RRC ID 6647
Author Swigart P, Insall R, Wilkins A, Cockcroft S.
Title Purification and cloning of phosphatidylinositol transfer proteins from Dictyostelium discoideum: homologues of both mammalian PITPs and Saccharomyces cerevisiae sec14p are found in the same cell.
Journal Biochem J
Abstract Soluble phosphatidylinositol transfer proteins (PITPs) have important roles in lipid-mediated signalling as well as in membrane traffic. Two PITPs (alpha and beta) have been cloned from mammalian cells, which are unrelated in sequence to yeast PITP (the product of the SEC14 gene). However, all three PITPs can perform interchangeably to reconstitute function in mammalian cells. We have now purified the major PITP from the cytoplasm of Dictyostelium discoideum and cloned the gene. This protein, DdPITP1, is homologous with mammalian PITPalpha and PITPbeta. We have also cloned a second gene (DdPITP2) related in sequence to DdPITP1. In addition, an independently cloned cDNA encodes a relative of the SEC14 family of yeast PITPs. DdPITP1, DdPITP2 and DdSec14 proteins were all able to mediate the transfer of PtdIns from one membrane compartment to another; they thus exhibited the hallmark of PITPs. Secondly, all three PITPs were able to rescue phospholipase C-mediated phosphoinositide hydrolysis in PITP-depleted HL60 cells, indicating that all three PITPs were capable of stimulating phosphoinositide synthesis. The identification of PITPs related to both mammalian PITPs and yeast Sec14p in a single organism will provide a unique opportunity to examine the functions of this class of protein with genetic approaches.
Volume 347 Pt 3(Pt 3)
Pages 837-43
Published 2000-5-1
PMID 10769190
PMC PMC1221023
MeSH Amino Acid Sequence Animals Carrier Proteins / chemistry Carrier Proteins / genetics* Carrier Proteins / isolation & purification* Carrier Proteins / metabolism Cell Line Cloning, Molecular Cross Reactions Cytosol / chemistry Cytosol / enzymology Dictyostelium / chemistry* Dictyostelium / cytology* Dictyostelium / genetics Dictyostelium / growth & development Escherichia coli Genes, Protozoan / genetics Genetic Complementation Test Golgi Apparatus / chemistry HL-60 Cells Humans Isoenzymes / metabolism Membrane Proteins* Molecular Sequence Data Molecular Weight Phosphatidylinositols / metabolism Phospholipase C beta Phospholipid Transfer Proteins Protozoan Proteins / chemistry Protozoan Proteins / genetics* Protozoan Proteins / isolation & purification* Protozoan Proteins / metabolism RNA, Messenger / analysis RNA, Messenger / genetics Recombinant Fusion Proteins / chemistry Recombinant Fusion Proteins / genetics Recombinant Fusion Proteins / isolation & purification Recombinant Fusion Proteins / metabolism Saccharomyces cerevisiae / chemistry Saccharomyces cerevisiae Proteins* Sequence Alignment Sequence Homology, Amino Acid Type C Phospholipases / metabolism
IF 4.097
Times Cited 24
Cellular slime molds G00173