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Hepatology Communications Jul 2023HCC characterizes malignant metastasis with high incidence and recurrence. Thus, it is pivotal to discover the mechanisms of HCC metastasis. TATA-box-binding protein...
BACKGROUND
HCC characterizes malignant metastasis with high incidence and recurrence. Thus, it is pivotal to discover the mechanisms of HCC metastasis. TATA-box-binding protein (TBP), a general transcriptional factor (TF), couples with activators and chromatin remodelers to sustain the transcriptional activity of target genes. Here, we investigate the key role of TBP in HCC metastasis.
METHODS
TBP expression was measured by PCR, western blot, and immunohistochemistry. RNA-sequencing was performed to identify downstream proteins. Functional assays of TBP and downstream targets were identified in HCC cell lines and xenograft models. Luciferase reporter and chromatin immunoprecipitation assays were used to demonstrate the mechanism mediated by TBP.
RESULTS
HCC patients showed high expression of TBP, which correlated with poor prognosis. Upregulation of TBP increased HCC metastasis in vivo and in vitro, and muscleblind-like-3 (MBNL3) was the effective factor of TBP, positively related to TBP expression. Mechanically, TBP transactivated and enhanced MBNL3 expression to stimulate exon inclusion of lncRNA-paxillin (PXN)-alternative splicing (AS1) and, thus, activated epithelial-mesenchymal transition for HCC progression through upregulation of PXN.
CONCLUSIONS
Our data revealed that TBP upregulation is an HCC enhancer mechanism that increases PXN expression to drive epithelial-mesenchymal transition.
Topics: Humans; Biological Assay; Carcinoma, Hepatocellular; Epithelial-Mesenchymal Transition; Liver Neoplasms; RNA-Binding Proteins; TATA-Box Binding Protein; Animals
PubMed: 37314767
DOI: 10.1097/HC9.0000000000000155 -
Archives of Razi Institute Feb 2023In the migration and metastasis of cancer cells, it is necessary to rotate the focal adhesion (FA). MAP4K4 plays a vital role in the formation of cytoskeletal...
In the migration and metastasis of cancer cells, it is necessary to rotate the focal adhesion (FA). MAP4K4 plays a vital role in the formation of cytoskeletal regeneration, but its role in regulating FA dynamics and cancer cell migration is not well understood. This present aimed to investigate the role of MAP4K4 in regulating FA dynamics and cell migration in the human breast cancer cell line. For this purpose, different variants, including MAP4K4 (wild type), partial active mutation kinase (MAP4K4-T178D), mutant with inactivated or reduced activity kinase (MAP4K4-T178A) and inactive kinase mutation (MAP4K4-K54R) was used in the evaluation. GFP-paxillin was also used as a marker in basal breast cancer cells (MDA-MB-231) in determining FA dynamics. Time-lapse and confocal microscopes were used to record FA dynamics and cell migration. The present study's findings showed that cells expressing MAP4K4-K54R, MAP4K4-T178D and MAP4K4-T178A type slowly down the FA turnover rates and had much larger FAs than those expressing WT MAP4K4 in MDA-MB-231 breast cancer cell line. Furthermore, inhibiting MAP4K4 strongly inhibited FA formation and reduced cell migration speed. In conclusion, MAP4K4 regulates FA dynamics and cancer cell migration, most probably through activating FA proteins and cytoskeleton.
Topics: Humans; Cell Movement; Focal Adhesions; Intracellular Signaling Peptides and Proteins; MCF-7 Cells; MDA-MB-231 Cells; Mutation; Neoplasms; Protein Serine-Threonine Kinases
PubMed: 37312710
DOI: 10.22092/ARI.2022.358953.2340 -
IScience Jun 2023The objective of this study is to develop a device to mimic a microfluidic system of human arterial blood vessels. The device combines fluid shear stress (FSS) and...
The objective of this study is to develop a device to mimic a microfluidic system of human arterial blood vessels. The device combines fluid shear stress (FSS) and cyclic stretch (CS), which are resulting from blood flow and blood pressure, respectively. The device can reveal real-time observation of dynamic morphological change of cells in different flow fields (continuous flow, reciprocating flow and pulsatile flow) and stretch. We observe the effects of FSS and CS on endothelial cells (ECs), including ECs align their cytoskeleton proteins with the fluid flow direction and paxillin redistribution to the cell periphery or the end of stress fibers. Thus, understanding the morphological and functional changes of endothelial cells on physical stimuli can help us to prevent and improve the treatment of cardiovascular diseases.
PubMed: 37305698
DOI: 10.1016/j.isci.2023.106927 -
Heliyon Jun 2023Cervical cancer is the most common gynecological pernicious tumor with high morbidity and mortality worldwide, especially in developing countries. Arctigenin (ARG), a...
CONTEXT
Cervical cancer is the most common gynecological pernicious tumor with high morbidity and mortality worldwide, especially in developing countries. Arctigenin (ARG), a nature-derived component, has exhibited anti-tumor activity in various tumors.
OBJECTIVE
To explore the effect of ARG on cervical cancer.
MATERIALS AND METHODS
The effect and mechanism of ARG on cervical cancer cells were explored by cell counting kit-8 (CCK-8), flow cytometry, transwell and Western blot assays. Additionally, experiment was conducted in xenografted mice by immunohistochemistry (IHC), terminal deoxynucleotidyl transferase deoxyuridine triphosphate (dUTP) nick end labeling (TUNEL) and Western blot assays.
RESULTS
ARG treatment induced both concentration-dependent and time-dependent reductions in the cell viability of SiHa and HeLa cells with a IC50 value of 9.34 μM and 14.45 μM, respectively. ARG increased the apoptosis rate and the protein levels of cleaved-caspase 3 and E-cadherin, but decreased the invaded cell numbers and the protein levels of Vimentin and N-cadherin . Mechanically, ARG inhibited the expression of focal adhesion kinase (FAK)/paxillin pathway, which was confirmed by the overexpression of FAK in SiHa cells. The inhibitory role of overexpression of FAK in proliferation and invasion, as well as its promoted role in apoptosis were reversed with ARG treatment. Meanwhile, ARG suppressed growth and metastasis, and enhanced apoptosis . Consistently, ARG administration reduced the relative protein level of -FAK/FAK and -paxillin/paxillin in tumor tissues of xenografted mice.
CONCLUSION
ARG inhibited proliferation, invasion and metastasis, but enhanced apoptosis of cervical cancer via the FAK/paxillin axis.
PubMed: 37292259
DOI: 10.1016/j.heliyon.2023.e16683 -
Frontiers in Neuroscience 2023The adhesion systems employed by migrating cortical neurons are not well understood. Genetic deletion studies of focal adhesion kinase (FAK) and paxillin in mice...
The adhesion systems employed by migrating cortical neurons are not well understood. Genetic deletion studies of focal adhesion kinase (FAK) and paxillin in mice suggested that these classical focal adhesion molecules control the morphology and speed of cortical neuron migration, but whether β1 integrins also regulate migration morphology and speed is not known. We hypothesized that a β1 integrin adhesion complex is required for proper neuronal migration and for proper cortical development. To test this, we have specifically deleted β1 integrin from postmitotic migrating and differentiating neurons by crossing conditional β1 integrin floxed mice into the transgenic line. Similar to our prior findings with conditional paxillin deficiency, we found that both homozygous and heterozygous deletion of β1 integrin causes transient mispositioning of cortical neurons in the developing cortex when analyzed pre- and perinatally. Paxillin and β1 integrin colocalize in the migrating neurons and deletion of paxillin in the migrating neuron causes an overall reduction of the β1 integrin immunofluorescence signal and reduction in the number of activated β1 integrin puncta in the migrating neurons. These findings suggest that these molecules may form a functional complex in migrating neurons. Similarly, there was an overall reduced number of paxillin+ puncta in the β1 integrin deficient neurons, despite the normal distribution of FAK and Cx26, a connexin required for cortical migration. The double knockout of paxillin and β1 integrin produces a cortical malpositioning phenotype similar to the paxillin or β1 integrin single knockouts, as would be expected if paxillin and β1 integrin function on a common pathway. Importantly, an isolation-induced pup vocalization test showed that β1 integrin mutants produced a significantly smaller number of calls compared to their littermate controls when analyzed at postnatal day 4 (P4) and revealed a several days trend in reduced vocalization development compared to controls. The current study establishes a role for β1 integrin in cortical development and suggests that β1 integrin deficiency leads to migration and neurodevelopmental delays.
PubMed: 37250402
DOI: 10.3389/fnins.2023.1158419 -
Journal of Cell Science Jun 2023Vinculin is an actin-binding protein present at cell-matrix and cell-cell adhesions, which plays a critical role in bearing force experienced by cells and dissipating it...
Vinculin is an actin-binding protein present at cell-matrix and cell-cell adhesions, which plays a critical role in bearing force experienced by cells and dissipating it onto the cytoskeleton. Recently, we identified a key tyrosine residue, Y822, whose phosphorylation plays a critical role in force transmission at cell-cell adhesions. The role of Y822 in human cancer remains unknown, even though Y822 is mutated to Y822C in uterine cancers. Here, we investigated the effect of this amino acid substitution and that of a phosphodeficient Y822F vinculin in cancer cells. We observed that the presence of the Y822C mutation led to cells that proliferate and migrate more rapidly and contained smaller focal adhesions when compared to cells with wild-type vinculin. In contrast, the presence of the Y822F mutation led to highly spread cells with larger focal adhesions and increased contractility. Furthermore, we provide evidence that Y822C vinculin forms a disulfide bond with paxillin, accounting for some of the elevated phosphorylated paxillin recruitment. Taken together, these data suggest that vinculin Y822 modulates the recruitment of ligands.
Topics: Humans; Vinculin; Paxillin; Ligands; Cell Adhesion; Cell Communication; Focal Adhesions
PubMed: 37248996
DOI: 10.1242/jcs.260104 -
Journal of Cell Communication and... Jun 2023Skin primarily comprises a collagen-rich extracellular matrix (ECM) that provides structural and functional support to the skin. Aging causes progressive loss and...
Skin primarily comprises a collagen-rich extracellular matrix (ECM) that provides structural and functional support to the skin. Aging causes progressive loss and fragmentation of dermal collagen fibrils, leading to thin and weakened skin (Dermal aging). We previously reported that CCN1 is elevated in naturally aged human skin, photoaged human skin, and acute UV-irradiated human skin dermal fibroblasts in vivo. Elevated CCN1 alters the expression of numerous secreted proteins that have deleterious effects on the dermal microenvironment, impairing the structural integrity and function of the skin. Here we show that CCN1 is predominantly elevated in the human skin dermis by UV irradiation and accumulated in the dermal extracellular matrix. Laser capture microdissection indicated that CCN1 is predominantly induced in the dermis, not in the epidermis, by acute UV irradiation in human skin in vivo. Interestingly, while UV-induced CCN1 in the dermal fibroblasts and in the medium is transient, secreted CCN1 accumulates in the ECM. We explored the functionality of the matrix-bound CCN1 by culturing dermal fibroblasts on an acellular matrix plate that was enriched with a high concentration of CCN1. We observed that matrix-bound CCN1 activates integrin outside-in signaling resulting in the activation of FAK and its downstream target paxillin and ERK, as well as elevated MMP-1 and inhibition of collagen, in human dermal fibroblasts. These data suggest that accumulation of CCN1 in the dermal ECM is expected to progressively promote the aging of the dermis and thereby negatively impact the function of the dermis.
PubMed: 37245186
DOI: 10.1007/s12079-023-00767-6 -
Biomedicines May 2023A chronic subdural hematoma (CSDH) is considered to be an inflammatory and angiogenic disease. The CSDH outer membrane, which contains inflammatory cells, plays an...
BACKGROUND
A chronic subdural hematoma (CSDH) is considered to be an inflammatory and angiogenic disease. The CSDH outer membrane, which contains inflammatory cells, plays an important role in CSDH development. Osteopontin (OPN) is an extracellular matrix protein that is cleaved by thrombin, generating the N-terminal half of OPN, which is prominently involved in integrin signal transduction. We explored the expression of the N-terminal half of OPN in CSDH fluid and the expression of integrins α9 and β1 and the downstream components of the angiogenic signaling pathways in the outer membrane of CSDHs.
METHODS
Twenty samples of CSDH fluid and eight samples of CSDH outer membrane were collected from patients suffering from CSDHs. The concentrations of the N-terminal half of OPN in CSDH fluid samples were measured using ELISA kits. The expression levels of integrins α9 and β1, vinculin, talin-1, focal adhesion kinase (FAK), paxillin, α-actin, Src and β-actin were examined by Western blot analysis. The expression levels of integrins α9 and β1, FAK and paxillin were also examined by immunohistochemistry. We investigated whether CSDH fluid could activate FAK in cultured endothelial cells in vitro.
RESULTS
The concentration of the N-terminal half of OPN in CSDH fluid was significantly higher than that in the serum. Western blot analysis confirmed the presence of these molecules. In addition, integrins α9 and β1, FAK and paxillin were localized in the endothelial cells of vessels within the CSDH outer membrane. FAK was significantly phosphorylated immediately after treatment with CSDH fluid.
CONCLUSION
Our data suggest that the N-terminal half of OPN in CSDH fluid promotes neovascularization in endothelial cells through integrins α9 and β1. The N-terminal half of OPN, which is part of the extracellular matrix, plays a critical role in the promotion of CSDHs.
PubMed: 37239111
DOI: 10.3390/biomedicines11051440 -
Cancer Medicine Jul 2023In lung cancer patients, most deaths are caused by the distant dissemination of cancer cells. Epithelial-mesenchymal transition (EMT) and collective cell migration are...
OBJECTIVE
In lung cancer patients, most deaths are caused by the distant dissemination of cancer cells. Epithelial-mesenchymal transition (EMT) and collective cell migration are distinct and important mechanisms involved in cancer invasion and metastasis. Additionally, microRNA dysregulation contributes significantly to cancer progression. In this study, we aimed to explore the function of miR-503 in cancer metastasis.
METHODS
Molecular manipulations (silencing or overexpression) were performed to investigate the biological functions of miR-503 including migration and invasion. Reorganization of cytoskeleton was assessed using immunofluorescence and the relationship between miR-503 and downstream protein tyrosine kinase 7 (PTK7) was assessed using quantitative real-time PCR, immunoblotting, and reporter assays. The tail vein metastatic animal experiments were performed.
RESULTS
Herein, we demonstrated that the downregulation of miR-503 confers an invasive phenotype in lung cancer cells and provided in vivo evidence that miR-503 significantly inhibits metastasis. We found that miR-503 inversely regulates EMT, identified PTK7 as a novel miR-503 target, and showed the functional effects of miR-503 on cell migration and invasion were restored upon reconstitution of PTK7 expression. As PTK7 is a Wnt/planar cell polarity protein crucial for collective cell movement, these results implicated miR-503 in both EMT and collective migration. However, the expression of PTK7 did not influence EMT induction, suggesting that miR-503 regulates EMT through mechanisms other than PTK7 inhibition. Furthermore, we discovered that PTK7 mechanistically activates focal adhesion kinase (FAK) and paxillin, thereby controlling the reorganization of the cortical actin cytoskeleton.
CONCLUSION
Collectively, miR-503 is capable of governing EMT and PTK7/FAK signaling independently to control the invasion and dissemination of lung cancer cells, indicating that miR-503 represents a pleiotropic regulator of cancer metastasis and hence a potential therapeutic target for lung cancer.
Topics: Animals; Epithelial-Mesenchymal Transition; Cell Line, Tumor; Lung Neoplasms; MicroRNAs; Signal Transduction; Cell Movement; Focal Adhesion Protein-Tyrosine Kinases; Gene Expression Regulation, Neoplastic; Neoplasm Invasiveness; Neoplasm Metastasis
PubMed: 37212485
DOI: 10.1002/cam4.6116 -
The Role of Paxillin Aberrant Expression in Cancer and Its Potential as a Target for Cancer Therapy.International Journal of Molecular... May 2023Paxillin is a multi-domain adaptor protein. As an important member of focal adhesion (FA) and a participant in regulating cell movement, paxillin plays an important role... (Review)
Review
Paxillin is a multi-domain adaptor protein. As an important member of focal adhesion (FA) and a participant in regulating cell movement, paxillin plays an important role in physiological processes such as nervous system development, embryonic development, and vascular development. However, increasing evidence suggests that paxillin is aberrantly expressed in many cancers. Many scholars have also recognized that the abnormal expression of paxillin is related to the prognosis, metastases, invasion, survival, angiogenesis, and other aspects of malignant tumors, suggesting that paxillin may be a potential cancer therapeutic target. Therefore, the study of how aberrant paxillin expression affects the process of tumorigenesis and metastasis will help to develop more efficacious antitumor drugs. Herein, we review the structure of paxillin and its function and expression in tumors, paying special attention to the multifaceted effects of paxillin on tumors, the mechanism of tumorigenesis and progression, and its potential role in tumor therapy. We also hope to provide a reference for the clinical prognosis and development of new tumor therapeutic targets.
Topics: Humans; Paxillin; Neoplasms; Cell Movement; Antineoplastic Agents; Carcinogenesis; Cell Line, Tumor
PubMed: 37175948
DOI: 10.3390/ijms24098245