Phosphatidylinositol 3,5-bisphosphate regulates Ca2+ transport during yeast vacuolar fusion through the Ca2+ ATPase Pmc1.,Traffic

当前位置： X-MOL 学术 › Traffic › 论文详情

Our official English website, www.x-mol.net, welcomes your feedback! (Note: you will need to create a separate account there.)

Phosphatidylinositol 3,5-bisphosphate regulates Ca2+ transport during yeast vacuolar fusion through the Ca2+ ATPase Pmc1.
Traffic ( IF 3.6 ) Pub Date : 2020-05-10 , DOI: 10.1111/tra.12736
Gregory E Miner ₁ , Katherine D Sullivan ₁ , Chi Zhang ₁ , David Rivera-Kohr ₁ , Annie Guo ₁ , Logan R Hurst ₁ , Ez C Ellis ₁ , Matthew L Starr ₁ , Brandon C Jones ₁ , Rutilio A Fratti _{1,

2}

Affiliation

The transport of Ca²⁺ across membranes precedes the fusion and fission of various lipid bilayers. Yeast vacuoles under hyperosmotic stress become fragmented through fission events that requires the release of Ca²⁺ stores through the TRP channel Yvc1. This requires the phosphorylation of phosphatidylinositol‐3‐phosphate (PI3P) by the PI3P‐5‐kinase Fab1 to produce transient PI(3,5)P₂ pools. Ca²⁺ is also released during vacuole fusion upon trans‐ SNARE complex assembly, however, its role remains unclear. The effect of PI(3,5)P₂ on Ca²⁺ flux during fusion was independent of Yvc1. Here, we show that while low levels of PI(3,5)P₂ were required for Ca²⁺ uptake into the vacuole, increased concentrations abolished Ca²⁺ efflux. This was as shown by the addition of exogenous dioctanoyl PI(3,5)P₂ or increased endogenous production of by the hyperactive fab1 ^T2250A mutant. In contrast, the lack of PI(3,5)P₂ on vacuoles from the kinase dead fab1 ^EEE mutant showed delayed and decreased Ca²⁺ uptake. The effects of PI(3,5)P₂ were linked to the Ca²⁺ pump Pmc1, as its deletion rendered vacuoles resistant to the effects of excess PI(3,5)P₂. Experiments with Verapamil inhibited Ca²⁺ uptake when added at the start of the assay, while adding it after Ca²⁺ had been taken up resulted in the rapid expulsion of Ca²⁺. Vacuoles lacking both Pmc1 and the H⁺/Ca²⁺ exchanger Vcx1 lacked the ability to take up Ca²⁺ and instead expelled it upon the addition of ATP. Together these data suggest that a balance of efflux and uptake compete during the fusion pathway and that the levels of PI(3,5)P₂ can modulate which path predominates.

中文翻译：

磷脂酰肌醇 3,5-二磷酸通过 Ca2+ ATPase Pmc1 调节酵母液泡融合过程中的 Ca2+ 转运。

Ca ²⁺跨膜转运先于各种脂质双层的融合和裂变。高渗胁迫下的酵母液泡通过裂变事件破碎，裂变事件需要通过 TRP 通道 Yvc1释放 Ca ²⁺储存。这需要 PI3P-5-激酶 Fab1 磷酸化磷脂酰肌醇 3-磷酸 (PI3P) 以产生瞬时 PI(3,5)P ₂池。Ca ²⁺在反式SNARE 复合体组装后的液泡融合过程中也会释放，但其作用尚不清楚。PI(3,5)P ₂对融合过程中Ca ²⁺通量的影响与Yvc1 无关。在这里，我们表明虽然低水平的 PI(3,5)P ₂Ca ²⁺吸收到液泡中是必需的，增加的浓度消除了Ca ²⁺流出。这通过添加外源性二辛^酰基PI(3,5)P ₂或由过度活跃的fab1 ^T2250A突变体增加的内源性产生来^证明。相比之下，激酶死亡fab1 ^EEE突变体液泡上PI(3,5)P ₂的缺乏显示出延迟和降低的 Ca ²⁺摄取。PI(3,5)P _{2 的}影响与Ca ²⁺泵Pmc1 相关，因为它的缺失使液泡对过量PI(3,5)P ₂的影响具有抗性。维拉帕米抑制钙离子的实验²⁺在测定开始时被吸收，而在 Ca ²⁺被吸收后添加导致 Ca ²⁺的快速排出。缺乏 Pmc1 和 H ⁺ /Ca ²⁺交换器 Vcx1 的液泡缺乏吸收 Ca ²⁺的能力，而是在添加 ATP 时将其排出。这些数据一起表明，在融合途径期间外排和摄取的平衡竞争，并且 PI(3,5)P _{2 的水平}可以调节哪种途径占主导地位。

更新日期：2020-06-29

点击分享查看原文

点击收藏

阅读更多本刊最新论文本刊介绍/投稿指南11