Possible involvement of phosphatidylinositol-specific phospholipase C related to pertussis toxin-sensitive GTP-binding proteins during adipocyte differentiation of 3T3-L1 fibroblasts: Negative regulation of protein kinase C

Takashi Uehara, Shin ichi Hoshino, Michio Ui, Yukiko Tokumitsu, Yasuyuki Nomura

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Abstract

Insulin/dexamethasone/methylisobutylxanthine (hormones/IBMX) induce 3T3-L1 fibroblasts to differentiate into adipocytes. Our previous study suggested that pertussis toxin (IAP)-sensitive GTP-binding protein(s) (G-protein) is involved in the process of differentiation by hormones/IBMX, accompanied by c-fos induction. Northern blotting indicated that among the IAP-sensitive G-proteins, the levels of Gi2α, Goα, and Gi3α mRNA were decreased, increased and unchanged, respectively. Gi1α was undetectable and IAP attenuated the decrease in Gi2α mRNA level but did not affect the change in Goα mRNA level during the adipocyte differentiation. These results indicate that IAP-sensitive Gi2α mRNA level is decreased during adipocyte differentiation. A combination of phosphatidylinositol-specific phospholipase C (PI-PLC) and IBMX induced c-fos expression in 3T3-L1 fibroblasts similar to that induced with hormones/IBMX. c-fos induced by both stimulators was also diminished by anti-inositolglycan antibody or anti-PI-PLC antiserum. Insulin stimulated the release of inositolproteoglycan and diacylglycerol from 3T3-L1 fibroblasts, which was suppressed by IAP treatment. These findings suggested that one of the pathways of adipocyte differentiation induced by hormones/IBMX occurs via the inositolglycan-specific PI-PLC cascade coupled to IAP-sensitive G-protein(s). Both activation of glycerophosphate dehydrogenase and stimulation of insulin-dependent 2-deoxyglucose uptake induced by hormones/IBMX were enhanced in protein kinase C-depleted cells exposed to phorbol 12-myristate 13-acetate (PMA), and attenuated in IAP-treated cells. The level of a 32P-labeled 52 kDa protein in plasma membrane fractions immunoprecipitated by anti-PI-PLC antiserum was increased by PMA stimulation, abolished in PMA-treated cells, and increased in IAP-treated cells. These findings suggest that protein kinase C phosphorylates PI-PLC, resulting in a decrease in PI-PLC activity related to the signal transduction pathway of adipocyte differentiation of 3T3-L1 fibroblasts.

Original languageEnglish
Pages (from-to)302-310
Number of pages9
JournalBiochimica et Biophysica Acta - Molecular Cell Research
Volume1224
Issue number2
DOIs
Publication statusPublished - Nov 10 1994
Externally publishedYes

Fingerprint

Phosphoinositide Phospholipase C
1-Methyl-3-isobutylxanthine
Pertussis Toxin
GTP-Binding Proteins
Adipocytes
Protein Kinase C
Fibroblasts
Hormones
Messenger RNA
Acetates
Insulin
Immune Sera
Glycerolphosphate Dehydrogenase
Diglycerides
Deoxyglucose
Northern Blotting
Dexamethasone
Anti-Idiotypic Antibodies
Signal Transduction
Cell Membrane

Keywords

  • 3T3-L1 cell
  • Differentiation
  • G protein
  • Insulin
  • Pertussis toxin (IAP)
  • PI-PLC

ASJC Scopus subject areas

  • Biophysics
  • Cell Biology
  • Molecular Biology

Cite this

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title = "Possible involvement of phosphatidylinositol-specific phospholipase C related to pertussis toxin-sensitive GTP-binding proteins during adipocyte differentiation of 3T3-L1 fibroblasts: Negative regulation of protein kinase C",
abstract = "Insulin/dexamethasone/methylisobutylxanthine (hormones/IBMX) induce 3T3-L1 fibroblasts to differentiate into adipocytes. Our previous study suggested that pertussis toxin (IAP)-sensitive GTP-binding protein(s) (G-protein) is involved in the process of differentiation by hormones/IBMX, accompanied by c-fos induction. Northern blotting indicated that among the IAP-sensitive G-proteins, the levels of Gi2α, Goα, and Gi3α mRNA were decreased, increased and unchanged, respectively. Gi1α was undetectable and IAP attenuated the decrease in Gi2α mRNA level but did not affect the change in Goα mRNA level during the adipocyte differentiation. These results indicate that IAP-sensitive Gi2α mRNA level is decreased during adipocyte differentiation. A combination of phosphatidylinositol-specific phospholipase C (PI-PLC) and IBMX induced c-fos expression in 3T3-L1 fibroblasts similar to that induced with hormones/IBMX. c-fos induced by both stimulators was also diminished by anti-inositolglycan antibody or anti-PI-PLC antiserum. Insulin stimulated the release of inositolproteoglycan and diacylglycerol from 3T3-L1 fibroblasts, which was suppressed by IAP treatment. These findings suggested that one of the pathways of adipocyte differentiation induced by hormones/IBMX occurs via the inositolglycan-specific PI-PLC cascade coupled to IAP-sensitive G-protein(s). Both activation of glycerophosphate dehydrogenase and stimulation of insulin-dependent 2-deoxyglucose uptake induced by hormones/IBMX were enhanced in protein kinase C-depleted cells exposed to phorbol 12-myristate 13-acetate (PMA), and attenuated in IAP-treated cells. The level of a 32P-labeled 52 kDa protein in plasma membrane fractions immunoprecipitated by anti-PI-PLC antiserum was increased by PMA stimulation, abolished in PMA-treated cells, and increased in IAP-treated cells. These findings suggest that protein kinase C phosphorylates PI-PLC, resulting in a decrease in PI-PLC activity related to the signal transduction pathway of adipocyte differentiation of 3T3-L1 fibroblasts.",
keywords = "3T3-L1 cell, Differentiation, G protein, Insulin, Pertussis toxin (IAP), PI-PLC",
author = "Takashi Uehara and Hoshino, {Shin ichi} and Michio Ui and Yukiko Tokumitsu and Yasuyuki Nomura",
year = "1994",
month = "11",
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language = "English",
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T1 - Possible involvement of phosphatidylinositol-specific phospholipase C related to pertussis toxin-sensitive GTP-binding proteins during adipocyte differentiation of 3T3-L1 fibroblasts

T2 - Negative regulation of protein kinase C

AU - Uehara, Takashi

AU - Hoshino, Shin ichi

AU - Ui, Michio

AU - Tokumitsu, Yukiko

AU - Nomura, Yasuyuki

PY - 1994/11/10

Y1 - 1994/11/10

N2 - Insulin/dexamethasone/methylisobutylxanthine (hormones/IBMX) induce 3T3-L1 fibroblasts to differentiate into adipocytes. Our previous study suggested that pertussis toxin (IAP)-sensitive GTP-binding protein(s) (G-protein) is involved in the process of differentiation by hormones/IBMX, accompanied by c-fos induction. Northern blotting indicated that among the IAP-sensitive G-proteins, the levels of Gi2α, Goα, and Gi3α mRNA were decreased, increased and unchanged, respectively. Gi1α was undetectable and IAP attenuated the decrease in Gi2α mRNA level but did not affect the change in Goα mRNA level during the adipocyte differentiation. These results indicate that IAP-sensitive Gi2α mRNA level is decreased during adipocyte differentiation. A combination of phosphatidylinositol-specific phospholipase C (PI-PLC) and IBMX induced c-fos expression in 3T3-L1 fibroblasts similar to that induced with hormones/IBMX. c-fos induced by both stimulators was also diminished by anti-inositolglycan antibody or anti-PI-PLC antiserum. Insulin stimulated the release of inositolproteoglycan and diacylglycerol from 3T3-L1 fibroblasts, which was suppressed by IAP treatment. These findings suggested that one of the pathways of adipocyte differentiation induced by hormones/IBMX occurs via the inositolglycan-specific PI-PLC cascade coupled to IAP-sensitive G-protein(s). Both activation of glycerophosphate dehydrogenase and stimulation of insulin-dependent 2-deoxyglucose uptake induced by hormones/IBMX were enhanced in protein kinase C-depleted cells exposed to phorbol 12-myristate 13-acetate (PMA), and attenuated in IAP-treated cells. The level of a 32P-labeled 52 kDa protein in plasma membrane fractions immunoprecipitated by anti-PI-PLC antiserum was increased by PMA stimulation, abolished in PMA-treated cells, and increased in IAP-treated cells. These findings suggest that protein kinase C phosphorylates PI-PLC, resulting in a decrease in PI-PLC activity related to the signal transduction pathway of adipocyte differentiation of 3T3-L1 fibroblasts.

AB - Insulin/dexamethasone/methylisobutylxanthine (hormones/IBMX) induce 3T3-L1 fibroblasts to differentiate into adipocytes. Our previous study suggested that pertussis toxin (IAP)-sensitive GTP-binding protein(s) (G-protein) is involved in the process of differentiation by hormones/IBMX, accompanied by c-fos induction. Northern blotting indicated that among the IAP-sensitive G-proteins, the levels of Gi2α, Goα, and Gi3α mRNA were decreased, increased and unchanged, respectively. Gi1α was undetectable and IAP attenuated the decrease in Gi2α mRNA level but did not affect the change in Goα mRNA level during the adipocyte differentiation. These results indicate that IAP-sensitive Gi2α mRNA level is decreased during adipocyte differentiation. A combination of phosphatidylinositol-specific phospholipase C (PI-PLC) and IBMX induced c-fos expression in 3T3-L1 fibroblasts similar to that induced with hormones/IBMX. c-fos induced by both stimulators was also diminished by anti-inositolglycan antibody or anti-PI-PLC antiserum. Insulin stimulated the release of inositolproteoglycan and diacylglycerol from 3T3-L1 fibroblasts, which was suppressed by IAP treatment. These findings suggested that one of the pathways of adipocyte differentiation induced by hormones/IBMX occurs via the inositolglycan-specific PI-PLC cascade coupled to IAP-sensitive G-protein(s). Both activation of glycerophosphate dehydrogenase and stimulation of insulin-dependent 2-deoxyglucose uptake induced by hormones/IBMX were enhanced in protein kinase C-depleted cells exposed to phorbol 12-myristate 13-acetate (PMA), and attenuated in IAP-treated cells. The level of a 32P-labeled 52 kDa protein in plasma membrane fractions immunoprecipitated by anti-PI-PLC antiserum was increased by PMA stimulation, abolished in PMA-treated cells, and increased in IAP-treated cells. These findings suggest that protein kinase C phosphorylates PI-PLC, resulting in a decrease in PI-PLC activity related to the signal transduction pathway of adipocyte differentiation of 3T3-L1 fibroblasts.

KW - 3T3-L1 cell

KW - Differentiation

KW - G protein

KW - Insulin

KW - Pertussis toxin (IAP)

KW - PI-PLC

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U2 - 10.1016/0167-4889(94)90204-6

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AN - SCOPUS:0028171870

VL - 1224

SP - 302

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JO - Biochimica et Biophysica Acta - Molecular Cell Research

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