Structural determinants of microtubule minus end preference in CAMSAP CKK domains

CAMSAP/Patronins regulate microtubule minus-end dynamics. Their end specificity is mediated by their CKK domains, which we proposed recognise specific tubulin conformations found at minus ends. To critically test this idea, we compared the human CAMSAP1 CKK domain (HsCKK) with a CKK domain from Naegleria gruberi (NgCKK), which lacks minus-end specificity. Here we report near-atomic cryo-electron microscopy structures of HsCKK- and NgCKK-microtubule complexes, which show that these CKK domains share the same protein fold, bind at the intradimer interprotofilament tubulin junction, but exhibit different footprints on microtubules. NMR experiments show that both HsCKK and NgCKK are remarkably rigid. However, whereas NgCKK binding does not alter the microtubule architecture, HsCKK remodels its microtubule interaction site and changes the underlying polymer structure because the tubulin lattice conformation is not optimal for its binding. Thus, in contrast to many MAPs, the HsCKK domain can differentiate subtly specific tubulin conformations to enable microtubule minus-end recognition

PROTECTIVE EFFECT OF A PHYLLANTHUS EMBLICA EXTRACT AGAINST CISPLATIN-INDUCED APOPTOSIS IN DERMAL PAPILLA FIBROBLASTS

Cisplatin is a widely prescribed anticancer agent that causes hair loss in patients. Since the dermal papilla (DP) fibroblasts are known as a key mediator in controlling hair growth and loss, protection against cisplatin-induced cell damage on these cells may lead to a new strategy for hair loss protection in chemotherapy patients. We herein reported that cisplatin induced DP cell death in a concentration-dependent manner through apoptosis mechanism, which was further found to be related to the intracellular reactive oxygen species (ROS) generation. Therefore, extract from emblica fruits (Phyllanthus emblica), a known natural antioxidant, was examined for its protective effect on cisplatin-induced DP fibroblast cell death. Our results indicated that emblica extract attenuated the intracellular ROS induction following cisplatin treatment and subsequently protected against cisplatin-induced DP fibroblast apoptotic cell death. Co-treatment of emblica extract (250-500 µg/mL) and cisplatin 100 µmol/L (LD50) markedly increased the DP fibroblast relative cell viability to the same level as the non-treated control

CURCUMIN INDUCES ANOIKIS IN H460 NON-SMALL LUNG CANCER CELLS THROUGH SUPEROXIDE ANION INDUCTION

Anoikis or apoptosis triggered by loss of cell anchorage plays an important role in prevention of tumor cell metastasis. During metastasis process, anticancer drugs are not able to completely eliminate cancer cells that suspended in the blood stream; therefore, the use of natural safety compounds to enhance apoptosis of these suspended cells may improve the therapy. Curcumin (diferuloylmethane), a major active component in turmeric, Curcuma longa, has been shown to inhibit neoplastic initiation, promotion and progression of a wide variety of tumor cells. However, the effect of curcumin on anoikis process has not been well characterized. We herein demonstrated that 10 µM of curcumin triggered 1.2-folds anoikis of detached lung carcinoma H460 cells compared with controlled cells. Cell viability was detected by XTT assay and anoikis cells were indicated by annexin V staning assay. Mechanism of curcumin induced anoikis was associated with its ability to generate intracellular reactive oxygen species (ROS). Curcumin caused 1.6-folds induction of intracellular ROS level as detected by DCF-DA and flow cytometry. Our inhibitory study revealed that superoxide anion generated by curcumin was a principle ROS responsible for anoikis induction in these cells. Thus, our results showed that curcumin induced anoikis in detached-H460 cells resulted from increasing of intracellular ROS

EMBLICA EXTRACT INDUCES TYPE I PRO-COLLAGEN SYNTHESIS AND INHIBITS COLLAGENASE ACTIVITY IN MOUSE FIBROBLASTS

Decrease of type I collagen generation as well as the increase of its degradation via matrix metalloproteinases (MMPs; e.g. gelatinases and collagenases) are considered as a major cause of skin aging. A potent natural antioxidant exblica extract from emblica fruits (Phyllanthus emblica) was herein evaluated for collagen promoting activity in mouse fibroblasts. We demonstrated that the level of intracellular type I pro-collagen increased following emblica extract treatment in concentration-dependent manner. Emblica extract at the concentration of 100 µg/mL exhibited the maximum effect on type I pro-collagen; approximately 7 fold-induction versus non-treated control determined by Western blot analysis. Moreover, our result indicated that emblica extract affected the MMPs function. Percentage of collagenase inhibition exceeded forty at the concentration of 100 µg/mL of emblica extract. Taken together, emblica extract has a promising anti-aging effect attributed to its positive effect on type I collagen formation as well as the protection against collagen degradation

SENSITIZING EFFECT OF CURCUMIN ON CISPLATIN-INDUCED APOPTOSIS INVOLVES SUPEROXIDEANION INDUCTION AND BCL-2 DEGRADATION

The possibility of using curcumin as a chemotherapeutic sensitizing agent has been intensively demonstrated in some cancers. However, the effect of curcumin on non-small cell lung cancer (NSCLC), one of the most resistant cancers, is largely unknown. The aim of this study was to investigate the sensitizing effect of curcumin on cisplatin-induced apoptosis in NSCLC cells. Curcumin was shown to induce intracellular superoxide anion generation, down-regulate anti-apoptotic Bcl-2 protein, and subsequently sensitize NSCLC H460 cells to cisplatin-induced apoptosis. Amplification and overexpression of bcl-2 protein has been implicated in chemotherapeutic resistance in many cancers and overexpression of this protein strongly rendered H-460 cells resistant to cisplatin-induced apoptosis. The present study showed that co-treatment of the cells with curcumin and cisplatin resulted in increased apoptosis and reversal of Bcl-2-mediated cisplatin resistance. The mechanism by which curcumin down-regulates Bcl-2 and sensitizes cells to cisplatin-induced apoptosis involves proteasomal degradation of Bcl-2, since a specific proteasome inhibitor lactacystin reversed effect of curcumin on bcl-2 level. These findings indicate a novel pathway for curcumin regulation of Bcl-2, which benefits the development of a cisplatin sensitizing agent