Dynamic Localization Of Alpha B-Crystallin At The Microtubule Cytoskeleton Network In Beating Heart Cells

alpha B-crystallin is highly expressed in the heart and slow skeletal muscle; however, the roles of alpha B-crystallin in the muscle are obscure. Previously, we showed that alpha B-crystallin localizes at the sarcomere Z-bands, corresponding to the focal adhesions of cultured cells. In myoblast cells, alpha B-crystallin completely colocalizes with microtubules and maintains cell shape and adhesion. In this study, we show that in beating cardiomyocytes alpha-tubulin and alpha B-crystallin colocalize at the I- and Z-bands of the myocardium, where it may function as a molecular chaperone for tubulin/microtubules. Fluorescence recovery after photobleaching (FRAP) analysis revealed that the striated patterns of GFP-alpha B-crystallin fluorescence recovered quickly at 37 degrees C. FRAP mobility assay also showed alpha B-crystallin to be associated with nocodazole-treated free tubulin dimers but not with taxol-treated microtubules. The interaction of alpha B-crystallin and free tubulin was further confirmed by immunoprecipitation and microtubule sedimentation assay in the presence of 1-100 mu M calcium, which destabilizes microtubules. Forster resonance energy transfer analysis showed that alpha B-crystallin and tubulin were at 1-10 nm apart from each other in the presence of colchicine. These results suggested that alpha B-crystallin may play an essential role in microtubule dynamics by maintaining free tubulin in striated muscles, such as the soleus or cardiac muscles.