The central spindle drives anaphase chromosome segregation.

Anaphase chromosome segregation depends on forces exerted by spindle microtubules. In the current model, forces on chromosomes are mediated through the spindle poles: sliding of antiparallel microtubules in the central spindle pushes poles apart, while kinetochore microtubule (kMT) depolymerization pulls chromosomes towards the poles. Here we show that the central spindle is directly linked to the chromosomes rather than the poles in anaphase, based on three lines of evidence. Chromosomes in monopolar spindles can move away from the pole, consistent with forces exerted by antiparallel microtubule sliding. In bipolar spindles, kMT depolymerization is constrained by suppressing central spindle sliding, indicating kinetochore linkage to the central spindle. Finally, increasing the rate of kMT depolymerization slows pole separation without increasing chromosome separation velocity. We conclude that central spindle sliding drives anaphase chromosome separation, while kMT depolymerization limits spindle elongation.

Conditional Localization Pharmacology Manipulates the Cell Cycle with Spatiotemporal Precision.

Traditional pharmacology has limited control of drug activity and localization in space and time. Herein, we described an approach for kinase regulation using conditional localization pharmacology (CLP), where an inactive caged inhibitor is localized to a site of interest in a dormant state using intracellular protein tethering. The activity of the inhibitor can be regulated with spatial and temporal precision in a live cellular environment using light. As a proof of concept, a photocaged MPS1 kinase inhibitor (reversine) bearing a Halo-tag ligand tether was designed to manipulate the cell cycle. We demonstrate that this new caged reversine halo probe (CRH) strategy is capable of efficient localization and exceptional spatiotemporal control over spindle assembly checkpoint (SAC) silencing and mitotic exit.

"Off-pore" nucleoporins relocalize heterochromatic breaks through phase separation.

Phase separation forms membraneless compartments in the nuclei, including by establishing heterochromatin "domains" and repair foci. Pericentromeric heterochromatin mostly comprises repeated sequences prone to aberrant recombination, and "safe" homologous recombination (HR) repair of these sequences requires the movement of repair sites to the nuclear periphery before Rad51 recruitment and strand invasion. How this mobilization initiates is unknown, and the contribution of phase separation to these dynamics is unclear. Here, we show that Nup98 nucleoporin is recruited to heterochromatic repair sites before relocalization through Sec13 or Nup88 nucleoporins, and downstream from the Smc5/6 complex and SUMOylation. Remarkably, the phase separation properties of Nup98 are required and sufficient to mobilize repair sites and exclude Rad51, thus preventing aberrant recombination while promoting HR repair. Disrupting this pathway results in heterochromatin repair defects and widespread chromosome rearrangements, revealing a novel "off-pore" role for nucleoporins and phase separation in nuclear dynamics and genome integrity in a multicellular eukaryote.

An ultrasensitive GSH-specific fluorescent probe unveils celastrol-induced ccRCC ferroptosis.

Glutathione (GSH) is closely related to the occurrence and development of tumors. The intracellular GSH levels are abnormally altered when tumor cells undergo programmed cell death. Therefore, real-time monitoring of the dynamic changes of intracellular GSH levels can better enable the early diagnosis of diseases and evaluate the effects of cell death-inducing drugs. In this study, a stable and highly selective fluorescent probe AR has been designed and synthesized for the fluorescence imaging and rapid detection of GSH in vitro and in vivo, as well as patient-derived tumor tissue. More importantly, the AR probe can be used to track changes in GSH levels and fluorescence imaging during the treatment of clear cell renal cell carcinoma (ccRCC) with celastrol (CeT) via inducing ferroptosis. These findings demonstrate that the developed fluorescent probe AR exhibits high selectivity and sensitivity, as well as good biocompatibility and long-term stability, which can be used to image endogenous GSH in living tumors and cells. Also, a significant decrease in GSH levels was observed by the fluorescent probe AR during the treatment of ccRCC with CeT-induced ferroptosis in vitro and in vivo. Overall, these findings will provide a novel strategy for celastrol targeting ferroptosis in the treatment of ccRCC and the application of fluorescent probes to help reveal the underlying mechanism of CeT in the treatment of ccRCC.

GSH-specific fluorescent probe for sensing, bioimaging, rapid screening of natural inhibitor Celastrol and ccRCC theranostics.

High level of intracellular glutathione (GSH) has been identified as a major barrier for cancer therapy. Therefore, effective regulation of GSH can be regarded as a novel approach for cancer therapy. In this study, an off-on fluorescent probe (NBD-P) is developed for selective and sensitive sensing GSH. NBD-P has a good cell membrane permeability that can be applied in bioimaging endogenous GSH in living cells. Moreover, the NBD-P probe is used to visualize GSH in animal models. In addition, a rapid drug screening method is successfully established using the fluorescent probe NBD-P. A potent natural inhibitor of GSH is identified as Celastrol from Tripterygium wilfordii Hook F, which effectively triggers mitochondrial apoptosis in clear cell renal cell carcinoma (ccRCC). More importantly, NBD-P can selectively respond to GSH fluctuations to distinguish cancer tissues from normal tissues. Thus, the present study provides insights into fluorescence probes for the screening GSH inhibitors and cancer diagnosis, as well as in-depth exploration of the anti-cancer effects of Traditional Chinese Medicine (TCM).

Involvement of LDL and ox-LDL in Cancer Development and Its Therapeutical Potential.

Lipid metabolism disorder is related to an increased risk of tumorigenesis and is involved in the rapid growth of cancer cells as well as the formation of metastatic lesions. Epidemiological studies have demonstrated that low-density lipoprotein (LDL) and oxidized low-density lipoprotein (ox-LDL) are closely associated with breast cancer, colorectal cancer, pancreatic cancer, and other malignancies, suggesting that LDL and ox-LDL play important roles during the occurrence and development of cancers. LDL can deliver cholesterol into cancer cells after binding to LDL receptor (LDLR). Activation of PI3K/Akt/mTOR signaling pathway induces transcription of the sterol regulatory element-binding proteins (SREBPs), which subsequently promotes cholesterol uptake and synthesis to meet the demand of cancer cells. Ox-LDL binds to the lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) and cluster of differentiation 36 (CD36) to induce mutations, resulting in inflammation, cell proliferation, and metastasis of cancer. Classic lipid-lowering drugs, statins, have been shown to reduce LDL levels in certain types of cancer. As LDL and ox-LDL play complicated roles in cancers, the potential therapeutic effect of targeting lipid metabolism in cancer therapy warrants more investigation.

Celastrol ameliorates vascular neointimal hyperplasia through Wnt5a-involved autophagy.

Neointimal hyperplasia caused by the excessive proliferation of vascular smooth muscle cells (VSMCs) is the pathological basis of restenosis. However, there are few effective strategies to prevent restenosis. Celastrol, a pentacyclic triterpene, has been recently documented to be beneficial to certain cardiovascular diseases. Based on its significant effect on autophagy, we proposed that celastrol could attenuate restenosis through enhancing autophagy of VSMCs. In the present study, we found that celastrol effectively inhibited the intimal hyperplasia and hyperproliferation of VSMCs by inducing autophagy. It was revealed that autophagy promoted by celastrol could induce the lysosomal degradation of c-MYC, which might be a possible mechanism contributing to the reduction of VSMCs proliferation. The Wnt5a/PKC/mTOR signaling pathway was found to be an underlying mechanism for celastrol to induce autophagy and inhibit the VSMCs proliferation. These observations indicate that celastrol may be a novel drug with a great potential to prevent restenosis.

Targeting HDL in tumor microenvironment: New hope for cancer therapy.

Epidemiological studies have shown that plasma HDL-C levels are closely related to the risk of prostate cancer, breast cancer, and other malignancies. As one of the key carriers of cholesterol regulation, high-density lipoprotein (HDL) plays an important role in tumorigenesis and cancer development through anti-inflammation, antioxidation, immune-modulation, and mediating cholesterol transportation in cancer cells and noncancer cells. In addition, the occurrence and progression of cancer are closely related to the alteration of the tumor microenvironment (TME). Cancer cells synthesize and secrete a variety of cytokines and other factors to promote the reprogramming of surrounding cells and shape the microenvironment suitable for cancer survival. By analyzing the effect of HDL on the infiltrating immune cells in the TME, as well as the relationship between HDL and tumor-associated angiogenesis, it is suggested that a moderate increase in the level of HDL in vivo with consequent improvement of the function of HDL in the TME and induction of intracellular cholesterol efflux may be a promising strategy for cancer therapy.

Autophagy: a killer or guardian of vascular smooth muscle cells.

Vascular smooth muscle cells (VSMCs) is one of the main intracellular components of the blood vessel wall. The abnormalities of VSMCs participate in the development of cardiovascular diseases such as atherosclerosis, hypertension, and restenosis, especially the formation and stability of atherosclerotic plaques. Autophagy is involved in the regulation of proliferation, migration and phenotype switching of VSMCs, which in turn affects the pathological process of atherosclerosis. However, the autophagy of VSMCs has a dual effect on cells survival. Autophagy is induced in VSMCs by various stimuli such as 7-ketocholesterol (7-KC), unsaturated lipid peroxidation-derived aldehyde and excess free cholesterol, thereby promoting VSMCs survival and stabilising atherosclerotic plaque. Conversely, autophagy caused by factors such as osteopontin (OPN), angiotensin II (Ang II) and nicotine can accelerate the death of VSMCs, further accelerating atherosclerotic lesions. In addition, mitophagy and lipophagy as selective autophagy are also involved in the outcome of VSMCs as well as progression of atherosclerotic lesion. Currently, there are only a few drugs available to induce VSMCs autophagy, such as atorvastatin, telmisartan and so on. Due to the important role of VSMCs autophagy in the progression of atherosclerosis plaques, drugs that directly target autophagy of VSMCs are urgently needed to be developed.

Nickel-Catalyzed Decarboxylative Generation and Asymmetric Allylation of 2-Azaallyl Anions.

The first nickel-catalyzed asymmetric decarboxylative allylation (DcA) of allyl 2,2-diarylglycinate imines is reported. This transformation utilizes a chiral ferrocenyl bidentate ligand and a Ni(0) precatalyst to mediate the decarboxylative generation and asymmetric allylation of 2-azaallyl anions, affording α-aryl homoallylic imines in modest-to-high yields and moderate-to-high enantiomeric ratios. The resulting Ni-catalyzed transformation proved to be less general in comparison to our previously reported analogous Pd-mediated protocol, but it still exhibited certain advantages in regard to the regio- and enantioselectivity of the C-C bond formation.