The interaction of titin and alpha-actinin is controlled by a phospholipid-regulated intramolecular pseudoligand mechanism.
The assembly of stable cytoskeletal structures from dynamically recycled molecules requires developmental and spatial regulation of protein interactions. In muscle, titin acts as a molecular ruler organizing the actin cytoskeleton via interactions with many sarcomeric proteins, including the crosslinking protein alpha-actinin. An interaction between the C-terminal domain of alpha-actinin and titin Z-repeat motifs targets alpha-actinin to the Z-disk. Here we investigate the cellular regulation of this interaction. alpha-actinin is a rod shaped head-to-tail homodimer. In contrast to C-terminal fragments, full-length alpha-actinin does not bind Z-repeats. We identify a 30-residue Z-repeat homologous sequence between the actin-binding and rod regions of alpha-actinin that binds the C-terminal domain with nanomolar affinity. Thus, Z-repeat binding is prevented by this 'pseudoligand' interaction between the subunits of the alpha-actinin dimer. This autoinhibition is relieved upon binding of the Z-disk lipid phosphatidylinositol-bisphosphate to the actin-binding domain. We suggest that this novel mechanism is relevant to control the site-specific interactions of alpha-actinin during sarcomere assembly and turnover. The intramolecular contacts defined here also constrain a structural model for intrasterical regulation of all alpha-actinin isoforms.
Young
P
P
European Molecular Biology Laboratory, Structural Biology Division, 69012 Heidelberg, Germany.
Gautel
M
M
D000185:Actinin / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D000831:Animals, Newborn; D001667:Binding, Competitive; D002151:Calorimetry; D002478:Cells, Cultured; D003001:Cloning, Molecular; D064211:Connectin; D018076:DNA, Complementary / Q000378:metabolism; D019281:Dimerization; D004591:Electrophoresis, Polyacrylamide Gel; D005455:Fluorescent Antibody Technique; D005982:Glutathione Transferase / Q000378:metabolism; D007136:Immunoglobulins / Q000737:chemistry; D008969:Molecular Sequence Data; D009124:Muscle Proteins / Q000378:metabolism*; D009206:Myocardium / Q000378:metabolism; D010710:Phosphates / Q000737:chemistry; D010716:Phosphatidylinositols / Q000737:chemistry; D010743:Phospholipids / Q000737:chemistry; D011485:Protein Binding; D020033:Protein Isoforms; D011494:Protein Kinases / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D051381:Rats; D011994:Recombinant Proteins / Q000378:metabolism; D012518:Sarcomeres / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013997:Time Factors; D020798:Two-Hybrid System Techniques
D064211:Connectin; D018076:DNA, Complementary; D007136:Immunoglobulins; D009124:Muscle Proteins; D010710:Phosphates; D010716:Phosphatidylinositols; D010743:Phospholipids; D020033:Protein Isoforms; D011994:Recombinant Proteins; D000185:Actinin; D005982:Glutathione Transferase; D011494:Protein Kinases