SummaryCytokinesis is initiated by constriction of the cleavage furrow, and completed with separation of the two daughter cells by abscission. Control of transition from constriction to abscission is therefore crucial for cytokinesis. However, the underlying mechanism is largely unknown. Here, we analyze the role of Citron kinase (Citron-K) that localizes at the cleavage furrow and the midbody, and dissect its action mechanisms during this transition. Citron-K forms a stable ring-like structure at the midbody and its depletion affects the maintenance of the intercellular bridge, resulting in fusion of two daughter cells after the cleavage furrow ingression. RNA interference (RNAi) targeting Citron-K reduced accumulation of RhoA, Anillin, and septins at the intercellular bridge in mid telophase, and impaired concentration and maintenance of KIF14 and PRC1 at the midbody in late telophase. RNAi rescue experiments revealed that these functions of Citron-K are mediated by its coiled-coil (CC) domain, and not by its kinase domain. The C-terminal part of CC contains a Rho-binding domain and a cluster-forming region and is important for concentrating Citron-K from the cleavage furrow to the midbody. The N-terminal part of CC directly binds to KIF14, and this interaction is required for timely transfer of Citron-K to the midbody after furrow ingression. We propose that the CC-domain-mediated translocation and actions of Citron-K ensure proper stabilization of the midbody structure during the transition from constriction to abscission.

The small GTPase Rho and mDia2, a Rho-regulated actin nucleator, function as critical regulators of cytokinesis in cultured cells. However, their involvement in cytokinesis during mammalian development remains unknown. Here, we generated mice deficient in mDia2 and examined the role of Rho signaling in cytokinesis during development. mDia2-deficient mice survive until embryonic day 11.5 (E11.5), exhibit severe anemia with multinucleate erythroblasts, and die in utero by E12.5. mDia2-deficient erythroid cells differentiate normally, though in a delayed manner, but exhibit cytokinesis failure with decreased accumulation of F-actin in the cleavage furrow during late differentiation from proerythroblasts. On the other hand, inactivation of Rho induces cytokinesis failure from the earlier progenitor stage. mDia2-deficient erythroblasts, however, are able to enucleate their nuclei. Our findings have thus revealed that mDia2 functions critically in cytokinesis in vivo during erythropoiesis and further suggest that the cytokinesis mechanism in development diverges downstream of Rho. They also demonstrate that cytokinesis and enucleation utilize different mechanisms.

The major obstacle to successful chemotherapy of cancer patients is drug resistance. Previously we explored the molecular mechanisms of curcumin cross-resistance in carboplatin resistant human laryngeal carcinoma 7T cells. Following curcumin treatment we found a reduction in curcumin accumulation, and reduced induction of reactive oxygen species (ROS) and their downstream effects, compared to parental HEp-2 cells. In order to shed more light on mechanisms involved in drug resistance of 7T cells, in the present study we applied Fourier transform infrared (FTIR) spectroscopy, a technique that provides information about the nature and quantities of all molecules present in the cell. By comparing the spectra from parental HEp-2 cells and their 7T subline, we found an increase in the intensity of ester vibrational bands in 7T cells. This implied an increase in the amount of cholesteryl esters in resistant cells, which we confirmed by an enzymatic assay. Since cholesteryl esters are localized in lipid droplets, we confirmed their higher quantity and serum dependency in 7T cells compared to HEp-2 cells. Moreover, treatment with oleic acid induced more lipid droplets in 7T when compared to HEp-2 cells, as shown by flow cytometry. We can conclude that along with previously determined molecular mechanisms of curcumin resistance in 7T cells, these cells exhibit an increased content of cholesteryl esters and lipid droplets, suggesting an alteration in cellular lipid metabolism as a possible additional mechanism of drug resistance. Furthermore, our results suggest the use of FTIR spectroscopy as a promising technique in drug resistance research.

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