Correlation analysis of cofilin-1 with renal prognosis in primary IgA nephropathy.

PURPOSE: The purpose of this study was to investigate the correlation between podocyte related biomarker cofilin-1 and renal function, and explore the value of cofilin-1 in predicting the risk of renal adverse prognosis in IgA nephropathy (IgAN). METHODS: Patients with primary IgAN diagnosed by initial renal biopsy performed in our hospital from January 2019 to February 2022 were included. This study was a prospective cohort study. All IgAN patients were detected the expression of cofilin-1 and other related biomarkers (RhoA, NGAL) in urine by enzyme-linked immunosorbent assay (ELISA) and follow-up at least 6 months. We also collected baseline clinicopathologial data of IgAN. The decreased renal function group was defined as baseline eGFR < 60 ml/min/1.73m2. Logistic and Cox regression model were used to analyze the correlation among cofilin-1 and renal prognosis. RESULTS: 133 IgAN patients were included, with a male-to-female ratio of 1.25:1 and an age of 37.67 ± 13.78 years, as well as an average of eGFR was 71.63 (40.42,109.33) ml/min/1.73m2. 56 patients (42.1%) had decreased renal function at baseline, with the average of eGFR was 34.07 (16.72, 49.21) ml/min/1.73 m2. 12 of which developed to renal adverse prognosis. The average of follow-up time was 22.035 ± 8.992 months. The multivariate regression analysis showed that increased urinary cofilin-1 was an independent risk factor associated with baseline renal function decline and renal adverse prognosis in IgAN patients (P < 0.05). ROC curves showed great efficacy of urinary cofilin-1 levels in diagnosing baseline renal function decline and predicting renal adverse prognosis (the area under the ROC curve was 0.708 and 0.803). CONCLUSION: Cofilin-1 as a novel biomarker of podocyte lesion is closely related to renal function decline in IgAN. Cofilin-1 has certain clinical value in predicting the risk of renal adverse prognosis. Podocyte fusion affects the renal prognosis of IgAN.

[The expression and prognosis prediction of CFL-1 in head and neck squamous cell carcinoma].

PURPOSE: The purpose of this study was to explore the expression, prognostic value and immune correlation of Cofilin 1 protein(CFL-1) in head and neck squamous cell carcinoma(HNSCC). METHODS: The expression and prognostic value of CFL-1 in head and neck squamous cell carcinoma(HNSCC) was explored in the cancer genome map database (TCGA) and gene expression total databases (GEO), and the potential immune pathway of CFL-1 in HNSCC was revealed by GESA and cibersoft analysis. The data were statistically analyzed using SPSS 26.0 software package. RESULTS: CFL-1 was significantly up-regulated in HNSCC tissue. The expression level of CFL-1 was significantly correlated with the overall survival status of HNSCC. High expression of CFL-1 was significantly associated with a lower overall survival rate. In addition, multivariate Cox survival analysis showed that CFL-1 expression was independent predictors of poor prognosis of HNSCC. GESA and cibersoft analysis showed that the imbalance of CFL-1 expression affected multiple signal pathways and infiltration of immune cells. CONCLUSIONS: CFL-1 is highly expressed in HNSCC and is significantly associated with poor prognosis of NHSCC. It is a potential prognostic marker of HNSCC.

LIMK1 promotes the development of cervical cancer by up-regulating the ROS/Src-FAK/cofilin signaling pathway.

OBJECTIVE: In this study, we investigated the mechanism of action of LIMK1 in cervical cancer progression. METHODS: The biological role of LIMK1 in regulating the growth, invasion, and metastasis of cervical cancer was studied in SiHa, CaSki cells and nude mice tumor models. The role of LIMK1 in the growth of cervical cancer was evaluated by HE staining. The role of LIMK1 in the invasion, metastasis, and proliferation of cervical cancer was evaluated by cell scratch, Transwell, and monoclonal experiments. The interaction among LIMK1, ROS, and Src was evaluated by Western blotting. The effects of regulating ROS and p-Src expression on LIMK1 in the migration/invasion and proliferation of cervical cancer cells were evaluated through cellular functional assays. RESULTS: Overexpression of LIMK1 promoted tumor growth in nude mice. Cell scratch, Transwell, and monoclonal experiments suggested that LIMK1 promoted the invasion, metastasis, and proliferation of cervical cancer cells. Western blotting suggested that LIMK1 can promote the expression of ROS-related proteins NOX2, NOX4, p-Src, and downstream proteins p-FAK, p-ROCK1/2, p-Cofilin-1, F-actin and inhibit the expression of p-SHP2 protein. Correction experiments showed that LIMK1 regulated the expression of p-FAK and p-Cofilin-1 proteins by regulating ROS and p-Src. Through the detection of cervical cancer cell functions, it was found that the activation of ROS and p-Src induced by LIMK1 is an early event that promotes the migration, proliferation, and invasion of cervical cancer cells. CONCLUSIONS: LIMK1 promotes the expression of F-actin and promotes the development of cervical cancer by regulating the oxidative stress/Src-mediated p-FAK/p-ROCK1/2/p-Cofilin-1 pathway.

The interaction of PRDX1 with Cofilin promotes oral squamous cell carcinoma metastasis.

Peroxiredoxin 1 (PRDX1) is an important member of the peroxiredoxin family (PRDX) and is upregulated in a variety of tumors. Previous studies have found that high PRDX1 expression is closely related to the metastasis of oral squamous cell carcinoma (OSCC), but the specific molecular mechanism is elusive. To elucidate the role of PRDX1 in the metastasis process of OSCC, we evaluated the expression of PRDX1 in OSCC clinical specimens and its impact on the prognosis of OSCC patients. Then, the effect of PRDX1 on OSCC metastasis and cytoskeletal reconstruction was explored in vitro and in nude mouse tongue cancer models, and the molecular mechanisms were also investigated. PRDX1 can directly interact with the actin-binding protein Cofilin, inhibiting the phosphorylation of its Ser3 site, accelerating the depolymerization and turnover of actin, promoting OSCC cell movement, and aggravating the invasion and metastasis of OSCC. In clinical samples and mouse tongue cancer models, PRDX1 also increased lymph node metastasis of OSCC and was negatively correlated with the phosphorylation of Cofilin; PRDX1 also reduced the overall survival rate of OSCC patients. In summary, our study identified that PRDX1 may be a potential therapeutic target to inhibit OSCC metastasis.

Impact of siRNA-Mediated Cofilin-1 Knockdown and Obesity Associated Microenvironment on the Motility of Natural Killer Cells.

The anti-tumor capacity of natural killer (NK) cells heavily relies on their ability to migrate towards their target cells. This process is based on dynamic actinrearrangement, so-called actin treadmilling, andis tightly regulated by proteins such as cofilin-1. The aim of the present study was to identify the role of cofilin-1 (CFL-1) in the migratory behavior of NK cells and to investigate a possible impact of an obesity-associated micromilieu on these cells, as it is known that obesity correlates with various impaired NK cell functions. CFL-1 was knocked-down via transfection of NK-92 cells with respective siRNAs. Obesity associated micromilieu was mimicked by incubation of NK-92 cells with adipocyte-conditioned medium from human preadipocyte SGBS cells or leptin. Effects on CFL-1 levels, the degree of phosphorylation to the inactive pCFL-1 as well as NK-92 cell motility were analyzed. Surprisingly, siRNA-mediated CFL-1 knockdown led to a significant increase of migration, as determined by enhanced velocity and accumulated distance of migration. No effect on CFL-1 nor pCFL-1 expression levels, proportion of phosphorylation and cell migratory behavior could be demonstrated under the influence of an obesity-associated microenvironment. In conclusion, the results indicate a significant effect of a CFL-1 knockdown on NK cell motility.

The amoeboid migration of monocytes in confining channels requires the local remodeling of the cortical actin cytoskeleton by cofilin-1.

Within the bloodstream, monocytes must traverse the microvasculature to prevent leukostasis, which is the entrapment of monocytes within the confines of the microvasculature. Using the model cell line, THP-1, and VCAM-1 coated channels to simulate the microvasculature surface, we demonstrate that monocytes predominantly adopt an amoeboid phenotype, which is characterized by the formation of blebs. As opposed to cortical actin flow in leader blebs, cell movement is correlated with myosin contraction at the cell rear. It was previously documented that cofilin-1 promotes cortical actin turnover at leader bleb necks in melanoma cells. In monocytes, our data suggest that cofilin-1 promotes the local upregulation of myosin contractility through actin cytoskeleton remodeling. In support of this concept, cofilin-1 is found to localize to a single cell edge. Moreover, the widespread upregulation of myosin contractility was found to inhibit migration. Thus, monocytes within the microvasculature may avoid entrapment by adopting an amoeboid mode of migration.

Deciphering the Cofilin Oligomers via Intermolecular Disulfide Bond Formation: A Coarse-Grained Molecular Dynamics Approach to Understanding Cofilin's Regulation on Actin Filaments.

Cofilin, a key actin-binding protein, orchestrates the dynamics of the actomyosin network through its actin-severing activity and by promoting the recycling of actin monomers. Recent experiments suggest that cofilin forms functionally distinct oligomers via thiol post-translational modifications (PTMs) that promote actin nucleation and assembly. Despite these advances, the structural conformations of cofilin oligomers that modulate actin activity remain elusive because there are combinatorial ways to oxidize thiols in cysteines to form disulfide bonds rapidly. This study employs molecular dynamics simulations to investigate human cofilin 1 as a case study for exploring cofilin dimers via disulfide bond formation. Utilizing a biasing scheme in simulations, we focus on analyzing dimer conformations conducive to disulfide bond formation. Additionally, we explore potential PTMs arising from the examined conformational ensemble. Using the free energy profiling, our simulations unveil a range of probable cofilin dimer structures not represented in current Protein Data Bank entries. These candidate dimers are characterized by their distinct population distributions and relative free energies. Of particular note is a dimer featuring an interface between cysteines 139 and 147 residues, which demonstrates stable free energy characteristics and intriguingly symmetrical geometry. In contrast, the experimentally proposed dimer structure exhibits a less stable free energy profile. We also evaluate frustration quantification based on the energy landscape theory in the protein-protein interactions at the dimer interfaces. Notably, the 39-39 dimer configuration emerges as a promising candidate for forming cofilin tetramers, as substantiated by frustration analysis. Additionally, docking simulations with actin filaments further evaluate the stability of these cofilin dimer-actin complexes. Our findings thus offer a computational framework for understanding the role of thiol PTM of cofilin proteins in regulating oligomerization, and the subsequent cofilin-mediated actin dynamics in the actomyosin network.

Multicomponent depolymerization of actin filament pointed ends by cofilin and cyclase-associated protein depends upon filament age.

Intracellular actin networks assemble through the addition of ATP-actin subunits at the growing barbed ends of actin filaments. This is followed by "aging" of the filament via ATP hydrolysis and subsequent phosphate release. Aged ADP-actin subunits thus "treadmill" through the filament before being released back into the cytoplasmic monomer pool as a result of depolymerization at filament pointed ends. The necessity for aging before filament disassembly is reinforced by preferential binding of cofilin to aged ADP-actin subunits over newly-assembled ADP-Pi actin subunits in the filament. Consequently, investigations into how cofilin influences pointed-end depolymerization have, thus far, focused exclusively on aged ADP-actin filaments. Using microfluidics-assisted Total Internal Reflection Fluorescence (mf-TIRF) microscopy, we reveal that, similar to their effects on ADP filaments, cofilin and cyclase-associated protein (CAP) also promote pointed-end depolymerization of ADP-Pi filaments. Interestingly, the maximal rates of ADP-Pi filament depolymerization by CAP and cofilin together remain approximately 20-40 times lower than for ADP filaments. Further, we find that the promotion of ADP-Pi pointed-end depolymerization is conserved for all three mammalian cofilin isoforms. Taken together, the mechanisms presented here open the possibility of newly-assembled actin filaments being directly disassembled from their pointed-ends, thus bypassing the slow step of Pi release in the aging process.

Identification of an immune-related gene signature for predicting prognosis and immunotherapy efficacy in liver cancer via cell-cell communication.

BACKGROUND: Liver cancer is one of the deadliest malignant tumors worldwide. Immunotherapy has provided hope to patients with advanced liver cancer, but only a small fraction of patients benefit from this treatment due to individual differences. Identifying immune-related gene signatures in liver cancer patients not only aids physicians in cancer diagnosis but also offers personalized treatment strategies, thereby improving patient survival rates. Although several methods have been developed to predict the prognosis and immunotherapeutic efficacy in patients with liver cancer, the impact of cell-cell interactions in the tumor microenvironment has not been adequately considered. AIM: To identify immune-related gene signals for predicting liver cancer prognosis and immunotherapy efficacy. METHODS: Cell grouping and cell-cell communication analysis were performed on single-cell RNA-sequencing data to identify highly active cell groups in immune-related pathways. Highly active immune cells were identified by intersecting the highly active cell groups with B cells and T cells. The significantly differentially expressed genes between highly active immune cells and other cells were subsequently selected as features, and a least absolute shrinkage and selection operator (LASSO) regression model was constructed to screen for diagnostic-related features. Fourteen genes that were selected more than 5 times in 10 LASSO regression experiments were included in a multivariable Cox regression model. Finally, 3 genes (stathmin 1, cofilin 1, and C-C chemokine ligand 5) significantly associated with survival were identified and used to construct an immune-related gene signature. RESULTS: The immune-related gene signature composed of stathmin 1, cofilin 1, and C-C chemokine ligand 5 was identified through cell-cell communication. The effectiveness of the identified gene signature was validated based on experimental results of predictive immunotherapy response, tumor mutation burden analysis, immune cell infiltration analysis, survival analysis, and expression analysis. CONCLUSION: The findings suggest that the identified gene signature may contribute to a deeper understanding of the activity patterns of immune cells in the liver tumor microenvironment, providing insights for personalized treatment strategies.

Neuronal CFL1 upregulation in head and neck squamous cell carcinoma enhances tumor-nerve crosstalk and promotes tumor growth.

Head and neck squamous cell carcinoma (HNSCC) is a prevalent cancer type, marked by a pronounced nerve density within the tumor microenvironment and a high rate of perineural invasion (PNI). Growing evidence suggests that the nervous system plays a vital role in HNSCC progression. Yet, the mechanisms governing cancer-nerve interactions remain largely elusive. Our research revealed that cofilin-1 (CFL1) is significantly overexpressed in HNSCC and correlates with both PNI and unfavorable prognosis. Utilizing multiplex fluorescent immunohistochemistry, we have localized CFL1 chiefly to the nerves adjacent to tumor sites. Significantly, it is the elevated expression of CFL1 in neuronal structures, rather than in the tumor cells, that aligns with diminished patient survival rates. We observed that HNSCC cells induced the expression of neuronal CFL1 and that the conditional knockout of neuronal CFL1 impedes tumor-nerve interactions. Both Gene Ontology functional enrichment analyses and Gene Set Enrichment Analysis demonstrate that CFL1 expression in HNSCC is associated with specific biological processes, including "RIBOSOME," "PROTEASOME," and "cadherin binding." In summary, HNSCC promotes the expression of CFL1 in nerves, which is essential for cancer-nerve interactions. The neuronal CFL1 is associated with PNI and may be a potential molecular prognostic marker of poor survival in HNSCC.

CFL1 restores the migratory capacity of bone marrow mesenchymal stem cells in primary Sjögren's syndrome by regulating CCR1 expression.

BACKGROUND: Primary Sjögren's syndrome (pSS) is a chronic systemic autoimmune disease. There is no relevant research on whether the migratory ability of bone marrow mesenchymal stem cells (BM-MSC) is impaired in patients with pSS (pSS-BMMSC). METHODS: Trajectories and velocities of BM-MSC were analyzed. Transwell migration assay and wound healing assay were used to investigate the migratory capacity of BM-MSC. The proliferative capacity of BM-MSC was evaluated by EDU and CCK8 assay. RNA-seq analysis was then performed to identify the underlying mechanism of lentivirus-mediated cofilin-1 overexpression BM-MSC (BMMSCCFL1). The therapeutic efficacy of BMMSCCFL1 was evaluated in NOD mice. RESULTS: The migratory capacity of pSS-BMMSC was significantly reduced compared to normal volunteers (HC-BMMSC). The expression of the motility-related gene CFL1 was decreased in pSS-BMMSC. Lentivirus-mediated CFL1 overexpression of pSS-BMMSC promoted the migration capacity of pSS-BMMSC. Furthermore, RNA-seq revealed that CCR1 was the downstream target gene of CFL1. To further elucidate the mechanism of CFL1 in regulating BM-MSC migration and proliferation via the CCL5/CCR1 axis, we performed a rescue experiment using BX431 (a CCR1-specific inhibitor) to inhibit CCR1. The results showed that CCR1 inhibitors suppressed the migration and proliferation capacity of MSC induced by CFL1. CONCLUSION: The pSS-BMMSC leads to impaired migration and proliferation, and overexpression of CFL1 can rescue the functional deficiency and alleviate disease symptoms in NOD mice. Mechanically, CFL1 can regulate the expression level of the downstream CCL5/CCR1 axis to enhance the migration and proliferation of BM-MSC.

Remodeling Serine Synthesis and Metabolism via Nanoparticles (NPs)-Mediated CFL1 Silencing to Enhance the Sensitivity of Hepatocellular Carcinoma to Sorafenib.

Tyrosine kinase inhibitors represented by sorafenib are the first-line treatment for hepatocellular carcinoma (HCC), but the low response rate of HCC patient has become a clinical pain-point. Emerging evidences have revealed that metabolic reprogramming plays an important role in regulating the sensitivity of tumor cells to various chemotherapeutics including sorafenib. However, the underlying mechanisms are very complex and are not fully elucidated. By comparing the transcriptome sequencing data of sorafenib-sensitive and -insensitive HCC patients, it is revealed that cofilin 1 (CFL1) is highly expressed in the tumor tissues of sorafenib-insensitive HCC patients and closely correlated with their poor prognosis. Mechanically, CFL1 can promote phosphoglycerate dehydrogenase transcription and enhance serine synthesis and metabolism to accelerate the production of antioxidants for scavenging the excessive reactive oxygen species induced by sorafenib, thereby impairing the sorafenib sensitivity of HCC. To translate this finding and consider the severe side effects of sorafenib, a reduction-responsive nanoplatform for systemic co-delivery of CFL1 siRNA (siCFL1) and sorafenib is further developed, and its high efficacy in inhibiting HCC tumor growth without apparent toxicity is demonstrated. These results indicate that nanoparticles-mediated co-delivery of siCFL1 and sorafenib can be a new strategy for the treatment of advanced HCC.

Cofilin promotes tau pathology in Alzheimer's disease.

The molecular mechanisms mediating the aggregation and transmission of tau in AD remain unclear. Here, we show that the actin-binding protein cofilin is cleaved by a cysteine protease asparagine endopeptidase (AEP) at N138 in the brains of patients with AD. The AEP-generated cofilin 1-138 fragment interacts with tau and promotes its aggregation. The mixed fibrils consisting of cofilin 1-138 and tau are more pathogenic to cells than pure tau fibrils. Furthermore, overexpression of cofilin 1-138 in the brain facilitates the propagation of pathological tau aggregates and promotes AD-like cognitive impairments in tau P301S mice. However, mice infected with adeno-associated viruses (AAVs) encoding an AEP-uncleavable cofilin mutant show attenuated tau pathology and cognitive impairments compared with mice injected with AAVs encoding wild-type cofilin. Together, these observations support the role of the cofilin 1-138 fragment in the aggregation and transmission of tau pathology during the onset and progression of AD.

Celastrus orbiculatus Thunb. extract inhibits EMT and metastasis of gastric cancer by regulating actin cytoskeleton remodeling.

ETHNOPHARMACOLOGICAL RELEVANCE: The traditional Chinese medicine herb Celastrus orbiculatus Thunb. is an important folk medicinal plant in China that has been used as an anti-inflammatory, antitumor, and analgesic in various diseases. Recent years, many studies have reported the significant effects of Celastrus orbiculatus Thunb. extract (COE) on gastric cancer. However, the specific mechanism by which COE regulates gastric cancer cytoskeleton remodeling and thus inhibits EMT has not yet been reported. AIM OF STUDY: To study the effect and mechanism of COE in inhibiting the epithelial-mesenchymal transition (EMT) and metastasis of gastric cancer cells, laying an experimental foundation for the clinical application and further development of COE. METHODS: The high-content cell dynamic tracking system was used to continuously track the trajectory of cell movement in real time. Through the high-content data, the average movement distance and movement speed of the cells are calculated. Additionally, the dynamic images of the cell movement in the high-content imaging system are derived to analyze the impact of COE on the movement of gastric cancer cells. Cytoskeleton staining experiment was performed to detect the effect of COE on the assembly of gastric cancer cell cytoskeleton proteins. Western blot was employed to detect the changes of EMT and metastasis-related proteins in the gastric cancer cells treated by COE. The effect of COE on the key regulatory protein Cofilin-1 (CFL1) of cell movement was examined by Western blot and protein degradation experiment. The effect of COE on EMT and metastasis of the gastric cancer cells lacking CFL1 was assessed by a transwell assay. The in vivo inhibitory effect of COE on EMT and metastasis of gastric cancer was determined by the animal living image system. IHC assays were used to detect the levels of EMT-related proteins in COE reversal in vivo. RESULT: The results showed that the movement distance and average movement speed of gastric cancer cells after COE treatment were significantly lower than those of the control group. Cytoskeleton staining experiments revealed that COE can significantly change the distribution of skeletal proteins in gastric cancer cells. Additionally, COE treatment significantly reduced the expression of Matrix metalloproteinases (MMP-2, MMP-9) and other proteins. Furthermore, COE can significantly accelerate the degradation of CFL1 protein, and both COE treatment and CFL1 deletion can significantly inhibit EMT and metastasis of gastric cancer cells. Lastly, the number of peritoneal metastases of gastric cancer cells was significantly reduced in animals after COE treatment. COE can reverse the levels of EMT-related proteins while reducing the expression levels of CFL1 protein in vivo. CONCLUSION: COE can significantly inhibit EMT and metastasis of gastric cancer cells in vivo and in vitro. This effect may be achieved by reducing the stability of CFL1 and inhibiting the assembly of actin in gastric cancer cells.

The phosphorylation level of Cofilin-1 is related to the pathological subtypes of gastric cancer.

The purpose of the study was to explore the relationship between multiple proteins belonging to the LIMK/Cofilin pathway, including LIMK1, LIMK2, Cofilin-1, and p-Cofilin-1 and clinical features of gastric cancer (GC) patients, including overall survival, TNM stages, and pathological subtypes. The expression of LIMK1, LIMK2, Cofilin-1 and p-Cofilin-1 in the GC tissues and adjacent normal stomach tissues from 141 patients were detected using immunohistochemistry (IHC) staining. Wilcoxon rank-sum test and Spearman rank correlation coefficients were used to measure the relationship between different TNM stages, pathological types, and selected parameters. OS was estimated using the Kaplan-Meier method and survival curves were compared using the log-rank test. Our results showed that, compared to those in the adjacent normal stomach tissues, LIMK1, LIMK2 and Cofilin-1 were up-regulated while p-Cofilin-1 was down-regulated in the GC tissues. LIMK1 level was positively correlated to the TNM stages of GC. According to the published dataset, the expression levels of both LIMK1 and LIMK2 were correlated to the overall survival time of GC patients. The level of Cofilin-1 was significantly different between GCs of different TNM stages. Moreover, most importantly, this is the first study to reveal that the level of Cofilin-1 is higher, and the level of p-Cofilin-1 is lower in the diffuse type of GC compared to that in intestinal type. Taken together, our study demonstrated that multiple factors in LIMK/Cofilin pathway including LIMK1, LIMK2, Cofilin-1, and p-Cofilin-1 were associated with the clinical and pathological features of GC, which is potentially helpful for the diagnosis and treatment of GC.

Magic angle spinning NMR structure of human cofilin-2 assembled on actin filaments reveals isoform-specific conformation and binding mode.

Actin polymerization dynamics regulated by actin-binding proteins are essential for various cellular functions. The cofilin family of proteins are potent regulators of actin severing and filament disassembly. The structural basis for cofilin-isoform-specific severing activity is poorly understood as their high-resolution structures in complex with filamentous actin (F-actin) are lacking. Here, we present the atomic-resolution structure of the muscle-tissue-specific isoform, cofilin-2 (CFL2), assembled on ADP-F-actin, determined by magic-angle-spinning (MAS) NMR spectroscopy and data-guided molecular dynamics (MD) simulations. We observe an isoform-specific conformation for CFL2. This conformation is the result of a unique network of hydrogen bonding interactions within the α2 helix containing the non-conserved residue, Q26. Our results indicate F-site interactions that are specific between CFL2 and ADP-F-actin, revealing mechanistic insights into isoform-dependent F-actin disassembly.

Estradiol inhibits HIV-1BaL infection and induces CFL1 expression in peripheral blood mononuclear cells and endocervical mucosa.

An inhibitory effect of estradiol (E2) on HIV-1 infection was suggested by several reports. We previously identified increased gene expression of actin-binding protein cofilin 1 (CFL1) in endocervix in the E2-dominated proliferative phase of the menstrual cycle. Actin cytoskeleton has an integral role in establishing and spreading HIV-1 infection. Herein, we studied in vitro effects of E2 on HIV-1 infection and on CFL1 expression to gain insight into the mechanism of HIV-1 inhibition by E2. E2 dose-dependently inhibited HIV-1BaL infection in peripheral blood mononuclear cells (PBMCs) and endocervix. In PBMCs and endocervix, E2 increased protein expression of total CFL1 and phosphorylated CFL1 (pCFL1) and pCFL1/CFL1 ratios. LIMKi3, a LIM kinase 1 and 2 inhibitor, abrogated the phenotype and restored infection in both PBMCs and endocervix; inhibited E2-induced expression of total CFL1, pCFL1; and decreased pCFL1/CFL1 ratios. Knockdown of CFL1 in PBMCs also abrogated the phenotype and partially restored infection. Additional analysis of soluble mediators revealed decreased concentrations of pro-inflammatory chemokines CXCL10 and CCL5 in infected tissues incubated with E2. Our results suggest a link between E2-mediated anti-HIV-1 activity and expression of CFL1 in PBMCs and endocervical mucosa. The data support exploration of cytoskeletal signaling pathway targets for the development of prevention strategies against HIV-1.

Cellular heterogeneity and transcriptomic profiles during intrahepatic cholangiocarcinoma initiation and progression.

BACKGROUND AND AIMS: Intrahepatic cholangiocarcinoma (ICC) is not fully investigated, and how stromal cells contribute to ICC formation is poorly understood. We aimed to uncover ICC origin, cellular heterogeneity, and critical modulators during ICC initiation/progression, and to decipher how fibroblast and endothelial cells in the stromal compartment favor ICC progression. APPROACH AND RESULTS: We performed single-cell RNA sequencing (scRNA-seq) using AKT/Notch intracellular domain-induced mouse ICC tissues at early, middle, and late stages. We analyzed the transcriptomic landscape, cellular classification and evolution, and intercellular communication during ICC initiation/progression. We confirmed the findings using quantitative real-time PCR, western blotting, immunohistochemistry or immunofluorescence, and gene knockout/knockdown analysis. We identified stress-responding and proliferating subpopulations in late-stage mouse ICC tissues and validated them using human scRNA-seq data sets. By integrating weighted correlation network analysis and protein-protein interaction through least absolute shrinkage and selection operator regression, we identified zinc finger, MIZ-type containing 1 (Zmiz1) and Y box protein 1 (Ybx1) as core transcription factors required by stress-responding and proliferating ICC cells, respectively. Knockout of either one led to the blockade of ICC initiation/progression. Using two other ICC mouse models (YAP/AKT, KRAS/p19) and human ICC scRNA-seq data sets, we confirmed the orchestrating roles of Zmiz1 and Ybx1 in ICC occurrence and development. In addition, hes family bHLH transcription factor 1, cofilin 1, and inhibitor of DNA binding 1 were identified as driver genes for ICC. Moreover, periportal liver sinusoidal endothelial cells could differentiate into tip endothelial cells to promote ICC development, and this was Dll4-Notch4-Efnb2 signaling-dependent. CONCLUSIONS: Stress-responding and ICC proliferating subtypes were identified, and Zmiz1 and Ybx1 were revealed as core transcription factors in these subtypes. Fibroblast-endothelial cell interaction promotes ICC development.

E2F4-induced AGAP2-AS1 up-regulation accelerates the progression of colorectal cancer via miR-182-5p/CFL1 axis.

BACKGROUND: Long non-coding RNAs (lncRNAs) are closely associated with the pathogenesis of numerous diseases including cancers. LncRNA AGAP2 Antisense RNA 1 (AGAP2-AS1) has been found to participate in the tumorigenesis of several kinds of human cancers. Nonetheless, its potential function in colorectal cancer (CRC) was still poorly investigated. METHODS: The expression level of RNAs or proteins was assessed by RT-qPCR or western blot analysis. Functional experiments were performed to analyze the role of AGAP2-AS1 in CRC in vitro and in vivo. Mechanism investigations were fulfilled to determine the potential mechanism of the molecules. RESULTS: AGAP2-AS1 expression was significantly elevated in CRC cells and could be transcriptionally activated by E2F Transcription Factor 4 (E2F4). Down-regulated AGAP2-AS1 could weaken CRC cell growth, migration, invasion, and epithelial-mesenchymal transition (EMT). MicroRNA-182-5p (miR-182-5p) was the target downstream molecule of AGAP2-AS1. Furthermore, Cofilin 1 (CFL1) was proved as the target of miR-182-5p. Mechanically, AGAP2-AS1 could boost the CFL1 expression via competitively binding to miR-182-5p in CRC. Importantly, CFL1 restoration could counteract the in vitro and in vivo suppression of depleted AGAP2-AS1 on CRC progression. CONCLUSION: E2F4-stimulated AGAP2-AS1 aggravated CRC development through regulating miR-182-5p/CFL1 axis, implying that AGAP2-AS1 might become a potent new target for future therapies for CRC.

Effects and mechanisms of FBXO31 on Taxol chemoresistance in esophageal squamous cell carcinoma.

Taxol is commonly used chemotherapy regimen for esophageal squamous cell carcinoma (ESCC). Study of the underlying mechanisms of Taxol chemoresistance provides better understanding of esophageal cancer treatment and may provide a rational molecular target for diagnosis and intervention. Here we showed FBXO31, which was reported to be highly expressed in ESCC and significantly associated with poor prognosis, could regulate ESCC chemosensitivity to Taxol. Silencing of FBXO31 in ESCC cells sensitized cells to Taxol treatment, evidenced by FACS analysis and TUNEL assay, showing as an increased apoptotic population in FBXO31-knockdown cells compared to the control cells. The mass spectrometry data and coimmunoprecipitation results showed FBXO31 could bind with cofilin-1. Cofilin-1 knockdown in FBXO31-overexpression cells reversed FBXO31-induced suppression of cell apoptosis, suggesting FBXO31-mediated Taxol chemoresistance is associated with cofilin-1. Furthermore, in vivo experiments confirmed that knockdown of FBXO31 sensitized ESCC to Taxol treatment. This finding substantiated a pivotal role of FBOX31 in ESCC chemoresistance, indicating that FBXO31 may be a potential indicator or target for drug resistance in ESCC.