A subpopulation of membrane tubulin is associated with Na+,K+-ATPase in neural and non-neural cells. This association both confers hydrophobic behavior to tubulin and inhibits the Na+,K+-ATPase activity. Treatment of cells with 1mM of L-glutamate provokes dissociation of the complex, leading to increment of active enzyme. The acetylated form of tubulin is present in the tubulin fraction associated with Na+,K+-ATPase, however we do not know whether this type of tubulin is indispensable for interaction with the enzyme. To investigate this question, we used 6-11B-1 antibody specific to acetyl group bound to Lys at position 40 of the ¥á chain of tubulin, and CAD cells, which lacks acetylated tubulin. In these cells, L-glutamate was unable to stimulate the Na+,K+-ATPase. Using immunoprecipitation procedure we showed that the Na+,K+-ATPase/tubulin complex was absent in CAD cells. Treatment of cells with deacetylases inhibitors (TSA and tubacin), led to appearance of a significant amount of acetylated tubulin. Under these conditions, the Na+,K+-ATPase/tubulin complex was found in membranes, and Na+,K+-ATPase activity was inhibited. In addition L-glutamate was now able to d¨¬¨¬¨¬issociate the complex and to increase the enzyme activity. These results indicates that tubulin must be acetylated at Lys 40 to interact with the plasma membrane Na+,K+-ATPase.
