Dataset Information

Extracellular matrix stiffness mediates uterine repair via the Rap1a/ARHGAP35/RhoA/F-actin/YAP axis.

ABSTRACT: The integrity of the structure and function of the endometrium is essential for the maintenance of fertility. However, the repair mechanisms of uterine injury remain largely unknown. Here, we showed that the disturbance of mechanical cue homeostasis occurs after uterine injury. Applying a multimodal approach, we identified YAP as a sensor of biophysical forces that drives endometrial regeneration. Through protein activation level analysis of the combinatorial space of mechanical force strength and of the presence of particular kinase inhibitors and gene silencing reagents, we demonstrated that mechanical cues related to extracellular matrix rigidity can turn off the Rap1a switch, leading to the inactivation of ARHGAP35and then induced activation of RhoA, which in turn depends on the polymerization of the agonist protein F-actin to activate YAP. Further study confirmed that mechanotransduction significantly accelerates remodeling of the uterus by promoting the proliferation of endometrial stromal cells in vitro and in vivo. These studies provide new insights into the dynamic regulatory mechanisms behind uterine remodeling and the function of mechanotransduction. Video Abstract.

SUBMITTER: Zhang T

PROVIDER: S-EPMC9869517 | biostudies-literature | 2023 Jan

REPOSITORIES: biostudies-literature

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Extracellular matrix stiffness mediates uterine repair via the Rap1a/ARHGAP35/RhoA/F-actin/YAP axis.

Zhang Tao T Hu Ruiting R Wang Yan Y Guo Shuai S Wu Zhimin Z Liu Junfeng J Han Chunyang C Qiu Changwei C Deng Ganzhen G

Cell communication and signaling : CCS 20230123 1

The integrity of the structure and function of the endometrium is essential for the maintenance of fertility. However, the repair mechanisms of uterine injury remain largely unknown. Here, we showed that the disturbance of mechanical cue homeostasis occurs after uterine injury. Applying a multimodal approach, we identified YAP as a sensor of biophysical forces that drives endometrial regeneration. Through protein activation level analysis of the combinatorial space of mechanical force strength a ...[more]

PMID: 36691027

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Project description:Rational: Increasing evidence indicates that the physical environment is a critical mediator of tumor behavior. Hepatocellular carcinoma (HCC) develops in an altered biomechanical environment, and increased matrix stiffness is a strong predictor of HCC development. C-X-C chemokine receptor type 4 (CXCR4) is known to trigger HCC progression. However, CXCR4 as a mediator of mechanical cues in HCC is not well characterized. Methods: qRT-PCR, Western blot and IHC were used to detect the CXCR4 expression in different matrix stiffness gels. MTT was used to measure the cell proliferation of HCC cells. Immunoblotting was used for detection of epithelial-to-mesenchymal transition (EMT) and stemness on the matrix stiffness. Immunofluorescence (IF) was used to detect the nuclear location in HCC cells. IP was used to show the interaction between YAP, UbcH5c and β-TrCP. Results: We identified CXCR4 as a critical intracellular signal transducer that relays matrix stiffness signals to control mechano-sensitive cellular activities through ubiquitin domain-containing protein 1 (UBTD1)-mediated YAP signaling pathway. We found that CXCR4 expression was remarkably up-regulated in HCC cells with increasing matrix stiffness and mediated proliferation, epithelial to mesenchymal transition, and stemness. Mechanistically, matrix stiffness acts through CXCR4 to decrease the levels of UBTD1, which is involved in the proteasome-dependent degradation of YAP, a major cell mechano-transducer. UBTD1 interacted with components of the YAP degradation complex and promoted the interaction between YAP and its E3 ubiquitin ligase β-TrCP. UBTD1 knockdown decreased YAP ubiquitylation and resulted in the activation of YAP-targeted genes and YAP downstream signaling. Downregulation of UBTD1 in HCC tissues correlated with malignant prognostic features and overall survival. Finally, luteolin, a natural product, suppressed matrix stiffness-induced biological effects and CXCR4-mediated YAP signaling pathway in HCC cells. Conclusion: Our findings reveal CXCR4 as a molecular switch in mechano-transduction, thereby defining a mechano-signaling pathway from matrix stiffness to the nucleus.

Project description:OBJECTIVE:Vascular calcification is a pathology characterized by arterial mineralization, which is a common late-term complication of atherosclerosis that independently increases the risk of adverse cardiovascular events by fourfold. A major source of calcifying cells is transdifferentiating vascular smooth muscle cells (VSMCs). Previous studies showed that deletion of the collagen-binding receptor, DDR1 (discoidin domain receptor-1), attenuated VSMC calcification. Increased matrix stiffness drives osteogenesis, and DDR1 has been implicated in stiffness sensing in other cell types; however, the role of DDR1 as a mechanosensor in VSMCs has not been investigated. Here, we test the hypothesis that DDR1 senses increased matrix stiffness and promotes VSMC transdifferentiation and calcification. Approach and Results: Primary VSMCs isolated from Ddr1+/+ (wild-type) and Ddr1-/- (knockout) mice were studied on collagen-I-coated silicon substrates of varying stiffness, culturing in normal or calcifying medium. DDR1 expression and phosphorylation increased with increasing stiffness, as did in vitro calcification, nuclear localization of Runx2 (Runt-related transcription factor 2), and expression of other osteochondrocytic markers. By contrast, DDR1 deficient VSMCs were not responsive to stiffness and did not undergo transdifferentiation. DDR1 regulated stress fiber formation and RhoA (ras homolog family member A) activation through the RhoGEF (rho guanine nucleotide exchange factor), Vav2. Inhibition of actomyosin contractility reduced Runx2 activation and attenuated in vitro calcification in wild-type VSMCs. Finally, a novel positive feedforward loop was uncovered between DDR1 and actomyosin contractility, important in regulating DDR1 expression, clustering, and activation. CONCLUSIONS:This study provides mechanistic insights into DDR1 mechanosignaling and shows that DDR1 activity and actomyosin contractility are interdependent in mediating stiffness-dependent increases in VSMC calcification.

Project description:BackgroundDespite the promise of dual BRAF/MEK inhibition as a therapy for BRAF-mutant (BRAF-mut) melanoma, heterogeneous responses have been observed in patients, thus predictors of benefit from therapy are needed. We have previously identified semaphorin 6A (SEMA6A) as a BRAF-mut-associated protein involved in actin cytoskeleton remodeling. The purpose of the present study is to dissect the role of SEMA6A in the biology of BRAF-mut melanoma, and to explore its predictive potential towards dual BRAF/MEK inhibition.MethodsSEMA6A expression was assessed by immunohistochemistry in melanoma cohort RECI1 (N = 112) and its prognostic potential was investigated in BRAF-mut melanoma patients from DFCI and TCGA datasets (N = 258). The molecular mechanisms regulated by SEMA6A to sustain tumor aggressiveness and targeted therapy resistance were investigated in vitro by using BRAF-mut and BRAF-wt melanoma cell lines, an inducible SEMA6A silencing cell model and a microenvironment-mimicking fibroblasts-coculturing model. Finally, SEMA6A prediction of benefit from dual BRAF/MEK inhibition was investigated in melanoma cohort RECI2 (N = 14).ResultsOur results indicate higher protein expression of SEMA6A in BRAF-mut compared with BRAF-wt melanoma patients and show that SEMA6A is a prognostic indicator in BRAF-mut melanoma from TCGA and DFCI patients cohorts. In BRAF-mut melanoma cells, SEMA6A coordinates actin cytoskeleton remodeling by the RhoA-dependent activation of YAP and dual BRAF/MEK inhibition by dabrafenib+trametinib induces SEMA6A/RhoA/YAP axis. In microenvironment-mimicking co-culture condition, fibroblasts confer to melanoma cells a proliferative stimulus and protect them from targeted therapies, whereas SEMA6A depletion rescues the efficacy of dual BRAF/MEK inhibition. Finally, in BRAF-mut melanoma patients treated with dabrafenib+trametinib, high SEMA6A predicts shorter recurrence-free interval.ConclusionsOverall, our results indicate that SEMA6A contributes to microenvironment-coordinated evasion of melanoma cells from dual BRAF/MEK inhibition and it might be a good candidate predictor of short-term benefit from dual BRAF/MEK inhibition.

Dataset Information

Extracellular matrix stiffness mediates uterine repair via the Rap1a/ARHGAP35/RhoA/F-actin/YAP axis.

Publications

Extracellular matrix stiffness mediates uterine repair via the Rap1a/ARHGAP35/RhoA/F-actin/YAP axis.

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