Microenvironment-induced CREPT expression by cancer-derived small extracellular vesicles primes field cancerization.

Rationale: Cancer local recurrence increases the mortality of patients, and might be caused by field cancerization, a pre-malignant alteration of normal epithelial cells. It has been suggested that cancer-derived small extracellular vesicles (CDEs) may contribute to field cancerization, but the underlying mechanisms remain poorly understood. In this study, we aim to identify the key regulatory factors within recipient cells under the instigation of CDEs. Methods: In vitro experiments were performed to demonstrate that CDEs promote the expression of CREPT in normal epithelial cells. TMT-based quantitative mass spectrometry was employed to investigate the proteomic differences between normal cells and tumor cells. Loss-of-function approaches by CRISPR-Cas9 system were used to assess the role of CREPT in CDEs-induced field cancerization. RNA-seq was performed to explore the genes regulated by CREPT during field cancerization. Results: CDEs promote field cancerization by inducing the expression of CREPT in non-malignant epithelial cells through activating the ERK signaling pathway. Intriguingly, CDEs failed to induce field cancerization when CREPT was deleted, highlighting the importance of CREPT. Transcriptomic analyses revealed that CDEs elicited inflammatory responses, primarily through activation of the TNF signaling pathway. CREPT, in turn, regulates the transduction of downstream signals of TNF by modulating the expression of TNFR2 and PI3K, thereby promoting inflammation-to-cancer transition. Conclusion: CREPT not only serves as a biomarker for field cancerization, but also emerges as a target for preventing the cancer local recurrence.

Lachnochromonin, a fungal metabolite from Lachnum virgineum, inhibits cell growth and promotes apoptosis in tumor cells through JAK/STAT3 signaling.

Natural compounds that interfere with tumor cell growth have potential to be used as therapeutic agents to treat cancers. Lachnochromonin (p71) is a small molecule isolated from Lachnum virgineum. Here, we reported the effect of p71 on human tumor cells, especially on breast cancer MCF-7 cells. We found that p71 significantly suppresses cell growth and induces apoptosis. The luciferase results demonstrated that p71 specifically attenuates the activation of JAK/STAT3 signaling. Biochemical analysis revealed that p71 blocks the phosphorylation of STAT3 tyrosine 705 and serine 727, resulting in down-regulation of c-Myc and Cyclin D1 expression level. Importantly, p71 inhibited cell growth, colony-formation, and migration through affecting STAT3 activity. These results implied that p71 may be used as a therapeutic agent against breast cancer.

CREPT/RPRD1B promotes tumorigenesis through STAT3-driven gene transcription in a p300-dependent manner.

BACKGROUND: Signal transducer and activator of transcription 3 (STAT3) has been shown to upregulate gene transcription during tumorigenesis. However, how STAT3 initiates transcription remains to be exploited. This study is to reveal the role of CREPT (cell cycle-related and elevated-expression protein in tumours, or RPRD1B) in promoting STAT3 transcriptional activity. METHODS: BALB/c nude mice, CREPT overexpression or deletion cells were employed for the assay of tumour formation, chromatin immunoprecipitation, assay for transposase-accessible chromatin using sequencing. RESULTS: We demonstrate that CREPT, a recently identified oncoprotein, enhances STAT3 transcriptional activity to promote tumorigenesis. CREPT expression is positively correlated with activation of STAT3 signalling in tumours. Deletion of CREPT led to a decrease, but overexpression of CREPT resulted in an increase, in STAT3-initiated tumour cell proliferation, colony formation and tumour growth. Mechanistically, CREPT interacts with phosphorylated STAT3 (p-STAT3) and facilitates p-STAT3 to recruit p300 to occupy at the promoters of STAT3-targeted genes. Therefore, CREPT and STAT3 coordinately facilitate p300-mediated acetylation of histone 3 (H3K18ac and H3K27ac), further augmenting RNA polymerase II recruitment. Accordingly, depletion of p300 abolished CREPT-enhanced STAT3 transcriptional activity. CONCLUSIONS: We propose that CREPT is a co-activator of STAT3 for recruiting p300. Our study provides an alternative strategy for the therapy of cancers related to STAT3.

CREPT is required for murine stem cell maintenance during intestinal regeneration.

Intestinal stem cells (ISCs) residing in the crypts are critical for the continual self-renewal and rapid recovery of the intestinal epithelium. The regulatory mechanism of ISCs is not fully understood. Here we report that CREPT, a recently identified tumor-promoting protein, is required for the maintenance of murine ISCs. CREPT is preferably expressed in the crypts but not in the villi. Deletion of CREPT in the intestinal epithelium of mice (Vil-CREPTKO) results in lower body weight and slow migration of epithelial cells in the intestine. Vil-CREPTKO intestine fails to regenerate after X-ray irradiation and dextran sulfate sodium (DSS) treatment. Accordingly, the deletion of CREPT decreases the expression of genes related to the proliferation and differentiation of ISCs and reduces Lgr5+ cell numbers at homeostasis. We identify that CREPT deficiency downregulates Wnt signaling by impairing ß-catenin accumulation in the nucleus of the crypt cells during regeneration. Our study provides a previously undefined regulator of ISCs.

A safety consideration of mesenchymal stem cell therapy on COVID-19.

Due to the multi-potential differentiation and immunomodulatory function, mesenchymal stem cells (MSCs) have been widely used in the therapy of chronic and autoimmune diseases. Recently, the novel coronavirus disease 2019 (COVID-19) has grown to be a global public health emergency but no effective drug is available to date. Several studies investigated MSCs therapy for COVID-19 patients. However, it remains unclear whether MSCs could be the host cells of SARS-CoV-2 (severe acute respiratory syndrome coronavirus-2) and whether they might affect the SARS-CoV-2 entry into other cells. Here, we report that human MSCs barely express ACE2 and TMPRSS2, two receptors required for the virus endocytosis, indicating that MSCs are free from SARS-CoV-2 infection. Furthermore, we observed that MSCs were unable to induce the expression of ACE2 and TMPRSS2 in epithelial cells and macrophages. Importantly, under different inflammatory challenge conditions, implanted human MSCs failed to up-regulate the expression of ACE2 and TMPRSS2 in the lung tissues of mice. Intriguingly, we showed that a SARS-CoV-2 pseudovirus failed to infect MSCs and co-cultured MSCs did not increase the risk of SARS-CoV-2 pseudovirus infection in epithelial cells. All these results suggest that human MSCs have no risk of assisting SARS-CoV-2 infection and the use of MSCs as the therapy for COVID-19 patients is feasible and safe.

NOK associates with c-Src and promotes c-Src-induced STAT3 activation and cell proliferation.

Signal transducers and activators of transcription 3 (STAT3) is reported to regulate cell proliferation, survival, and differentiation, and thus plays a central role in development and carcinogenesis. Accumulating evidence demonstrated the involvement of cellular Src (c-Src) tyrosine kinase in the activation of STAT3. Additionally, novel oncogene with kinase-domain (NOK), a receptor protein tyrosine kinase that involves in cell transformation and tumorigenesis, was found to activate STAT3 signaling by a JAK2-dependent mechanism. However, whether the existence of the interaction between c-Src/STAT3 and NOK/STAT3 signals is still unknown. In this study, we showed that NOK formed a complex with c-Src and facilitated the interaction between c-Src and STAT3. In the complex, NOK greatly elevated the c-Src-mediated STAT3 activation by increasing the phosphorylation level of STAT3 on Tyr705. Truncated and mutation experiments further demonstrated that the kinase activity was responsible for the synergistic effect of NOK and c-Src on STAT3 activation. In addition, NOK and c-Src synergistically promoted cell proliferation and tumor growth in nude mice. Taken together, our results indicate that NOK associates with c-Src and promotes c-Src-induced STAT3 activation in a kinase-dependent manner. We proposed that the axis that NOK promoted c-Src-induced STAT3 activation is critical in cell proliferation and tumorigenesis.

Genome-Wide Analysis of Methylation-Driven Genes and Identification of an Eight-Gene Panel for Prognosis Prediction in Breast Cancer.

BACKGROUND: Aberrant DNA methylation is a crucial epigenetic regulator that is closely related to the occurrence and development of various cancers, including breast cancer (BC). The present study aimed to identify a novel methylation-based prognosis biomarker panel by integrally analyzing gene expression and methylation patterns in BC patients. METHODS: DNA methylation and gene expression data of breast cancer (BRCA) were downloaded from The Cancer Genome Atlas (TCGA). R packages, including ChAMP, SVA, and MethylMix, were applied to identify the unique methylation-driven genes. Subsequently, these genes were subjected to Metascape for GO analysis. Univariant Cox regression was used to identify survival-related genes among the methylation-driven genes. Robust likelihood-based survival modeling was applied to define the prognosis markers. An independent data set (GSE72308) was used for further validation of our risk score system. RESULTS: A total of 879 DNA methylation-driven genes were identified from 765 BC patients. In the discovery cohort, we identified 50 survival-related methylation-driven genes. Finally, we built an eight-methylation-driven gene panel that serves as prognostic predictors. CONCLUSIONS: Our analysis of transcriptome and methylome variations associated with the survival status of BC patients provides a further understanding of basic biological processes and a basis for the genetic etiology in BC.

Transcription Factor Myeloid Zinc-Finger 1 Suppresses Human Gastric Carcinogenesis by Interacting with Metallothionein 2A.

PURPOSE: Metallothionein 2A (MT2A) suppresses the progression of human gastric cancer potentially through an "MT2A-NF-κB pathway" with unclear mechanisms. This study explored the role of a transcription factor, myeloid zinc-finger 1 (MZF1), in MT2A-NF-κB pathway and its clinical significance in gastric cancer. EXPERIMENTAL DESIGN: MZF1 expression and function in gastric cancer were investigated in vitro and in vivo. The relationship between MZF1 and MT2A was determined by gain-of-function and loss-of-function assays in gastric cancer cells and an immortalized gastric cell line GES-1. The prognostic value of MZF1 expression in association with MT2A was evaluated using IHC in two cohorts. RESULTS: MZF1 was epigenetically silenced in human gastric cancer cell lines and primary tumors. Overexpression of MZF1 in gastric cancer cells suppressed cell proliferation and migration, as well as the growth of xenograft tumors in nude mice. Knocking-down of MZF1 transformed GES-1 cells into a malignant phenotype characterized by increased cell growth and migration. Mechanistically, MZF1 was upregulated in both GC and GES-1 cells by MT2A ectopically expressed or induced upon treatment with a garlic-derived compound, diallyl trisulfide (DATS). MZF1 associated with MT2A was colocalized in the nuclei of GES-1 cells to target the promoter of NF-κB inhibitor alpha (NFKBIA). Clinically, MT2A and MZF1 were progressively downregulated in clinical specimens undergoing gastric malignant transformation. Downregulation of MT2A and MZF1 was significantly correlated with poorer patient prognosis. CONCLUSIONS: MT2A exerts its anti-gastric cancer effects by complexing with MZF1 to target NFKBIA. MT2A/MZF1 may serve as a valuable prognostic marker and a novel therapeutic target for human gastric cancer.

CREPT/RPRD1B associates with Aurora B to regulate Cyclin B1 expression for accelerating the G2/M transition in gastric cancer.

Gastric cancer, like most of other cancers, has an uncontrolled cell cycle regulated by cyclins and cyclin-dependent kinases (CDKs). In this study, we reported that gastric cancer cells showed an accelerated G2/M transition promoted by CREPT/RPRD1B and Aurora kinase B (Aurora B). We found that CREPT/RPRD1B and Aurora B were coordinately expressed during the cell cycle in gastric cancer cells. Deletion of CREPT/RPRD1B disturbed the cell progression and extended the length of cell cycle, leading to a significant accumulation of mitotic cells. Mechanistically, we revealed that CREPT/RPRD1B interacted with Aurora B to regulate the expression of Cyclin B1 in gastric cancer cells. Interestingly, Aurora B phosphorylates S145 in a well-conserved motif of CREPT/RPRD1B. We proposed that phosphorylation of CREPT/RPRD1B by Aurora B is required for promoting the transcription of Cyclin B1, which is critical for the regulation of gastric tumorigenesis. Our study provides a mechanism by which gastric tumor cells maintain their high proliferation rate via coordination of Aurora B and CREPT/RPRD1B on the expression of Cyclin B1. Targeting the interaction of Aurora B and CREPT/RPRD1B might be a strategy for anti-gastric cancer therapy in the future.

Mesenchymal stromal cells ameliorate acute allergic rhinitis in rats.

Mesenchymal stromal cells (MSCs) have been extensively investigated as a potential antiinflammatory treatment in many inflammatory-related diseases; however, it remains unclear whether MSCs could be used to treat acute allergic rhinitis. A rat model of allergic rhinitis was treated with MSCs. The effect of MSCs on the inflammation of allergic rhinitis was evaluated by sneezing, nose rubbing, the pathology of the nasal mucosa, and the expression of interleukin 4, tumour necrosis factor alpha, and immunoglobulin E in the serum of rats. Also, the population of MSCs isolated from umbilical cords of humans was evaluated to determine if they could inhibit the symptoms and inflammation of acute allergic rhinitis in a rat model. We observed that this population of cells inhibited sneezing, nose rubbing, and changes in the pathology of the nasal mucosa. Intriguingly, we observed that MSCs reduced the expression of interleukin 4, tumour necrosis factor alpha, and immunoglobulin E in the serum. Furthermore, MSCs reduced the expression of histamine and the recruitment of macrophages in the nasal mucosa of allergic rhinitis rats. We reasoned that the effect of MSCs on allergic rhinitis might be through its regulation of the secretion of related cytokines from macrophages during the process of acute allergic rhinitis. This work suggested that MSCs from the umbilical cords of humans could be used as a positive clinical therapy for the human disease.

REGγ is critical for skin carcinogenesis by modulating the Wnt/ß-catenin pathway.

Here we report that mice deficient for the proteasome activator, REGγ, exhibit a marked resistance to TPA (12-O-tetradecanoyl-phorbol-13-acetate)-induced keratinocyte proliferation, epidermal hyperplasia and onset of papillomas compared with wild-type counterparts. Interestingly, a massive increase of REGγ in skin tissues or cells resulting from TPA induces activation of p38 mitogen-activated protein kinase (MAPK/p38). Blocking p38 MAPK activation prevents REGγ elevation in HaCaT cells with TPA treatment. AP-1, the downstream effector of MAPK/p38, directly binds to the REGγ promoter and activates its transcription in response to TPA stimulation. Furthermore, we find that REGγ activates Wnt/ß-catenin signalling by degrading GSK-3ß in vitro and in cells, increasing levels of CyclinD1 and c-Myc, the downstream targets of ß-catenin. Conversely, MAPK/p38 inactivation or REGγ deletion prevents the increase of cyclinD1 and c-Myc by TPA. This study demonstrates that REGγ acts in skin tumorigenesis mediating MAPK/p38 activation of the Wnt/ß-catenin pathway.

Oxidative challenge enhances REGγ-proteasome-dependent protein degradation.

Elimination of oxidized proteins is important to cells as accumulation of damaged proteins causes cellular dysfunction, disease, and aging. Abundant evidence shows that the 20S proteasome is largely responsible for degradation of oxidative proteins in both ubiquitin-dependent and ubiquitin-independent pathways. However, the role of the REGγ-proteasome in degrading oxidative proteins remains unclear. Here, we focus on two of the well-known REGγ-proteasome substrates, p21(Waf1/Cip1) and hepatitis C virus (HCV) core protein, to analyze the impact of oxidative stress on REGγ-proteasome functions. We demonstrate that REGγ-proteasome is essential for oxidative stress-induced rapid degradation of p21 and HCV proteins. Silencing REGγ abrogated this response in multiple cell lines. Furthermore, pretreatment with proteasome inhibitor MG132 completely blunted oxidant-induced p21 degradation, indicating a proteasome-dependent action. Cellular oxidation promoted REGγ-proteasome-dependent trypsin-like activity by enhancing the interaction between REGγ and 20S proteasome. Antioxidant could counteract oxidation-induced protein degradation, indicating that REGγ-proteasome activity may be regulated by redox state. This study provides further insights into the actions of a unique proteasome pathway in response to an oxidative stress environment, implying a novel molecular basis for REGγ-proteasome functions in antioxidation.