کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
1950510 1055644 2015 11 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
Class III PI 3-kinase is the main source of PtdIns3P substrate and membrane recruitment signal for PIKfyve constitutive function in podocyte endomembrane homeostasis
موضوعات مرتبط
علوم زیستی و بیوفناوری بیوشیمی، ژنتیک و زیست شناسی مولکولی زیست شیمی
پیش نمایش صفحه اول مقاله
Class III PI 3-kinase is the main source of PtdIns3P substrate and membrane recruitment signal for PIKfyve constitutive function in podocyte endomembrane homeostasis
چکیده انگلیسی


• PIKfyve is critical for endomembrane homeostasis.
• PIKfyve uses PtdIns3P as both an enzyme substrate and a membrane recruitment signal.
• We find that Vps34 is a main PtdIns3P supplier for constitutive PIKfyve functionality.
• Cytoplasmic vacuoles due to Vps34 KO in podocytes are secondary to PIKfyve dysfunction.
• Podocyte endomembrane homeostasis requires a functional link between Vps34 and PIKfyve.

The evolutionarily conserved PIKfyve, which synthesizes PtdIns5P from PtdIns, and PtdIns(3,5)P2 from PtdIns3P, requires PtdIns3P as both an enzyme substrate and a membrane recruitment signal. Whereas the PtdIns3P source is undetermined, class III PI3K (Vps34), the only evolutionarily conserved of the eight mammalian PI3Ks, is presumed as a main candidate. A hallmark of PIKfyve deficiency is formation of multiple translucent cytoplasmic vacuoles seen by light microscopy in cells cultured in complete media. Such an aberrant phenotype is often observed in cells from conditional Vps34 knockout (KO) mice. To clarify the mechanism of Vps34 KO-triggered vacuolation and the PtdIns3P source for PIKfyve functionality, here we have characterized a podocyte cell type derived from Vps34fl/fl mice, which, upon Cre-mediated gene KO, robustly formed cytoplasmic vacuoles resembling those in PikfyveKO MEFs. Vps34wt, expressed in Vps34KO podocytes restored the normal morphology, but only if the endogenous PIKfyve activity was intact. Conversely, expressed PIKfyvewt rescued completely the vacuolation only in PikfyveKO MEFs but not in Vps34KO podocytes. Analyses of phosphoinositide profiles by HPLC and localization patterns by a PtdIns3P biosensor revealed that Vps34 is the main supplier of localized PtdIns3P not only for PIKfyve activity but also for membrane recruitment. Concordantly, Vps34KO podocytes had severely reduced steady-state levels of both PtdIns(3,5)P2 and PtdIns5P, along with PtdIns3P. We further revealed a plausible physiologically-relevant Vps34-independent PtdIns3P supply for PIKfyve, operating through activated class I PI3Ks. Our data provide the first evidence that the vacuolation phenotype in Vps34KO podocytes is due to PIKfyve dysfunction and that Vps34 is a main PtdIns3P source for constitutive PIKfyve functionality.

ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Biochimica et Biophysica Acta (BBA) - Molecular Cell Research - Volume 1853, Issue 5, May 2015, Pages 1240–1250
نویسندگان
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