Occurrence of the a subunits of G proteins in cerebral cortex synaptic membrane and postsynaptic density fractions

Modulation of ADP-ribosylation by Ca2+/calmodulin

Wu Kuo, Sanjay K. Nigam, Marie Leuoux, Yung Yu Huang, Chiye Aoki, Philip Siekevitz

Research output: Contribution to journalArticle

Abstract

We have examined the isolated postsynaptic density (PSD) fraction for the presence of a G protein. First, we found specific binding of guanosine 5′-[γ-[35S]thio]triphosphate to the PSD. Second, pertussis toxin-activated ADP-ribosylation of the isolated PSD fraction resulted in the appearance of a G protein with an apparent molecular mass of 41 kDa, and two G proteins with apparent molecular masses of 41 kDa and 39 kDa in synaptic membrane (SM) fraction and total homogenate (H). The amount of the 41-kDa G protein per unit protein was in the order of SM > H > PSD. Anti-Gi antibodies recognized the 41-kDa G protein in both PSD and SM, whereas anti-Go antibodies reacted with the 39-kDa G protein in the SM. The absence of Go protein in the PSD suggested that there was no contamination with SM. Moreover, unlabeled PSD incubated with an extract of SM that contained the labeled G proteins resulted in no label in the subsequently reisolated PSD, suggesting that the G protein found in the PSD was not due to adsorption of the G protein onto the PSD during its isolation from the SM. PSD pretreated with EGTA gave an 11-fold increase in the ADP-ribosylation reaction of the Gi protein; similar effects on the G1 and Go proteins of SM were obtained. Restoration of Ca2+/calmodulin to the PSD, but not of either Ca2+ or calmodulin alone, removed the effect of EGTA, indicating a strong complex formation between Gi and Ca2+/calmodulin that decreased the ADP-ribosylation reaction. Preincubation with the Ca2+-channel blocker nifedipine decreased the ADP-ribosylation reaction in the PSD. We conclude that Gi is present in the PSD, that it may interact with calmodulin and that it is involved in the regulation of voltage-dependent Ca2+ channel. We present a theory of the involvement of the G protein and calmodulin in postsynaptic neurophysiological events.

Original languageEnglish (US)
Pages (from-to)8686-8690
Number of pages5
JournalProceedings of the National Academy of Sciences of the United States of America
Volume89
Issue number18
StatePublished - 1992

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Post-Synaptic Density
Synaptic Membranes
Calmodulin
GTP-Binding Proteins
Cerebral Cortex
Adenosine Diphosphate
Egtazic Acid
Anti-Idiotypic Antibodies
Proteins
Guanosine
Pertussis Toxin
Nifedipine
Adsorption

Keywords

  • Signaling

ASJC Scopus subject areas

  • Genetics
  • General

Cite this

Occurrence of the a subunits of G proteins in cerebral cortex synaptic membrane and postsynaptic density fractions : Modulation of ADP-ribosylation by Ca2+/calmodulin. / Kuo, Wu; Nigam, Sanjay K.; Leuoux, Marie; Huang, Yung Yu; Aoki, Chiye; Siekevitz, Philip.

In: Proceedings of the National Academy of Sciences of the United States of America, Vol. 89, No. 18, 1992, p. 8686-8690.

Research output: Contribution to journalArticle

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AU - Aoki, Chiye

AU - Siekevitz, Philip

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