beta1 integrin cytoplasmic domain residues selectively modulate fibronectin matrix assembly and cell spreading through talin and Akt-1.
The integrin beta(1) cytoplasmic domain (tail) serves as a scaffold for numerous intracellular proteins. The mechanisms by which the tail coordinates these proteins to facilitate extracellular matrix assembly and cell spreading are not clear. This study demonstrates that the beta(1) cytoplasmic domain can regulate cell spreading on fibronectin and fibronectin matrix assembly through Akt- and talin-dependent mechanisms, respectively. To identify these mechanisms, we characterized GD25 cells expressing the beta(1) integrin cytoplasmic domain mutants W775A and R760A. Although cell spreading appears normal in R760A mutant-integrin cells compared with wild type, it is inhibited in W775A mutant cells. In contrast, both mutant cell lines show defective fibronectin matrix assembly. Inhibition of cell spreading, but not matrix assembly, in the W775A mutant cells is due to a specific defect in Akt-1 activation. In addition, we find that both W775A and R760A mutant integrins have reduced surface expression of the 9EG7 epitope that correlates with reduced recruitment of talin to beta(1) integrin cytoplasmic complexes. Down-regulation of talin with small interfering RNA or expression of green fluorescent protein-talin head domain inhibits matrix assembly in beta(1) wild-type cells, mimicking the defect seen with the W775A and R760A mutant cells. These results demonstrate distinct mechanisms by which integrins regulate cell spreading and matrix assembly through the beta(1) integrin cytoplasmic tail.
Green
J Angelo
JA
Laboratory of Cell and Developmental Biology, NIDCR, National Institutes of Health, Bethesda, Maryland 20892, USA. angelo.green@nih.gov
Berrier
Allison L
AL
Pankov
Roumen
R
Yamada
Kenneth M
KM
D019943:Amino Acid Substitution; D000818:Animals; D002460:Cell Line; D004789:Enzyme Activation; D005109:Extracellular Matrix / Q000235:genetics / Q000378:metabolism*; D005353:Fibronectins / Q000235:genetics / Q000378:metabolism*; D019012:Integrin beta1 / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D020125:Mutation, Missense; D017434:Protein Structure, Tertiary / Q000502:physiology; D051057:Proto-Oncogene Proteins c-akt / Q000235:genetics / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000235:genetics; D016608:Talin / Q000235:genetics / Q000378:metabolism*
D005353:Fibronectins; D019012:Integrin beta1; D034741:RNA, Small Interfering; D016608:Talin; C514862:Akt1 protein, mouse; D051057:Proto-Oncogene Proteins c-akt