Measurement of Subcellular Sites of Polyphosphoinositide Metabolism in Isolated Rat Hepatocytes

William W. Wells, Mark A. Seyfred, Charles Smith, Masahiro Sakai

Research output: Contribution to journalArticle

10 Citations (Scopus)

Abstract

The products of phosphatidylinositol bisphosphate (PIP2) degradation––diacylglycerol and D-myo-inositol 1, 4, 5-trisphosphate (IP3)––have both been identified as intracellular second messengers. Numerous cell types respond to a variety of stimuli with the enhanced hydrolysis of PIP2 and to a lesser extent of phosphatidylinositol 4-phosphate (PIP). The plasma membrane PIP2 phosphodiesterase does not appear to hydrolyze phosphatidylinositol (PI) in liver. One approach to the investigation of the subcellular location of polyphosphoinositide metabolic reactions is to fractionate cell homogenates by the best procedures available and to assay the respective enzymes with added substrates under Vmax conditions. A second approach for determining the subcellular sites of the metabolism of PIP and PIP2 is to analyze the distribution of PIP and PIP2, the products of PI and PIP kinase, in a cell; however, their low levels make analysis by existing chemical means virtually impossible. The chapter explains the preparation of phosphatidylinositol [4-32P] phosphate and phosphatidylinositol [4,5-32p]bisphosphate.

Original languageEnglish (US)
Pages (from-to)92-99
Number of pages8
JournalMethods in enzymology
Volume141
Issue numberC
DOIs
StatePublished - Jan 1 1987

Fingerprint

Phosphatidylinositol Phosphates
Phosphatidylinositols
Metabolism
Rats
Hepatocytes
Inositol 1,4,5-Trisphosphate
Phosphoric Diester Hydrolases
Enzyme Assays
Second Messenger Systems
Cell membranes
Liver
Hydrolysis
Assays
Cell Membrane
phosphatidylinositol 4-phosphate
Substrates
Enzymes

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology

Cite this

Wells, William W. ; Seyfred, Mark A. ; Smith, Charles ; Sakai, Masahiro. / Measurement of Subcellular Sites of Polyphosphoinositide Metabolism in Isolated Rat Hepatocytes. In: Methods in enzymology. 1987 ; Vol. 141, No. C. pp. 92-99.
@article{8daa92a2722f422cbd472868c0a4da75,
title = "Measurement of Subcellular Sites of Polyphosphoinositide Metabolism in Isolated Rat Hepatocytes",
abstract = "The products of phosphatidylinositol bisphosphate (PIP2) degradation––diacylglycerol and D-myo-inositol 1, 4, 5-trisphosphate (IP3)––have both been identified as intracellular second messengers. Numerous cell types respond to a variety of stimuli with the enhanced hydrolysis of PIP2 and to a lesser extent of phosphatidylinositol 4-phosphate (PIP). The plasma membrane PIP2 phosphodiesterase does not appear to hydrolyze phosphatidylinositol (PI) in liver. One approach to the investigation of the subcellular location of polyphosphoinositide metabolic reactions is to fractionate cell homogenates by the best procedures available and to assay the respective enzymes with added substrates under Vmax conditions. A second approach for determining the subcellular sites of the metabolism of PIP and PIP2 is to analyze the distribution of PIP and PIP2, the products of PI and PIP kinase, in a cell; however, their low levels make analysis by existing chemical means virtually impossible. The chapter explains the preparation of phosphatidylinositol [4-32P] phosphate and phosphatidylinositol [4,5-32p]bisphosphate.",
author = "Wells, {William W.} and Seyfred, {Mark A.} and Charles Smith and Masahiro Sakai",
year = "1987",
month = "1",
day = "1",
doi = "10.1016/0076-6879(87)41058-6",
language = "English (US)",
volume = "141",
pages = "92--99",
journal = "Methods in Enzymology",
issn = "0076-6879",
publisher = "Academic Press Inc.",
number = "C",

}

Measurement of Subcellular Sites of Polyphosphoinositide Metabolism in Isolated Rat Hepatocytes. / Wells, William W.; Seyfred, Mark A.; Smith, Charles; Sakai, Masahiro.

In: Methods in enzymology, Vol. 141, No. C, 01.01.1987, p. 92-99.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Measurement of Subcellular Sites of Polyphosphoinositide Metabolism in Isolated Rat Hepatocytes

AU - Wells, William W.

AU - Seyfred, Mark A.

AU - Smith, Charles

AU - Sakai, Masahiro

PY - 1987/1/1

Y1 - 1987/1/1

N2 - The products of phosphatidylinositol bisphosphate (PIP2) degradation––diacylglycerol and D-myo-inositol 1, 4, 5-trisphosphate (IP3)––have both been identified as intracellular second messengers. Numerous cell types respond to a variety of stimuli with the enhanced hydrolysis of PIP2 and to a lesser extent of phosphatidylinositol 4-phosphate (PIP). The plasma membrane PIP2 phosphodiesterase does not appear to hydrolyze phosphatidylinositol (PI) in liver. One approach to the investigation of the subcellular location of polyphosphoinositide metabolic reactions is to fractionate cell homogenates by the best procedures available and to assay the respective enzymes with added substrates under Vmax conditions. A second approach for determining the subcellular sites of the metabolism of PIP and PIP2 is to analyze the distribution of PIP and PIP2, the products of PI and PIP kinase, in a cell; however, their low levels make analysis by existing chemical means virtually impossible. The chapter explains the preparation of phosphatidylinositol [4-32P] phosphate and phosphatidylinositol [4,5-32p]bisphosphate.

AB - The products of phosphatidylinositol bisphosphate (PIP2) degradation––diacylglycerol and D-myo-inositol 1, 4, 5-trisphosphate (IP3)––have both been identified as intracellular second messengers. Numerous cell types respond to a variety of stimuli with the enhanced hydrolysis of PIP2 and to a lesser extent of phosphatidylinositol 4-phosphate (PIP). The plasma membrane PIP2 phosphodiesterase does not appear to hydrolyze phosphatidylinositol (PI) in liver. One approach to the investigation of the subcellular location of polyphosphoinositide metabolic reactions is to fractionate cell homogenates by the best procedures available and to assay the respective enzymes with added substrates under Vmax conditions. A second approach for determining the subcellular sites of the metabolism of PIP and PIP2 is to analyze the distribution of PIP and PIP2, the products of PI and PIP kinase, in a cell; however, their low levels make analysis by existing chemical means virtually impossible. The chapter explains the preparation of phosphatidylinositol [4-32P] phosphate and phosphatidylinositol [4,5-32p]bisphosphate.

UR - http://www.scopus.com/inward/record.url?scp=0023072222&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0023072222&partnerID=8YFLogxK

U2 - 10.1016/0076-6879(87)41058-6

DO - 10.1016/0076-6879(87)41058-6

M3 - Article

VL - 141

SP - 92

EP - 99

JO - Methods in Enzymology

JF - Methods in Enzymology

SN - 0076-6879

IS - C

ER -