Gene Profiling of Mta1 Identifies Novel Gene Targets and Functions

BACKGROUND: Metastasis-associated protein 1 (MTA1), a master dual co-regulatory protein is found to be an integral part of NuRD (Nucleosome Remodeling and Histone Deacetylation) complex, which has indispensable transcriptional regulatory functions via histone deacetylation and chromatin remodeling. Emerging literature establishes MTA1 to be a valid DNA-damage responsive protein with a significant role in maintaining the optimum DNA-repair activity in mammalian cells exposed to genotoxic stress. This DNA-damage responsive function of MTA1 was reported to be a P53-dependent and independent function. Here, we investigate the influence of P53 on gene regulation function of Mta1 to identify novel gene targets and functions of Mta1. METHODS: Gene expression analysis was performed on five different mouse embryonic fibroblasts (MEFs) samples (i) the Mta1 wild type, (ii) Mta1 knock out (iii) Mta1 knock out in which Mta1 was reintroduced (iv) P53 knock out (v) P53 knock out in which Mta1 was over expressed using Affymetrix Mouse Exon 1.0 ST arrays. Further Hierarchical Clustering, Gene Ontology analysis with GO terms satisfying corrected p-value<0.1, and the Ingenuity Pathway Analysis were performed. Finally, RT-qPCR was carried out on selective candidate genes. SIGNIFICANCE/CONCLUSION: This study represents a complete genome wide screen for possible target genes of a coregulator, Mta1. The comparative gene profiling of Mta1 wild type, Mta1 knockout and Mta1 re-expression in the Mta1 knockout conditions define "bona fide" Mta1 target genes. Further extensive analyses of the data highlights the influence of P53 on Mta1 gene regulation. In the presence of P53 majority of the genes regulated by Mta1 are related to inflammatory and anti-microbial responses whereas in the absence of P53 the predominant target genes are involved in cancer signaling. Thus, the presented data emphasizes the known functions of Mta1 and serves as a rich resource which could help us identify novel Mta1 functions

Combination of photothermal, prodrug and tumor cell camouflage technologies for triple-negative breast cancer treatment

Triple-negative breast cancer (TNBC) remains the most challenging breast cancer subtype. In the presented work, we have combined several emerging technologies to build up a nanoplatform for TNBC treatment: photothermal therapy, prodrug design and tumor cell camouflage formulation. First, we synthesized a paclitaxel (PTX) based prodrug PTX-SS, and then conjugated it to the surface of gold nanorod (Au NR) @ mesoporous silica (MSN) core-shell nanoparticles (Au@MSN-NH2 NPs). Subsequently, doxorubicin (DOX) was loaded into the Au@PTXSS-MSN NPs and further coated with cell membranes isolated from MDA-MB-231 cells to form cell camouflaged Au@PTXSS-MSN/DOX@CM NPs. The Au@PTXSS-MSN/DOX@CM NPs exhibited very good DOX loading capacity and the prodrug strategy enabled the precise adjustability of PTX-SS loading to achieve the optimized ratio between PTX and DOX to maximize the synergistic effect of these two drugs, as well as enabled GSH-responsive intracellular drug release. More interestingly, the cell membrane coating not only protected the drug from premature release, but also significantly improved the targeting ability of NPs to breast cancer MDA-MB-231 cells. The NPs also showed good photothermal responsiveness with clear improvement in inhibiting MDA-MB231 cell proliferation under laser irradiation. The in vivo studies further confirmed the effectiveness of Au@PTXSS-MSN/DOX@CM NPs on TNBC tumor inhibition in 4T1 cell grafted tumor mice model. (c) 2021 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/)

Expression of Multiple Artificial MicroRNAs from a Chicken miRNA126-Based Lentiviral Vector

Background: The use of RNAi in both basic and translational research often requires expression of multiple siRNAs from the same vector. Methods/Principal Findings: We have developed a novel chicken miR126-based artificial miRNA expression system that can express one, two or three miRNAs from a single cassette in a lentiviral vector. We show that each of the miRNAs expressed from the same lentiviral vector is capable of potent inhibition of reporter gene expression in transient transfection and stable integration assays in chicken fibroblast DF-1 cells. Transduction of Vero cells with lentivirus expressing two or three different anti-influenza miRNAs leads to inhibition of influenza virus production. In addition, the chicken miR126-based expression system effectively inhibits reporter gene expression in human, monkey, dog and mouse cells. These results demonstrate that the flanking regions of a single primary miRNA can support processing of three different stem-loops in a single vector. Conclusions/Significance: This novel design expands the means to express multiple miRNAs from the same vector for potent and effective silencing of target genes and influenza virus.National Institutes of Health (U.S.) (Grant R01AI056267)Cobb-Vantress, inc

Ectopic PDX-1 Expression Directly Reprograms Human Keratinocytes along Pancreatic Insulin-Producing Cells Fate

BACKGROUND: Cellular differentiation and lineage commitment have previously been considered irreversible processes. However, recent studies have indicated that differentiated adult cells can be reprogrammed to pluripotency and, in some cases, directly into alternate committed lineages. However, although pluripotent cells can be induced in numerous somatic cell sources, it was thought that inducing alternate committed lineages is primarily only possible in cells of developmentally related tissues. Here, we challenge this view and analyze whether direct adult cell reprogramming to alternate committed lineages can cross the boundaries of distinct developmental germ layers. METHODOLOGY/PRINCIPAL FINDINGS: We ectopically expressed non-integrating pancreatic differentiation factors in ectoderm-derived human keratinocytes to determine whether these factors could directly induce endoderm-derived pancreatic lineage and β-cell-like function. We found that PDX-1 and to a lesser extent other pancreatic transcription factors, could rapidly and specifically activate pancreatic lineage and β-cell-like functional characteristics in ectoderm-derived human keratinocytes. Human keratinocytes transdifferentiated along the β cell lineage produced processed and secreted insulin in response to elevated glucose concentrations. Using irreversible lineage tracing for KRT-5 promoter activity, we present supporting evidence that insulin-positive cells induced by ectopic PDX-1 expression are generated in ectoderm derived keratinocytes. CONCLUSIONS/SIGNIFICANCE: These findings constitute the first demonstration of human ectoderm cells to endoderm derived pancreatic cells transdifferentiation. The study represents a proof of concept which suggests that transcription factors induced reprogramming is wider and more general developmental process than initially considered. These results expanded the arsenal of adult cells that can be used as a cell source for generating functional endocrine pancreatic cells. Directly reprogramming somatic cells into alternate desired tissues has important implications in developing patient-specific, regenerative medicine approaches

Prediction of Drought-Resistant Genes in Arabidopsis thaliana Using SVM-RFE

Background: Identifying genes with essential roles in resisting environmental stress rates high in agronomic importance. Although massive DNA microarray gene expression data have been generated for plants, current computational approaches underutilize these data for studying genotype-trait relationships. Some advanced gene identification methods have been explored for human diseases, but typically these methods have not been converted into publicly available software tools and cannot be applied to plants for identifying genes with agronomic traits. Methodology: In this study, we used 22 sets of Arabidopsis thaliana gene expression data from GEO to predict the key genes involved in water tolerance. We applied an SVM-RFE (Support Vector Machine-Recursive Feature Elimination) feature selection method for the prediction. To address small sample sizes, we developed a modified approach for SVM-RFE by using bootstrapping and leave-one-out cross-validation. We also expanded our study to predict genes involved in water susceptibility. Conclusions: We analyzed the top 10 genes predicted to be involved in water tolerance. Seven of them are connected to known biological processes in drought resistance. We also analyzed the top 100 genes in terms of their biological functions. Our study shows that the SVM-RFE method is a highly promising method in analyzing plant microarray data for studyin

Activation of PyMT in β Cells Induces Irreversible Hyperplasia, but Oncogene-Dependent Acinar Cell Carcinomas When Activated in Pancreatic Progenitors

It is unclear whether the cellular origin of various forms of pancreatic cancer involves transformation or transdifferentiation of different target cells or whether tumors arise from common precursors, with tumor types determined by the specific genetic alterations. Previous studies suggested that pancreatic ductal carcinomas might be induced by polyoma middle T antigen (PyMT) expressed in non-ductal cells. To ask whether PyMT transforms and transdifferentiates endocrine cells toward exocrine tumor phenotypes, we generated transgenic mice that carry tetracycline-inducible PyMT and a linked luciferase reporter. Induction of PyMT in β cells causes β-cell hyperplastic lesions that do not progress to malignant neoplasms. When PyMT is de-induced, β cell proliferation and growth cease; however, regression does not occur, suggesting that continued production of PyMT is not required to maintain the viable expanded β cell population. In contrast, induction of PyMT in early pancreatic progenitor cells under the control of Pdx1 produces acinar cell carcinomas and β-cell hyperplasia. The survival of acinar tumor cells is dependent on continued expression of PyMT. Our findings indicate that PyMT can induce exocrine tumors from pancreatic progenitor cells, but cells in the β cell lineage are not transdifferentiated toward exocrine cell types by PyMT; instead, they undergo oncogene-dependent hyperplastic growth, but do not require PyMT for survival

Relative Roles of Grey Squirrels, Supplementary Feeding, and Habitat in Shaping Urban Bird Assemblages

Non-native species are frequently considered to influence urban assemblages. The grey squirrel Sciurus carolinensis is one such species that is widespread in the UK and is starting to spread across Europe; it predates birds’ nests and can compete with birds for supplementary food. Using distance sampling across the urbanisation intensity gradient in Sheffield (UK) we test whether urban grey squirrels influence avian species richness and density through nest predation and competition for supplementary food sources. We also assess how urban bird assemblages respond to supplementary feeding. We find that grey squirrels slightly reduced the abundance of breeding bird species most sensitive to squirrel nest predation by reducing the beneficial impact of woodland cover. There was no evidence that grey squirrel presence altered relationships between supplementary feeding and avian assemblage structure. This may be because, somewhat surprisingly, supplementary feeding was not associated with the richness or density of wintering bird assemblages. These associations were positive during the summer, supporting advocacy to feed birds during the breeding season and not just winter, but explanatory capacity was limited. The amount of green space and its quality, assessed as canopy cover, had a stronger influence on avian species richness and population size than the presence of grey squirrels and supplementary feeding stations. Urban bird populations are thus more likely to benefit from investment in improving the availability of high quality habitats than controlling squirrel populations or increased investment in supplementary feeding

Localization and Androgen Regulation of Metastasis-Associated Protein 1 in Mouse Epididymis

BACKGROUND: Metastasis-associated protein 1 (MTA1), the founding member of the MTA family of genes, can modulate transcription by influencing the status of chromatin remodeling. Despite its strong correlation with the metastatic potential of cancer cells, MTA1 can also regulate crucial cellular pathways by modifying the acetylation status. We have previously reported the presence of MTA1/MTA1 in human and mouse testes, providing the evidence for its involvement in the regulation of testicular function during murine spermatogenesis. The objective of present study was to further assess the localization of MTA1 in mouse epididymis on both transcriptional and translational level, and then to explore whether MTA1 expression is regulated by androgens and postnatal epididymal development. METHODOLOGY/PRINCIPAL FINDINGS: Mice were deprived of circulating androgen by bilaterally castration and were then supplemented with exogenous testosterone propionate for one week. MTA1 was immunolocalized in the epithelium of the entire epididymis with the maximal expression in the nuclei of principal cells and of clear cells in proximal region. Its expression decreased gradually after castration, whereas testosterone treatment could restore the expression, indicating that the expression of this gene is dependent on androgen. During postnatal development, the protein expression in the epididymis began to appear from day 7 to day 14, increased dramatically from postnatal day 28, and peaked at adulthood onwards, coinciding with both the well differentiated status of epididymis and the mature levels of circulating androgens. This region- and cell-specific pattern was also conservative in normal human epididymis. CONCLUSIONS: Our data suggest that the expression of MTA1 protein could be regulated by androgen pathway and its expression level is closely associated with the postnatal development of the epididymis, giving rise to the possibility that this gene plays a potential role in sperm maturation and fertility