Talin, a protein localized primarily in cell-matrix type adherens junctions, is involved in linking the cytoskeleton to the cell membrane. The major objectives of my study were to (1) determine the structural basis for the effect of pH and ionic strength on the direct talin-actin interaction, (2) examine the effects of other factors, such as specific phospholipids, on the talin-actin interaction, and (3) compare abilities of human platelet and avian smooth muscle talins to interact with actin. Negative staining results showed that at pH 6.4 and low ionic strength, talin extensively crosslinked actin filaments into both tight networks and bundles. Some of the bundles consisted of parallel actin filaments with an interfilament spacing of ~13 nm, and talin crossbridges spaced at ~36 nm intervals along the actin bundles. As pH and/or ionic strength was increased, talin's actin bundling activity was decreased first, then its networking activity. Chemical crosslinking indicated that talin was present primarily as a dimer of two ~225 kDa subunits under all ionic conditions tested. Talin crosslinked actin filaments into networks and bundles after only five minutes of actin polymerization, with no evidence of shorter actin filaments formed in comparison to actin controls. Talin also crosslinked preformed actin filaments as well as it did actin filaments polymerized in its presence. The ~190 kDa talin fragment was able to crosslink actin filaments into networks and bundles, but required a slightly lower pH than did intact talin;The phosphoinositides, PIP2, PIP, or PI, but not IP3 or the phosphoglycerides, PS or PC, greatly inhibited the ability of talin to crosslink actin filaments. The phosphoinositides bound talin to form talin/phosphoinositide complexes, which prevented talin from effectively interacting with actin filaments. The detergent Triton X-100 or divalent cations, such as Ca2+, decreased the effect of the phosphoinositides on the talin-actin interaction;Cosedimentation assays, low shear viscometry, and negative staining showed that platelet talin interacted with actin in a fashion similar to that of smooth muscle talin. However, some differences were noted between the two talins, with platelet talin exhibiting lower actin crosslinking activity.