Annexin A6 at the cardiac myocyte sarcolemma--evidence for self-association and binding to actin.
The plasma membrane of the heart muscle cell and its underlying cytoskeleton are vitally important to the function of the heart. Annexin A6 is a major cellular calcium and phospholipid binding protein. Here we show that annexin A6 copurifies with sarcolemma isolated from pig heart. Two pools of annexin A6 are present in the sarcolemma fraction, one dependent on calcium and one that resists extraction by the calcium chelator EGTA. Potential annexin A6 binding proteins in the sarcolemma fraction were identified using Far Western blotting. Two major annexin A6 binding proteins were identified as actin and annexin A6 itself. Annexin A6 bound to itself both in the presence and in the absence of calcium ions. Sites for self association were mapped by performing Western blots on proteolytic fragments of recombinant annexin A6. Annexin A6 bound preferentially not only to the N terminal fragment (domains I-IV, residues 1-352) but also to C-terminal fragments corresponding to domains V+VI and domains VII+VIII. Actin binding to annexin A6 was calcium-dependent and exclusively to the N-terminal fragment of annexin A6. A calcium-dependent complex of annexin A6 and actin may stabilize the cardiomyocyte sarcolemma during cell stimulation.
Locate
Salma
S
Faculty of Biological Sciences, School of Biochemistry and Microbiology, University of Leeds, Leeds, UK.
Colyer
John
J
Gawler
Debra J
DJ
Walker
John H
JH
D000199:Actins / Q000378:metabolism*; D000818:Animals; D017317:Annexin A6 / Q000737:chemistry / Q000378:metabolism*; D026481:Blotting, Far-Western; D002118:Calcium / Q000378:metabolism; D020013:Calcium Signaling / Q000502:physiology; D046911:Macromolecular Substances / Q000378:metabolism*; D032383:Myocytes, Cardiac / Q000378:metabolism* / Q000648:ultrastructure; D010447:Peptide Hydrolases / Q000737:chemistry; D011485:Protein Binding / Q000502:physiology; D017434:Protein Structure, Tertiary / Q000502:physiology; D040901:Proteomics / Q000379:methods; D051381:Rats; D012508:Sarcolemma / Q000378:metabolism* / Q000648:ultrastructure; D034421:Sus scrofa
D000199:Actins; D017317:Annexin A6; D046911:Macromolecular Substances; D010447:Peptide Hydrolases; D002118:Calcium