Dataset Information

Stress induced ? subunit of heterotrimeric G-proteins from Pisum sativum interacts with mitogen activated protein kinase.

ABSTRACT: We here report in Pisum sativum system a novel protein-protein interaction of ?-subunit of heterotrimeric G-proteins (PsG?) with a Mitogen activated protein kinase (PsMPK3) during cDNA library screening by yeast-two-hybrid assay. The transcript of these two genes also showed co-regulation under abscisic acid (ABA) and methyl jasmonate (MeJA) treatments. The protein-protein interaction was further validated by performing one-to-one interaction and ?-galactosidase assay in yeast system. ?-subunit of G-proteins from a heterologous system Oryzae sativa also showed interaction with PsMPK3. The interaction between PsG? and PsMPK3 was further confirmed by in vitro protein-protein interaction. This suggested that MPK3 function as effector molecule for G?, which may helps in the regulation of stomatal functioning. These findings also provide an evidence for a possible cross-talk between MPK3 and G-protein-mediated signaling pathways in plants.

SUBMITTER: Bhardwaj D

PROVIDER: S-EPMC3121990 | biostudies-literature | 2011 Feb

REPOSITORIES: biostudies-literature

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Stress induced β subunit of heterotrimeric G-proteins from Pisum sativum interacts with mitogen activated protein kinase.

Bhardwaj Deepak D Sheikh Arsheed Hussain AH Sinha Alok Krishna AK Tuteja Narendra N

Plant signaling & behavior 20110201 2

We here report in Pisum sativum system a novel protein-protein interaction of β-subunit of heterotrimeric G-proteins (PsGβ) with a Mitogen activated protein kinase (PsMPK3) during cDNA library screening by yeast-two-hybrid assay. The transcript of these two genes also showed co-regulation under abscisic acid (ABA) and methyl jasmonate (MeJA) treatments. The protein-protein interaction was further validated by performing one-to-one interaction and β-galactosidase assay in yeast system. β-subunit ...[more]

PMID: 21350337

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Project description:Skeletal muscle phosphorylase kinase (PhK) is an (alphabetagammadelta) 4 hetero-oligomeric enzyme complex that phosphorylates and activates glycogen phosphorylase b (GP b) in a Ca (2+)-dependent reaction that couples muscle contraction with glycogen breakdown. GP b is PhK's only known in vivo substrate; however, given the great size and multiple subunits of the PhK complex, we screened muscle extracts for other potential targets. Extracts of P/J (control) and I/lnJ (PhK deficient) mice were incubated with [gamma- (32)P]ATP with or without Ca (2+) and compared to identify potential substrates. Candidate targets were resolved by two-dimensional polyacrylamide gel electrophoresis, and phosphorylated glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was identified by matrix-assisted laser desorption ionization mass spectroscopy. In vitro studies showed GAPDH to be a Ca (2+)-dependent substrate of PhK, although the rate of phosphorylation is very slow. GAPDH does, however, bind tightly to PhK, inhibiting at low concentrations (IC 50 approximately 0.45 microM) PhK's conversion of GP b. When a short synthetic peptide substrate was substituted for GP b, the inhibition was negligible, suggesting that GAPDH may inhibit predominantly by binding to the PhK complex at a locus distinct from its active site on the gamma subunit. To test this notion, the PhK-GAPDH complex was incubated with a chemical cross-linker, and a dimer between the regulatory beta subunit of PhK and GAPDH was formed. This interaction was confirmed by the fact that a subcomplex of PhK missing the beta subunit, specifically an alphagammadelta subcomplex, was unable to phosphorylate GAPDH, even though it is catalytically active toward GP b. Moreover, GAPDH had no effect on the conversion of GP b by the alphagammadelta subcomplex. The interactions described herein between the beta subunit of PhK and GAPDH provide a possible mechanism for the direct linkage of glycogenolysis and glycolysis in skeletal muscle.

Dataset Information

Stress induced ? subunit of heterotrimeric G-proteins from Pisum sativum interacts with mitogen activated protein kinase.

Publications

Stress induced β subunit of heterotrimeric G-proteins from Pisum sativum interacts with mitogen activated protein kinase.

Similar Datasets

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