Talin is an essential cytoskeletal protein present in cell-matrix type adherens junctions where it helps in attachment of the actin cytoskeleton to the plasma membrane. The nature of the interaction between talin and actin was examined in this dissertation by investigating three actin binding sites (ABS1-3), with ABS1 located in the 47 kDa N-terminal head and ABS2 and ABS3 present in the 190 kDA C-terminal tail of the talin molecule. Cosedimentation of expressed GST-fusion deletion constructs from the talin head with F-actin showed that actin binding activity of the talin head is contained within residues 271-320. Talin constructs containing the narrowed ABS1 exhibited less pH dependence compared to those containing ABS3. Both ABS1 and ABS3 showed high dependence on ionic strength in ability to bind actin. Nearly all of the N-terminal head fragment purified from calpain-cleaved talin cosedimented with F-actin under conditions (pH 6.4 and low ionic strength) found to be optimal for the talin-actin interaction. EGFP-fusion talin constructs containing ABS1 were primarily colocalized with actin stress fibers when transfected into COS cells, whereas EGFP-ABS3 was predominantly colocalized with actin-rich membrane ruffles. The ABS2 and ABS3 within the C-terminal tail were further defined to residues 1051-1250 (ABS2) and 2331-2530 (ABS3), respectively. ABS2 exhibited high dependence on both pH and ionic strength in interacting with actin, which is similar to that found with ABS3. Individual transfections of EFGP-ABS2 and EGFP-ABS3 into COS cells showed that ABS2 was primarily colocalized with actin stress fibers, whereas ABS3 was colocalized with actin filaments at both membrane ruffles and microspike-like cell extensions. Site-directed mutagenesis studies on ABS1 showed that substitution of alanine for the five positively charged residues K295, R297, K300, R303 and K306, or for all six residues within 295-300 resulted in significant reduction in actin binding. Taken together, these results suggest that the ABS1 located within the head domain, and the ABS2 and ABS3 present in the C-terminal tail domain, of talin each may play distinct roles in the interaction of talin with actin. These differences may be important in regulating integrin-mediated cell adhesion and migration.

Actin is one of the most widespread proteins in eukaryotic cells. This book and its companion (Molecular Interactions of Actin. Actin-Myosin Interaction, Actin-Based Regulation) provide an authoritative and opinionated view of the structure and function of this essential protein. Each section includes an historical perspective and a detailed commentary on actin protein chemistry, molecular and cell biology of actin. While some chapters review the body of knowledge of the subject, others contain new experimental data. This book will appeal to research scientists seeking contemporary overviews of actin and its binding proteins. Contributors include senior scientists as well as the new breed of younger scientists.

Actin is one of the most widespread proteins in eukaryotic cells. This book and its companion ("Molecular Interactions of Actin. Actin Structure and Actin-Binding Proteins") provide an authoritative and opinionated view of the structure and function of this essential protein. Each section includes an historical perspective and a detailed commentary on actin protein chemistry, molecular and cell biology of actin. While some chapters review the body of knowledge of the subject, others contain new experimental data. This book will appeal to research scientists seeking contemporary overviews of actin-myosin interaction and actin-based regulation. Contributors include senior scientists as well as the new breed of younger scientists.

Actin is one of the most widespread proteins in eukaryotic cells. This book and its companion (Molecular Interactions of Actin. Actin-Myosin Interaction, Actin-Based Regulation) provide an authoritative and opinionated view of the structure and function of this essential protein. Each section includes an historical perspective and a detailed commentary on actin protein chemistry, molecular and cell biology of actin. While some chapters review the body of knowledge of the subject, others contain new experimental data. This book will appeal to research scientists seeking contemporary overviews of actin and its binding proteins. Contributors include senior scientists as well as the new breed of younger scientists.

Since the discovery of actin by Straub in the 1950’s and the pioneering work of Oosawa on actin self-assembly in helical laments in the 1960’s, many books and conference proceedings have been published. As one of the most essential p- teins in life, essential for movement in organisms rangingfrom bacteria to higher eukaryotes, it is no surprise that actin has fascinated generations of scientists from many different elds. Actin can be considered as a “living treasure” of biology; the kinetics and thermodynamics of self-assembly, the dissipative nature of actin po- merization, the molecular interactions of monomeric and polymerized actin with regulators, the mechanical properties of actin gels, and more recently the force p- ducing motile and morphogenetic processes organized by the actin nanomachine in response to signaling, are all milestones in actin research. Discoveries that directly derive from and provide deeper insight into the fundamental properties of actin are constantly being made, making actin an ever appealing research molecule. At the same time, the explosion in new technologies and techniques in biological sciences has served to attract researchers from an expanding number of disciplines, to study actin. This book presents the latest developments of these new multiscale approaches of force and movement powered by self-assembly processes, with the hope to opening our perspectives on the many areas of actin-based motility research.