Send to

Choose Destination
See comment in PubMed Commons below
J Biol Chem. 2002 Dec 6;277(49):47276-84. Epub 2002 Sep 20.

Phosphatidylinositol 5-phosphate biosynthesis is linked to PIKfyve and is involved in osmotic response pathway in mammalian cells.

Author information

Department of Physiology, Wayne State University School of Medicine, Detroit, Michigan 48201, USA.


The cellular functions, regulation and enzymology of phosphatidylinositol (PtdIns) 5-P, the newest addition to the family of phosphoinositides (PI), are still elusive. Whereas a kinase that uses PtdIns-5-P as an intracellular substrate has been assigned, a kinase that produces it remained to be identified. Here we report that PIKfyve, the enzyme found to synthesize PtdIns-5-P in vitro and PtdIns-3,5-P(2) in vitro and in vivo, is responsible for PtdIns-5-P production in a cellular context. Evidence is based on examination of two groups of cell types by two independent approaches. First, [(32)P]orthophosphate-labeled cells (Sf9, 3T3-L1 fibroblasts, and 3T3-L1 adipocytes) that show a high pressure liquid chromatography (HPLC)-detectable peak of the PtdIns-5-P head group at basal conditions demonstrated a 20-50% increase in radioactive PtdIns-5-P amounts upon expression of PIKfyve(WT). Second, cell types (HEK293), in which the basal levels of radioactive PtdIns-5-P were undetectable by HPLC head group analysis, demonstrated higher in vitro type II PIP kinase-directed conversion of the endogenous PtdIns-5-P pool into PtdIns-4,5-P(2), when induced to express PIKfyve(WT). Conversely, a decrease by 60% in the conversion of PtdIns-5-P to PtdIns-4,5-P(2) was associated with induced expression of the dominant-negative kinase-deficient PIKfyve(K1831E) mutant in HEK293 cells. When 3T3-L1 fibroblasts and 3T3-L1 adipocytes were subjected to osmotic shock, levels of PtdIns-5-P measured by both approaches were found to decrease profoundly upon a hypo-osmotic stimulus. Together, these results identify PIKfyve as an enzyme responsible for PtdIns-5-P biosynthesis and indicate a role for PtdIns-5-P in osmotic response pathways in mammalian cells.

[Indexed for MEDLINE]
Free full text
PubMed Commons home

PubMed Commons

How to join PubMed Commons

    Supplemental Content

    Full text links

    Icon for HighWire
    Loading ...
    Support Center