PC phosphorylation increases the ability of AFAP-110 to cross-link actin filaments.
The actin filament-associated protein and Src-binding partner, AFAP-110, is an adaptor protein that links signaling molecules to actin filaments. AFAP-110 binds actin filaments directly and multimerizes through a leucine zipper motif. Cellular signals downstream of Src(527F) can regulate multimerization. Here, we determined recombinant AFAP-110 (rAFAP-110)-bound actin filaments cooperatively, through a lateral association. We demonstrate rAFAP-110 has the capability to cross-link actin filaments, and this ability is dependent on the integrity of the carboxy terminal actin binding domain. Deletion of the leucine zipper motif or PKC phosphorylation affected AFAP-110's conformation, which correlated with changes in multimerization and increased the capability of rAFAP-110 to cross-link actin filaments. AFAP-110 is both a substrate and binding partner of PKC. On PKC activation, stress filament organization is lost, motility structures form, and AFAP-110 colocalizes strongly with motility structures. Expression of a deletion mutant of AFAP-110 that is unable to bind PKC blocked the effect of PMA on actin filaments. We hypothesize that upon PKC activation, AFAP-110 can be cooperatively recruited to newly forming actin filaments, like those that exist in cell motility structures, and that PKC phosphorylation effects a conformational change that may enable AFAP-110 to promote actin filament cross-linking at the cell membrane.
The Mary Babb Randolph Cancer Center and the Department of Microbiology and Immunology, West Virginia University, Morgantown, West Virginia 26506-9300, USA.
Yong
Y
Qian
Joseph M
JM
Baisden
Lidia
L
Cherezova
Justin M
JM
Summy
Anne
A
Guappone-Koay
Xianglin
X
Shi
Tom
T
Mast
Jennifer
J
Pustula
Henry G
HG
Zot
Nayef
N
Mazloum
Marietta Y
MY
Lee
Daniel C
DC
Flynn
D000185:Actinin / Q000378:metabolism; D000199:Actins / Q000378:metabolism*; D000818:Animals; D001665:Binding Sites; D002460:Cell Line; D003432:Cross-Linking Reagents / Q000378:metabolism; D004789:Enzyme Activation; D016350:Leucine Zippers; D008840:Microfilament Proteins / Q000235:genetics / Q000378:metabolism*; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D011108:Polymers / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D011493:Protein Kinase C / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism
C084510:AFAP 110; D000199:Actins; D003432:Cross-Linking Reagents; D008840:Microfilament Proteins; D010750:Phosphoproteins; D011108:Polymers; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D000185:Actinin; D011493:Protein Kinase C