Cell spreading, adhesion and remodelling of the extracellular matrix (ECM) involve bi-directional signalling and physical linkages between the ECM, integrins and the cell cytoskeleton. The actin-binding proteins talin1 and 2 link ligand-bound integrins to the actin cytoskeleton and increase the affinity of integrin for the ECM. Here we report that depletion of talin2 in talin1-null (talin1(-/-)) cells did not affect the initiation of matrix-activated spreading or Src family kinase (SFK) activation, but abolished the ECM-integrin-cytoskeleton linkage and sustained cell spreading and adhesion. Specifically, focal adhesion assembly, focal adhesion kinase (FAK) signalling and traction force generation on substrates were severely affected. The talin1 head domain restored beta1 integrin activation but only full-length talin1 restored the ECM-cytoskeleton linkage and normal cytoskeleton organization. Our results demonstrate three biochemically distinct steps in fibronectin-activated cell spreading and adhesion: (1) fibronectin-integrin binding and initiation of spreading, (2) fast cell spreading and (3) focal adhesion formation and substrate traction. We suggest that talin is not required for initial cell spreading. However, talin provides the important mechanical linkage between ligand-bound integrins and the actin cytoskeleton required to catalyse focal adhesion-dependent pathways.