TALIN CONTAINS THREE ACTIN-BINDING SITES EACH OF WHICH IS ADJACENT TO A VINCULIN-BINDING SITE

摘要

We have determined the sequence of chicken talin (2,541 amino acids, M(r) 271,881) which is very similar (89% identity) to that of the mouse protein, Alignments with the Caenorhabditis elegans and Dictyostelium discoideum talin sequences show that the N- and C-terminal regions of the protein are conserved whereas the central part of the molecule is more divergent, By expressing overlapping talin polypeptides as fusion proteins, we have identified at least three regions of the protein which can bind F-actin: residues 102-497, 951-1,327 and 2,269-2,541. The N-terminal binding site contains a region with homology to the ERM family of actin-binding proteins, and the C-terminal site is homologous to the yeast actin-binding protein Sla2p, Each of the actin-binding sites is close to, but distinct from a binding site for vinculin, a protein which also binds actin, The Pro1176 to Thr substitution found in talin from Wistar-Furth rats does not destroy the capacity of this region of the protein to hind actin or vinculin, Microinjection studies showed that a fusion protein containing the N-terminal actin-binding site localised weakly to stress fibres, whereas one containing the C-terminal site initially localised predominantly to focal adhesions, The former was readily solubilised, and the latter was resistant to Triton extraction, The N-terminal talin polypeptide eventually disrupted actin stress fibres whereas the C-terminal polypeptide was without effect, However, a larger C-terminal fusion protein also containing a vinculin-binding site did disrupt stress fibres and focal adhesions, The results suggest that, although both the N- and C-terminal regions of talin bind actin, the properties of these two regions of the protein are distinct. [References: 46]