Interactive Domains in the Molecular Chaperone Human αB Crystallin Modulate Microtubule Assembly and Disassembly

Background. Small heat shock proteins regulate microtubule assembly during cell proliferation and in response to stress through interactions that are poorly understood. Methodology. Novel functions for five interactive sequences in the small heat shock protein and molecular chaperone, human alpha B crystallin, were investigated in the assembly/disassembly of microtubules and aggregation of tubulin using synthetic peptides and mutants of human alpha B crystallin. Principal Findings. The interactive sequence 113FISREFHR120 exposed on the surface of alpha B crystallin decreased microtubule assembly by similar to 45%. In contrast, the interactive sequences, (131)LTITSSLSSDGV(142) and 156ERTIPITRE164, corresponding to the beta 8 strand and the C-terminal extension respectively, which are involved in complex formation, increased microtubule assembly by similar to 34-45%. The alpha B crystallin peptides, 113FISREFHR120 and 156ERTIPITRE164, inhibited microtubule disassembly by similar to 26-36%, and the peptides 113FISREFHR120 and (131)LTITSSLSSDGV(142) decreased the thermal aggregation of tubulin by similar to 42-44%. The (131)LTITSSLSSDGV(142) and 156ERTIPITRE164 peptides were more effective than the widely used anti-cancer drug, Paclitaxel, in modulating tubulin <-> microtubule dynamics. Mutagenesis of these interactive sequences in wt human alpha B crystallin confirmed the effects of the alpha B crystallin peptides on microtubule assembly/disassembly and tubulin aggregation. The regulation of microtubule assembly by alpha B crystallin varied over a narrow range of concentrations. The assembly of microtubules was maximal at alpha B crystallin to tubulin molar ratios between 1:4 and 2:1, while molar ratios >2:1 inhibited microtubule assembly. Conclusions and Significance. Interactive sequences on the surface of human alpha B crystallin collectively modulate microtubule assembly through a dynamic subunit exchange mechanism that depends on the concentration and ratio of alpha B crystallin to tubulin. These are the first experimental results in support of the functional importance of the dynamic subunit model of small heat shock proteins.