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Analysis of Fyn function in hemostasis and alphaIIbbeta3-integrin signaling.

Abstract：

:Recent studies have shown that Src-family kinases (SFKs) play an important role in mediating integrin signalling, and the beta3 subunit of alphaIIbbeta3 integrin has been shown to interact with multiple SFK members. Here, we analyzed the interactions and functional consequences of Fyn and Src binding to alphaIIbbeta3. Fyn associated with the beta3 subunit in resting and thrombin-aggregated platelets, whereas interaction between Src and alphaIIbbeta3 was seen predominantly in resting but not in thrombin-aggregated platelets. We have also observed that Fyn but not Src localized to focal adhesions in CHO cells adherent to fibrinogen through alphaIIbbeta3. On the basis of these differences, we wanted to determine the sequence requirements for the interaction of Fyn and Src within the beta3-cytoplasmic domain. Whereas Src association required the C-terminal region of beta3, Fyn continued to interact with mutants that could no longer associate with Src and that contained as few as 13 membrane-proximal amino acids of the beta3-cytoplasmic tail. Using deletion mutants of beta3-cytoplasmic tails expressed as GST-fusion proteins, we narrowed down the Fyn-binding site even further to the amino acid residues 721-725 (IHDRK) of the beta3-cytoplasmic domain. On the basis of these observations, we explored whether Fyn-/- mice exhibited any abnormalities in hemostasis and platelet function. We found that Fyn-/- mice significantly differed in their second bleeding times compared with wild-type mice, and platelets from Fyn-/- mice exhibited delayed spreading on fibrinogen-coated surfaces. Using mutant forms of Fyn, it appears that its kinase activity is required for its localization to focal adhesions and to mediate alphaIIbbeta3-dependent cell spreading. Our results suggest that Fyn and Src have distinct requirements for interaction with alphaIIbbeta3; and, consequently, the two SFK can mediate different functional responses.

journal_name

J Cell Sci

journal_title

Journal of cell science

authors

Reddy KB,Smith DM,Plow EF

doi

10.1242/jcs.014076

subject

Has Abstract

pub_date

2008-05-15 00:00:00

pages

1641-8

issue

Pt 10

eissn

0021-9533

issn

1477-9137

pii

jcs.014076

journal_volume

121

pub_type

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