Biochimica et biophysica acta. Molecular cell research最新文献

Liver fibrosis and cirrhosis generate a stiff extracellular matrix (ECM) niche that is closely associated with hepatocellular carcinoma (HCC) initiation and progression. Although multiple pathways and molecules are implicated in ECM rigidity-induced mechanical force transduction, the precise mechanism by which ECM rigidity drives HCC progression remain to be fully elucidated. In this study, we identified nuclear prelamin A recognition factor (NARF) as a novel matrix stiffness-responsive gene, whose transcription is directly regulated by the mechanosensor Yes-associated protein (YAP). Clinically, NARF exhibited high expressions in HCC tissues, and its overexpression was closely correlated with poor prognostic outcomes in HCC patients. Functionally, NARF knockdown significantly inhibited the proliferation, migration, invasion, and epithelial-mesenchymal transition (EMT) of HCC cells in vitro, whereas NARF overexpression enhanced these cellular processes. NARF silencing attenuated HCC cell growth and lung metastasis in vivo. RNA-sequencing analysis revealed a strong correlation of NARF with Wnt signaling activation. Further experiments confirmed that NARF positively regulated the expression of key Wnt target genes (MYC, CCND1, SNAIL, and TWIST1) in HCC cells. Mechanistically, NARF recruited acetyltransferase EP300 to enhance H3K27 acetylation at lymphoid enhancer binding factor 1 (LEF1)-binding sites, thereby amplifying LEF1-dependent transcriptional activity. LEF1 knockdown markedly abrogated NARF-mediated oncogenic activity in vitro and in vivo, confirming LEF1 as a critical downstream effector of NARF. Collectively, our findings identify NARF as stiffness-responsive driver that is transcriptionally regulated by YAP protein in HCC. By activating LEF1-mediated Wnt signaling in an EP300 dependent manner, NARF promotes HCC growth and metastasis, highlighting its potential as a prognostic biomarker and therapeutic target for HCC.

The retinal pigment epithelium (RPE) performs key roles in preserving retinal integrity and must continuously manage oxidative stress (OS). We previously demonstrated that the canonical phospholipase D isoforms, PLD1 and PLD2, mediate the RPE inflammatory response triggered by inflammatory injury. This study explores the mechanisms of modulation of OS mediated by PLD inhibition in RPE cells exposed to high glucose (HG) levels. ARPE-19, D407 and the novel human RPE cell line ABC were cultured under HG (33 mM) or normal glucose (NG, 5.5 mM) conditions. To inhibit PLD1, PLD2, and NADPH oxidase (NOX), VU0359595 (PLD1i), VU0285655-1 (PLD2i), and diphenyleneiodonium chloride (DPI) were used, respectively. HG exposure significantly increased reactive oxygen species (ROS) levels and reduced mitochondrial membrane potential (MMP) in ARPE-19 and D407 cells. These effects were prevented by PLD1i and PLD2i in an Nrf-2 and cyclooxygenase-2 -independent manner. In ARPE-19 cells, DPI prevented OS induced by HG as well as the stress triggered by the combination of phosphatidic acid + diacylglycerol, bioactive lipids generated through the PLD pathway-. Similarly, HG elevated ROS levels in ABC cells, and this increase was prevented by PLD1i and DPI. RNAseq analysis showed differential expression of NOX family members (NOX1,2 and 4 and DUOX1 and 2) in ARPE-19 and ABC cells. Our results demonstrate that PLDs inhibition prevent HG-induced OS in RPE cells, possibly by reducing NOX activity. The PLD pathway constitutes a novel pharmacological target to simultaneously mitigate OS and the inflammatory response, two hallmarks of retinal degenerative diseases.

The Eps15 Homology Domain protein-1 (EHD1) is an ATPase and key endocytic regulatory protein required for optimal receptor recycling, and primary ciliogenesis. Over the past decade, a central role for EHD1 has been identified in the fission of endosomes. Despite these findings, additional evidence has also pointed at a potential function of EHD1 in the regulation of microtubules. Herein, we demonstrate that EHD1 regulates the distribution of endosomes, and conversely, centrosome depletion alters EHD1 localization in cells. We show that endogenous EHD1 is found in a complex with various endogenous tubulins including TUBB3, TUBB1, α-tubulin and γ-tubulin, interactions that are independent of intact microtubules, and appear to be indirect. Depletion of key individual EHD1 interaction partners that are known to bind tubulin fail to impede EHD1-tubulin interactions, suggesting that either several proteins are capable of mediating EHD1's connection with microtubules, or that the bridging interaction partner remains to be identified. Functionally, EHD1 depletion leads to impaired microtubule regrowth and decreased end-binding protein displacement, suggesting a role for EHD1 in modulating microtubule plus-end dynamics. Finally, EHD1's role in microtubule regulation appears to be evolutionarily conserved, as single-cell stage C. elegans embryos with a dysfunctional EHD1/RME-1 protein displayed enhanced tubulin accumulation at metaphase spindle poles. Our findings strongly support a previously unaddressed role for EHD1 in microtubule regulation.

In melanoma, SOX9 and SOX10 are markers of the mesenchymal and melanocytic state, respectively. Using a panel of BRAF^V600E positive YUMM lines, we find that, following chronic vemurafenib treatment, SOX10 is lost whereas SOX9 is induced. Overexpression or knock-down of either SOX9 or SOX10 had no impact on vemurafenib sensitivity. However, we find that SOX9 is necessary to program a vemurafenib-resistance memory state following a drug holiday in vitro. RNA-Seq studies show that the loss of Sox10 represents an intermediate state that is accompanied by the loss of Sox6 and the induction of Sox7, Sox9 and other phenotype switching markers. However, SOX7 expression is not sufficient to induce vemurafenib resistance. Upon acquired drug resistance, we observed differential chromatin accessibility in the Sox9 and Sox10 upstream regions, supporting their activation and repression, respectively. Overall, our data show that the loss of SOX10 and SOX9 induction are critical to program drug resistance. Furthermore, we show that the YUMM cell lines represent a good murine model to investigate transitions to an acquired drug resistant state.