The mitotic tensegrity guardian tau protects mammary epithelia from katanin-like1-induced aneuploidy

Abstract

The microtubule associated-protein tau has been identified as an effective positive prognostic indicator in breast cancer. To explore the physiological function of tau in early carcinogenesis, endogenous tau was knocked down in primary cultured human mammary epithelial cells. This resulted in chromosome-bridging during anaphase followed by micronucleation, both of which were suppressed by a further katanin-like1 knockdown. We also detected that the exogenously expressed katanin-like1 induction of cellular transformation is prevented by exogenous tau in rat fibroblasts. The mutant katanin-like1 (L123V) identified in breast cancer showed an increase in this transformation capacity as well as microtubule severing activity resistant to tau. The tau knockdown resulted in a loss of the kinetochore fibers on which tau is normally localized. This physical fragility was also observed in isolated tau-knockdown mitotic spindles, supporting the relevance of microtubule damage to the onset of transformation. The karyotyping of tau-knockdown cells showed increased frequency of loss of one X chromosome, further suggesting the involvement of tau in breast tumorigenesis. We propose that tau may contribute to tumor progression by protecting spindle microtubules from excess severing by katanin-like1. We also present data indicating that the microtubule-binding octapeptide NAP is a candidate modifier against the tau deficiency in tumor cells.

Microtubule Hyperacetylation Enhances KL1-Dependent Micronucleation under a Tau Deficiency in Mammary Epithelial Cells

“…The rat fibroblast (RFL)-6, a cell line previously employed in studies of microtubule-severing proteins [ 20 , 21 , 24 , 25 , 26 ] was used in the present analyses. Fibroblasts are useful for high-resolution imaging of microtubules because they have a flat morphology.…”

“…Fibroblasts are useful for high-resolution imaging of microtubules because they have a flat morphology. In addition, they do not endogenously express tau which would lower the sensitivity of the microtubules to severing [ 20 , 24 , 26 ].…”

“…Katanin was included, the enhanced activity of which under tubacin treatment was observed in our previous study [ 21 ]. Cells were sorted with regard to the katanin or KL1-expression levels, and a focus was placed on medium expressors, as we did previously [ 20 ]. Representative images are shown in Figure 1 B with the quantitative data on the microtubule levels indicated in the graph.…”

“…Our previous study has suggested that KL1, one of the microtubule severing proteins, is abnormally up-regulated in early breast carcinogenesis when tau is decreased and contributes to tumor progression by inducing abnormal karyotypes that are relevant to the basal cell subtype [ 20 ]. Regarding the perturbation of the regulatory mechanisms underlying microtubule severing, we have also reported that microtubule hyperacetylation enhances the activity of katanin but not spastin [ 21 ].…”

“…The mitotic spindle and micronucleation were analyzed in response to microtubule hyperacetylation in the same cells. The micronucleation/microtubule acetylation relationship was further explored in normal human mammary epithelial cells (HMECs), which express endogenous tau, and in which the loss of tau has been suggested to promote early stage breast carcinogenesis [ 20 ]. It was observed that only under an imbalance between tau and KL1 did increased microtubule acetylation enhance abnormal chromosome segregation and micronucleation.…”