PT  - JOURNAL ARTICLE
AU  - Hari, Malathi
AU  - Wang, Yaqing
AU  - Veeraraghavan, Sudha
AU  - Cabral, Fernando
TI  - Mutations in α- and β-Tubulin That Stabilize Microtubules and Confer Resistance to Colcemid and Vinblastine&lt;a id=&quot;xref-fn-5-1&quot; class=&quot;xref-&quot; href=&quot;#fn-5&quot;&gt;&lt;sup&gt;1&lt;/sup&gt;&lt;/a&gt;
DP  - 2003 Jul 01
TA  - Molecular Cancer Therapeutics
PG  - 597--605
VI  - 2
IP  - 7
4099  - http://mct.aacrjournals.org/content/2/7/597.short
4100  - http://mct.aacrjournals.org/content/2/7/597.full
SO  - Mol Cancer Ther2003 Jul 01; 2
AB  - Single-step selections were used to obtain Chinese hamster ovary cell lines resistant to Colcemid and vinblastine. Verapamil 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 paclitaxel. The concomitant resistance to microtubule-destabilizing drugs and enhanced sensitivity to paclitaxel suggested that these cell lines have changes in microtubule assembly. Consistent with this interpretation, drug-resistant cell lines exhibited altered α- or β-tubulin mobility on two-dimensional gels and higher levels (47–54%) of assembled tubulin compared with wild-type (39%) or paclitaxel-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 (D45Y, C211F, D224N, S234N, and K350N) in β-tubulin and four different alterations (H283Y, E55K, A383V, and R390C) in α-tubulin. The amino acid 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 α- and β-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.