Sano Tomoki, Nagano Makoto, Shimamura Hiroki, Yamamoto Wataru, Tamada Tomoyuki, Toshima Junko Y, Toshima Jiro
Department of Biological Science and Technology, Tokyo University of Science, 6-3-1 Niijyuku, Katsushika-ku, Tokyo 125-8585, Japan.
School of Health Science, Tokyo University of Technology, 5-23-22 Nishikamata, Ota-ku, Tokyo 144-8535, Japan.
J Biol Chem. 2025 Jun 21:110410. doi: 10.1016/j.jbc.2025.110410.
Growing evidence suggests that counter-transport of phosphatidylinositol-4-phosphate (PtdIns(4)P) and phosphatidylserine (PS) at endoplasmic reticulum (ER)-plasma membrane (PM) contact sites is required for intracellular vesicle transport. PtdIns(4)P is metabolized by Stt4 PI 4-kinase residing at the PM and by Sac1 PtdIns(4)P phosphatase at the ER, and ER-PM contact sites are believed to be important for its efficient turnover. Recently, Stt4 has been shown to extensively localize to ER-PM contact sites. However, the precise location of Stt4 and the mechanism of localization to these sites have not been clarified. Additionally, although several studies have suggested a requirement for PS/PtdIns(4)P and sterol/PtdIns(4)P exchange at ER-PM contact sites in endocytosis, it is still unclear whether contact between the ER and PM, turnover of PtdIns(4)P or PS, or maintenance of PtdIns(4)P or PS levels is more important. Here we found that Stt4 localizes to the cER regions where Scs2 and Ist2 are localized abundantly, and that localization of Stt4 is maintained in the Δtether mutant, which has a reduced number of ER-PM contact sites. We also demonstrated that the Δtether and sac1Δ mutants showed defects at different stages of endocytosis, and that inactivation mutation of Stt4 restored the endocytosis defect only in the Δtether mutant. Furthermore, these mutants exhibited defective transport in the secretory and recycling pathways, and inactivation of Stt4 restored the secretory pathway in the Δtether mutant, but not the recycling pathway in either mutant. These results suggest that endocytosis, secretion, and recycling pathways are regulated directly or indirectly by different PtdIns(4)P-mediated mechanisms.
越来越多的证据表明，在内质网（ER）-质膜（PM）接触位点处磷脂酰肌醇-4-磷酸（PtdIns(4)P）和磷脂酰丝氨酸（PS）的反向转运是细胞内囊泡运输所必需的。PtdIns(4)P由位于质膜的Stt4 PI 4-激酶和内质网的Sac1 PtdIns(4)P磷酸酶代谢，并且内质网-质膜接触位点被认为对其有效周转很重要。最近，已表明Stt4广泛定位于内质网-质膜接触位点。然而，Stt4的确切位置及其定位于这些位点的机制尚未阐明。此外，尽管多项研究表明在内吞作用中内质网-质膜接触位点处PS/PtdIns(4)P和固醇/PtdIns(4)P交换是必需的，但内质网与质膜之间的接触、PtdIns(4)P或PS的周转，还是PtdIns(4)P或PS水平的维持哪个更重要仍不清楚。在这里，我们发现Stt4定位于Scs2和Ist2大量存在的cER区域，并且Stt4的定位在具有减少数量的内质网-质膜接触位点的Δtether突变体中得以维持。我们还证明，Δtether和sac1Δ突变体在内吞作用的不同阶段表现出缺陷，并且Stt4的失活突变仅在Δtether突变体中恢复了内吞作用缺陷。此外，这些突变体在分泌和再循环途径中表现出运输缺陷，并且Stt4的失活恢复了Δtether突变体中的分泌途径，但未恢复任何一个突变体中的再循环途径。这些结果表明，内吞作用、分泌和再循环途径直接或间接受不同的PtdIns(4)P介导机制调节。
