, DivIVA was delocalized, suggesting that SecA is required for DivIVA targeting. To further corroborate this, we performed SecA depletion and inhibition experiments, which provided further indications that DivIVA localization depends on SecA. Cell fractionation experiments showed that SecA is important for binding of DivIVA to the cell membrane. This was unexpected since DivIVA does not contain a signal sequence, and is able to bind to artificial lipid membranes in vitro without support of other proteins. SecA is required for protein secretion and membrane insertion, and therefore its role in DivIVA localization is likely indirect. Possible alternative roles of SecA in DivIVA folding and/or targeting are discussed.