Mutations on the Switch III region and the alpha3 helix of Galpha16 differentially affect receptor coupling and regulation of downstream effectorsReportar como inadecuado




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Journal of Molecular Signaling

, 3:17

First Online: 22 November 2008Received: 13 August 2008Accepted: 22 November 2008

Abstract

BackgroundGα16 can activate phospholipase Cβ PLCβ directly like Gαq. It also couples to tetratricopeptide repeat 1 TPR1 which is linked to Ras activation. It is unknown whether PLCβ and TPR1 interact with the same regions on Gα16. Previous studies on Gαq have defined two minimal clusters of amino acids that are essential for the coupling to PLCβ. Cognate residues in Gα16 might also be essential for interacting with PLCβ, and possibly contribute to TPR1 interaction and other signaling events.

ResultsAlanine mutations were introduced to the two amino acid clusters 246–248 and 259–260 in the switch III region and α3 helix of Gα16. Regulations of PLCβ and STAT3 were partially weakened by each cluster mutant. A mutant harboring mutations at both clusters generally produced stronger suppressions. Activation of Jun N-terminal kinase JNK by Gα16 was completely abolished by mutating either clusters. Contrastingly, phosphorylations of extracellular signal-regulated kinase ERK and nuclear factor κB NF-κB were not significantly affected by these mutations. The interactions between the mutants and PLCβ2 and TPR1 were also reduced in co-immunoprecipitation assays. Coupling between G16 and different categories of receptors was impaired by the mutations, with the effect of switch III mutations being more pronounced than those in the α3 helix. Mutations of both clusters almost completely abolished the receptor coupling and prevent receptor-induced Gβγ release.

ConclusionThe integrity of the switch III region and α3 helix of Gα16 is critical for the activation of PLCβ, STAT3, and JNK but not ERK or NF-κB. Binding of Gα16 to PLCβ2 or TPR1 was reduced by the mutations of either cluster. The same region could also differentially affect the effectiveness of receptor coupling to G16. The studied region was shown to bear multiple functionally important roles of G16.

AbbreviationsA1R, A2AR, A2BRtypes 1, 2A and 2B adenosine receptor

ACadenylyl cyclase

CHAN-cyclohexyladenosine

D1Rtype 1 dopamine receptor

ERKextracellular signal-regulated kinases 1 and 2

fMLPN-formyl-methioinyl-leucyl-phenylalanine

GPCRG protein coupled receptor

GTPaseGTP hydrolase

IBMXisobutylmethylxanthine

IKKinhibitor of κB kinase

IPinositol phosphates

JNKc-jun N-terminal kinase

MAPKmitogen-activating protein kinase

MEKMAPK-ERK kinase

MEMEagle-s minimal essential medium

MKKMAPK kinase

NF-κBnuclear factor κB

PKCprotein kinase C

PLCβphospholipase Cβ isoform

PTXpertussis toxin

STAT3type 3 signal transducers and activator of transcription protein

TPR1tetratricopeptide repeat protein 1

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Autor: May YM Yu - Maurice KC Ho - Andrew MF Liu - Yung H Wong

Fuente: https://link.springer.com/







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