RESUMO
@# Objective：To investigate the role and mechanism of chromosomal region maintenance 1 (CRM1) inhibitor sulforaphene (LFS-01) in killing triple negative breast cancer (TNBC) cells by inhibiting signal transducer and activator of transcription 3 (STAT3) signaling pathways. Methods: Whether LFS-01 could combine with the NES pocket of CRM1 was verified by molecular dynamics simulation techniques. The killing activity of LFS-01 on four different breast cancer cell lines was detected by CCK-8 method. TNBC MDA-MB-468 and MDA-MB-231 cells were treated with different concentrations of LFS-01, and the intracellular localization of CRM1 cargo protein STAT3 and protein with NES sequence was detected by immunofluorescence; WB was used to detect the effect of LFS-01 on the expression of STAT-3 signaling pathway and its downstream proteins; WB, cellular immunofluorescence and transmission electron microscopy were adopted to detect the occurrence of autophagy; the effect of LFS-01 on cell cycle and apoptosis was detected by flow cytometry. Results: Molecular dynamics simulations showed that LFS-01 can bind to the NES pocket of CRM1, indicating that it may structurally affect the latter's protein transport function. LFS-01 could specifically kill TNBC MDA-MB-468 and MDA-MB-231 cells. STAT3 and NES-tagged proteins were mainly blocked in the nucleus of TNBC cells after the treatment with 10 μmol/L LFS-01, while they were evenly distributed in the cytoplasm in the control group. The expressions of phosphorylated STAT3 protein, Bcl-xL and Cylin D1 were decreased in MDA-MB-468 and MDA-MB-231 cells with the increase of LFS-01 dose and the prolongation of treatment time; the expression of autophagy marker protein LC3B increased, and highdensity, multi-layered autophagosomes appeared at the same time; cell cycle arrest was observed in S phase and apoptosis rate was significantly increased (P<0.05 or P<0.01). Conclusion: LFS-01 blocks the export of CRM1 carrier protein, thereby inhibiting the activation of STAT3 signaling pathway and promoting autophagy, cell cycle arrest and apoptosis in TNBC MDA-MB-468 and MDAMB-231 cells.
RESUMO
Mantle cell lymphoma (MCL) is an aggressive histotype of B-cell non-Hodgkin lymphoma. The disease has no known cure, which prompts the urgent need for novel therapeutic agents. Chromosomal region maintenance 1 (CRM1) may play a role in human neoplasia and serve as a novel target of cancer treatment. This study summarizes MCL pathogenesis and determines the involvement of CRM1 in the regulation of several vital signaling pathways contributing to MCL pathogenesis, including the pathways of cell cycle progression, DNA damage response, phosphoinositide kinase-3, nuclear factor-κB activation, and chromosomal stability. A preclinical study is also presented to compare the CRM1 status in MCL cell lines and primary MCL cells with normal B cells, as well as the therapeutic efficiency of CRM1 inhibition in MCL in vitro and in vivo, which make these agents potential targets of novel MCL treatments.
RESUMO
The function of the herpes simplex virus type 1(HSV-1)UL4 protein is still elusive. Our objective is to investigate the subcellular transport mechanism of the UL4 protein. In this study,fluorescence microscopy was employed to investigate the subcellular localization of UL4 and characterize the transport mechanism in living cells. By constructing a series of deletion mutants fused with enhanced yellow fluorescent protein(EYFP),the nuclear export signals(NES)of UL4 were for the first time mapped to amino acid residues 178 to 186. In addition,the N-terminal 19 amino acids are identified to be required for the granule-like cytoplasmic pattern of UL4.Furthermore,the UL4 protein was demonstrated to be exported to the cytoplasm through the NES in a chromosomal region maintenance 1(CRM l)-dependent manner involving RanGTP hydrolysis.
