Identification of centaurin-alpha2: a phosphatidylinositide-binding protein present in fat, heart and skeletal muscle.
We describe here the cloning, expression and characterisation of centaurin-alpha2 from a rat adipocyte cDNA library. The centaurin-alpha2 cDNA contains an open reading frame, which codes for a protein of 376 amino acids with predicted mass of 43.5 kDa. Centaurin-alpha2 shares 51-59% identity with centaurin-alpha1 proteins and has the same domain organisation, consisting of a predicted N-terminal ArfGAP domain followed by two successive pleckstrin homology domains. Despite the sequence similarity, there are a number of notable differences between the previously characterised centaurin-alpha1 proteins and the newly described centaurin-alpha2: (i) in vitro lipid binding experiments with centaurin-alpha2 do not reveal the same selectivity for phosphatidylinositol 3,4,5-trisphosphate over phosphatidylinositol 4,5-bisphosphate that has been shown for centaurin-alpha; (ii) unlike centaurin-alpha1 which is expressed mainly in the brain, centaurin-alpha2 has a broad tissue distribution, being particularly abundant in fat, heart and skeletal muscle; (iii) in contrast to centaurin-alpha1 which is found in both membrane and cytosolic fractions, endogenous centaurin-alpha2 is exclusively present in the dense membrane fractions of cell extracts, suggesting a constitutive membrane association. Insulin stimulation, which stimulates phosphatidylinositol 3,4,5-trisphosphate production, does not alter the subcellular distribution of centaurin-alpha2 between adipocyte membrane fractions. This observation is consistent with the lack of specificity of centaurin-alpha2 for phosphatidylinositol 3,4,5-trisphosphate over phosphatidylinositol 4,5-bisphosphate.
Whitley
Paul
P
Department of Biology and Biochemistry, University of Bath, United Kingdom.
Gibbard
Alison M
AM
Koumanov
Françoise
F
Oldfield
Susan
S
Kilgour
Elaine E
EE
Prestwich
Glenn D
GD
Holman
Geoffrey D
GD
D048868:Adaptor Proteins, Signal Transducing; D000273:Adipose Tissue / Q000737:chemistry* / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002352:Carrier Proteins / Q000145:classification / Q000235:genetics* / Q000378:metabolism*; D002458:Cell Fractionation; D003001:Cloning, Molecular; D020690:GTPase-Activating Proteins; D006801:Humans; D008024:Ligands; D008081:Liposomes / Q000378:metabolism; D008969:Molecular Sequence Data; D018482:Muscle, Skeletal / Q000737:chemistry* / Q000378:metabolism; D009206:Myocardium / Q000737:chemistry* / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000145:classification / Q000235:genetics* / Q000378:metabolism*; D016366:Open Reading Frames; D010716:Phosphatidylinositols / Q000378:metabolism*; D010802:Phylogeny; D011485:Protein Binding; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D051381:Rats; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D013379:Substrate Specificity; D014018:Tissue Distribution
C100956:ADAP1 protein, human; C470610:ADAP2 protein, human; C491062:Adap1 protein, rat; C470609:Adap2 protein, rat; D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; D020690:GTPase-Activating Proteins; D008024:Ligands; D008081:Liposomes; D009419:Nerve Tissue Proteins; D010716:Phosphatidylinositols; D020033:Protein Isoforms; D011994:Recombinant Proteins