Mutations in alpha- and beta-tubulin that stabilize microtubules and confer resistance to colcemid and XXXX .

@entity72722 @entity72719 @entity72724 @entity72720 @entity1855 @entity535 @entity72721 @entity1311 @entity16200 @entity72725 @entity72723 @entity72726 @entity3569 @entity5713
Single-step selections were used to obtain @entity3569 ovary cell lines resistant to Colcemid and @entity5713 . @entity1855 was included in the selections to circumvent the isolation of cells with P-glycoprotein-mediated multidrug resistance and thereby enrich for cells with tubulin alterations. The isolated cell lines were 2-fold resistant to the selecting drug, exhibited cross-resistance to other drugs that inhibit microtubule assembly, and had enhanced sensitivity to the microtubule-stabilizing drug @entity1311 . The concomitant resistance to microtubule-destabilizing drugs and enhanced sensitivity to @entity1311 suggested that these cell lines have changes in microtubule assembly. Consistent with this interpretation, drug-resistant cell lines exhibited altered alpha- or beta-tubulin mobility on two-dimensional gels and higher levels (47-54%) of assembled tubulin compared with wild-type (39%) or @entity1311 -resistant cells (25%). Some drug-resistant cells also had bundled microtubules as judged by immunofluorescence. Genomic sequencing of 11 drug-resistant cell lines predicted five different alterations ( @entity72719 , @entity72720 , @entity72721 , @entity72722 , and @entity72723 ) in beta-tubulin and four different alterations ( @entity72724 , @entity72725 , @entity72726 , and @entity16200 ) in alpha-tubulin. The @entity535 substitutions are dispersed on the primary and tertiary structures of tubulin and, together with the other mutant properties, argue against a mechanism involving changes in drug binding. Rather, we propose that the alterations in alpha- and beta-tubulin increase microtubule stability by promoting longitudinal interdimer and intradimer interactions and/or lateral interactions between protofilaments. This enhanced stability of microtubules increases their resistance to drugs that inhibit assembly.
