Interactive sequences in the stress protein and molecular chaperone human αB crystallin recognize and modulate the assembly of filaments

Molecular chaperones including the small heat shock proteins, αB crystallin and sHSP27 participate in the assembly, disassembly, and reorganization of the cytoskeleton during cell development and differentiation. While αB crystallin and sHSP27 stabilize and modulate filament assembly and re-organization, the sequences and structural domains mediating interactions between these proteins and filaments are unknown. It is important to define these interactive domains in order to understand differential interactions between chaperones and stable or unfolding filaments and their function in the cellular stress response. Protein pin arrays identified sequences in human αB crystallin that selectively interacted with native or partially unfolded filament proteins desmin, glial-fibrillary acidic protein, and actin. Circular dichroism spectroscopy determined differences in the structure of these filaments at 23 and 45 °C. Seven αB crystallin sequences had stronger interactions with desmin and six sequences had stronger interactions with glial-fibrillary acidic protein at 23 °C than at 45 °C. The αB crystallin sequences 33LESDLFPTSTSLSPFYLRPPSFLR56 and 129DPLTITSSLSSDGV145 had the strongest interactions with actin at 23 °C, while 57APSWFDTG64, 111HGFISREF118, 145VNGPRKQVSG154, and 155PERTIPITREEK165 had the strongest interactions with actin at 45 °C. The actin interactive sequences of αB crystallin overlapped with previously identified αB crystallin chaperone sequences and were synthesized to evaluate their effect on the assembly and aggregation of actin. Full-length αB crystallin and the core domain chaperone sequence 131LTITSSLSSDGV143 promoted actin polymerization at 37 °C and inhibited depolymerization and aggregation at 50 °C. The results support the hypothesis that interactive domains in αB crystallin have multiple functions in stabilizing the cytoskeleton and protecting cytosolic proteins from unfolding.