scholarly journals Transcription Factor RFX3 Stabilizes Mammary Basal Cell Identity

2021 ◽  
Author(s):  
Ian Macara ◽  
Erica M. Tross ◽  
Christian de Caestecker ◽  
Ken Lau
Keyword(s):  
Mammary Gland ◽  
Basal Cell ◽  
Regulatory Networks ◽  
Cell Tracking ◽  
Molecular Mechanisms ◽  
Myoepithelial Cells ◽  
Mammary Fat Pad ◽  
Transplantation Assay ◽  
Differential Gene

The myoepithelial cell compartment of the murine postnatal mammary gland is generated from basal cap cells in the terminal end bud and maintained by self-renewal. Transdifferentiation to the luminal lineage does not normally occur but can be induced by DNA damage, luminal cell death or transplantation into a recipient mammary fat pad. Myoepithelial cells cultivated in vitro can also transdifferentiate towards the luminal lineage. Little is known about the molecular mechanisms and gene regulatory networks underlying this plasticity. Using a transgenic mouse (Tg11.5kb-GFP) that marks cap cells with GFP, we discovered that mature myoepithelial cells placed in culture begin to express GFP within ~24 hrs and later express the Keratin 8 (K8) luminal marker. Cell tracking showed that most K8+ cells arose from GFP+ cells, suggesting that myoepithelial cells de-differentiate towards a progenitor state before changing lineage. Differential gene expression analysis, comparing pure GFP+ cap cells with mature myoepithelial cells, identified multiple transcription factors that iRegulon predicted might regulate the myoepithelial to cap cell transition. Knockout of one of these genes, Regulatory Factor 3 (Rfx3), significantly reduced the population of GFP+ cells and increased differentiation to the K8+ luminal lineage. Rfx3 knockout also reduced mammosphere growth and mammary gland regeneration efficiency in a transplantation assay, but had no effect on proliferation in vitro. Together, these data support a key role for Rfx3 in the stabilization of the mammary basal cell lineages.

scholarly journals Discovering therapeutic activities from venoms using differential gene expression

2019 ◽  
Author(s):  
Joseph D. Romano ◽  
Hai Li ◽  
Ronald Realubit ◽  
Charles Karan ◽  
Nicholas P. Tatonetti
Keyword(s):  
Gene Expression ◽  
Differential Gene Expression ◽  
Regulatory Networks ◽  
Preliminary Evidence ◽  
Natural Toxins ◽  
Differential Gene ◽  
Public Repositories ◽  
Therapeutic Activities ◽  
Validation Experiments

AbstractVenoms are a diverse and complex group of natural toxins that have been adapted to treat many types of human disease, but rigorous informatics approaches for discovering new therapeutic activities are scarce. We have designed and validated a new platform—named VenomSeq—to systematically generate putative associations between venoms and drugs/diseases via high-throughput transcriptomics and perturbational differential gene expression analysis. In this study, we describe the architecture of VenomSeq, and its evaluation using the crude venoms from 25 diverse animal species. By integrating comparisons to public repositories of differential expression, associations between regulatory networks and disease, and existing knowledge of venom activity, we provide a number of new therapeutic hypotheses linking venoms to human diseases supported by multiple layers of preliminary evidence. We are currently performing validation experiments in vitro to corroborate these findings.

scholarly journals Protocol: Adeno-Associated Virus-Mediated Gene Transfer in Ex Vivo Cultured Embryonic Mammary Gland

2020 ◽  
Author(s):  
Qiang Lan ◽  
Marja L. Mikkola
Keyword(s):  
Mammary Gland ◽  
Gene Transfer ◽  
Regulatory Networks ◽  
Molecular Mechanisms ◽  
Ex Vivo ◽  
Transcriptional Profiling ◽  
Branching Morphogenesis ◽  
Culture Technique ◽  
Adeno Associated Virus ◽  
Profiling Techniques

Abstract Branching morphogenesis of the murine mammary gland starts during late embryogenesis. It is regulated by the signals emanating both from the epithelium and the mesenchyme, yet the molecular mechanisms regulating this process remain poorly understood. We have previously developed a unique whole organ culture technique for embryonic mammary glands, which provides a powerful tool to monitor and manipulate branching morphogenesis ex vivo. Nowadays, RNA sequencing and other transcriptional profiling techniques provide robust methods to identify components of gene regulatory networks driving branching morphogenesis. However, validation of the candidate genes still mainly depends on the use of the transgenic mouse models, especially in mammary gland studies. By comparing different serotypes of recombinant adeno-associated virus (rAAVs), we found out that rAAVs provide sufficient efficiency for gene transfer with different tissue preferences depending on the serotypes of the virus. AAV-2 and AAV-8 preferentially target epithelial and mesenchymal compartments, respectively, while AAV-9 infects both tissues. Here, we describe a protocol for AAV-mediated gene transfer in ex vivo cultured murine embryonic mammary gland facilitating gene function studies on mammary gland branching morphogenesis.

Uncovering of Key Pathways and miRNAs for Intracranial Aneurysm Based on Weighted Gene Co-Expression Network Analysis

2022 ◽  
pp. 1-12
Author(s):  
Zhengfei Ma ◽  
Ping Zhong ◽  
Peidong Yue ◽  
Zhongwu Sun
Keyword(s):  
Network Analysis ◽  
Intracranial Aneurysm ◽  
Differentially Expressed Genes ◽  
Regulatory Networks ◽  
Molecular Mechanisms ◽  
Area Under The Curve ◽  
Enrichment Analysis ◽  
Functional Enrichment ◽  
Differentially Expressed

<b><i>Background:</i></b> Intracranial aneurysm (IA) is a serious cerebrovascular disease. The identification of key regulatory genes can provide research directions for early diagnosis and treatment of IA. <b><i>Methods:</i></b> Initially, the miRNA and mRNA data were downloaded from the Gene Expression Omnibus database. Subsequently, the limma package in R was used to screen for differentially expressed genes. In order to investigate the function of the differentially expressed genes, a functional enrichment analysis was performed. Moreover, weighted gene co-expression network analysis (WGCNA) was performed to identify the hub module and hub miRNAs. The correlations between miRNAs and mRNAs were assessed by constructing miRNA-mRNA regulatory networks. In addition, in vitro validation was performed. Finally, diagnostic analysis and electronic expression verification were performed on the GSE122897 dataset. <b><i>Results:</i></b> In the present study, 955 differentially expressed mRNAs (DEmRNAs, 480 with increased and 475 with decreased expression) and 46 differentially expressed miRNAs (DEmiRNAs, 36 with increased and 10 with decreased expression) were identified. WGCNA demonstrated that the yellow module was the hub module. Moreover, 16 hub miRNAs were identified. A total of 1,124 negatively regulated miRNA-mRNA relationship pairs were identified. Functional analysis demonstrated that DEmRNAs in the targeted network were enriched in vascular smooth muscle contraction and focal adhesion pathways. In addition, the area under the curve of 16 hub miRNAs was &#x3e;0.8. It is implied that 16 hub miRNAs may be used as potential diagnostic biomarkers of IA. <b><i>Conclusion:</i></b> Hub miRNAs and key signaling pathways were identified by bioinformatics analysis. This evidence lays the foundation for understanding the underlying molecular mechanisms of IA and provided potential therapeutic targets for the treatment of this disease.

scholarly journals CBIO-20. MOLECULAR MECHANISMS OF OLIG2 TRANSCRIPTION FACTOR IN GLIOBLASTOMA

2020 ◽  
Vol 22 (Supplement_2) ◽  
pp. ii19-ii20
Author(s):  
Norihiko Saito ◽  
Sho Sato ◽  
Yu Hiramoto ◽  
Satoshi Fujita ◽  
Haruo Nakayama ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Transcription Factor ◽  
Regulatory Networks ◽  
Cell Cycle Progression ◽  
Molecular Mechanisms ◽  
Cell Cycle Control ◽  
Neural Lineage ◽  
Lineage Differentiation ◽  
Neural Stem Progenitor Cells

Abstract Oligodendrocyte lineage transcription factor 2 (OLIG2) promotes proliferation of normal neural stem/progenitor cells and glioma cells. However, the mechanisms underlying the regulation of OLIG2 remain largely unknown. Here, we show that a comprehensive analysis of the critical gene regulatory networks involving OLIG2 in glioma initiating cell (GIC) lines. In vitro differentiation studies showed that proneural GIC lines possess the potential to differentiate into astrocytic, neuronal, and oligodendrocytic lineages, whereas mesenchymal GICs exhibited limited potential for neural lineage differentiation following retinoic acid induction. We also showed that CDK2-mediated OLIG2 phosphorylation stabilizes OLIG2 protein from proteasomal degradation. Phosphorylated OLIG2 binds to the E-Box regions of p27 promoter and represses p27 transcription, which in turn activates CDK2 in positive feedback manner. CDK2-mediated OLIG2 phosphorylation promotes cell cycle progression, cell proliferation, and tumorigenesis. OLIG2 inhibition disrupted cell cycle control mechanism by decreasing CDK2 and elevating apoptosis-related molecules. Inhibition of CDK2 activity disrupted OLIG2-CDK2 interactions and attenuated OLIG2 protein stability.　In addition, OLIG2-high glioma initiating cells are highly sensitive to CDK2 inhibitor treatment, indicating that OLIG2 can be a biomarker for personalized treatment for glioblastoma patients with CDK2 inhibitors. In conclusion, we have identified OLIG2-CDK2 interactions in glioma stem cells that can be targeted by CDK2 inhibitors and this may allow the selection of patients with high likelihood of responding to this therapy.

ΔNp63α is a super enhancer-enriched master factor controlling the basal-to-luminal differentiation transcriptional program and gene regulatory networks in nasopharyngeal carcinoma

2019 ◽  
Vol 41 (9) ◽  
pp. 1282-1293 ◽  
Author(s):  
Jing Cai ◽  
Shengnan Chen ◽  
Mei Yi ◽  
Yixin Tan ◽  
Qian Peng ◽  
...  
Keyword(s):  
Gene Expression ◽  
Nasopharyngeal Carcinoma ◽  
Basal Cell ◽  
Regulatory Networks ◽  
Regulation Of Gene Expression ◽  
Gene Expression Signature ◽  
Molecular Signatures ◽  
Chip Sequencing

Abstract Nasopharyngeal carcinoma (NPC) originates via malignant transformation of the pseudostratified nasopharyngeal epithelium, composed of basal and luminal cells. Super enhancers (SEs) are large clusters of cis-elements involved in the regulation of gene expression through epigenetic regulatory mechanisms. In this study, we demonstrated that basal cell-specific proteins are highly expressed, whereas luminal cell proteins are downregulated in NPC, implying a perturbation of basal-to-luminal differentiation during NPC development. We characterized NPC cell models according to different molecular signatures associated with their differentiation status and found that distinct SE landscapes are tightly associated with basal or luminal-like molecular signatures in NPC cells. Furthermore, the transcription of ΔNP63α, a prominent isoform of TP63, was found to be driven by SEs in NPC cells. Data from chromatin immunoprecipitation (ChIP)-sequencing showed that ΔNP63α largely occupied regions of SEs associated with basal cell-specific genes. Silencing of ΔNP63α led to a loss of H3K27ac occupancy at basal-type SEs and triggered a basal-to-luminal gene expression signature switch, suggesting that ΔNP63α is a master factor contributing to the perturbation of luminal differentiation. Integrative transcriptomics analysis also revealed that ΔNP63α acts as a core factor involved in the dysregulation of gene expression in NPC. Furthermore, ΔNP63α enhanced EGF-stimulated NF-κB activation in NPC cells by activating SE-mediated EGFR transcription. Finally, depletion of ΔNP63α in NPC cells induced robust growth inhibition of NPC cells in vitro and in vivo. Our data revealed that ΔNP63α-dependent SE reprogramming contributes to the blockade of luminal differentiation and uncontrolled proliferation in NPC.

scholarly journals Immunocytochemical identification of proliferating cell types in mouse mammary gland.

1990 ◽  
Vol 38 (11) ◽  
pp. 1541-1547 ◽  
Author(s):  
A Sapino ◽  
L Macrì ◽  
P Gugliotta ◽  
G Bussolati
Keyword(s):  
Cell Proliferation ◽  
Mammary Gland ◽  
Smooth Muscle Actin ◽  
Cell Types ◽  
Myoepithelial Cells ◽  
Mouse Mammary Gland ◽  
Staining Procedure ◽  
Secretory Cells ◽  
Alpha Smooth Muscle Actin

To study cell proliferation in different cell types and segments of the mammary gland, we devised a dual staining procedure, combining nuclear labeling by 5-bromo-2'-deoxy-uridine (BrdU) uptake (revealed by a dark-brown precipitate) and an alternative (red or blue) cytoplasmic labeling by antibodies specific for the differentiation proteins of epithelial, myoepithelial, and secretory cell types. The following markers, revealed by APAAP or beta-galactosidase procedure, were selected: alpha-smooth muscle actin for the myoepithelial cells, keratin (detected by AE1 monoclonal) for the luminal epithelial cells, alpha-lactalbumin and beta-casein for the secretory cells. To follow the full process of organogenesis, the study was conducted in mouse mammary glands from virgin, primed, and lactating animals and from glands cultured in vitro under specific hormone stimulation. Cell proliferation was localized mainly in focal areas (end buds), and mostly corresponded to "null" undifferentiated cells. Estrogen and progestin stimulation induced a relative increase of proliferating differentiated cells of either epithelial or myoepithelial type, localized in ducts and alveolar structures. Prolactin stimulation induced proliferation in secretory cells.

scholarly journals Differential Gene Expression Between the Biotrophic-Like and Saprotrophic Mycelia of the Witches' Broom Pathogen Moniliophthora perniciosa

2008 ◽  
Vol 21 (7) ◽  
pp. 891-908 ◽  
Author(s):  
Johana Rincones ◽  
Leandra M. Scarpari ◽  
Marcelo F. Carazzolle ◽  
Jorge M. C. Mondego ◽  
Eduardo F. Formighieri ◽  
...  
Keyword(s):  
Gene Expression ◽  
Oxidative Phosphorylation ◽  
Differential Gene Expression ◽  
Molecular Mechanisms ◽  
Expressed Sequence Tag ◽  
Lower Growth ◽  
Moniliophthora Perniciosa ◽  
Differential Gene ◽  
Combined Strategy

Moniliophthora perniciosa is a hemibiotrophic fungus that causes witches' broom disease (WBD) in cacao. Marked dimorphism characterizes this fungus, showing a monokaryotic or biotrophic phase that causes disease symptoms and a later dikaryotic or saprotrophic phase. A combined strategy of DNA microarray, expressed sequence tag, and real-time reverse-transcriptase polymerase chain reaction analyses was employed to analyze differences between these two fungal stages in vitro. In all, 1,131 putative genes were hybridized with cDNA from different phases, resulting in 189 differentially expressed genes, and 4,595 reads were clusterized, producing 1,534 unigenes. The analysis of these genes, which represent approximately 21% of the total genes, indicates that the biotrophic-like phase undergoes carbon and nitrogen catabolite repression that correlates to the expression of phytopathogenicity genes. Moreover, downregulation of mitochondrial oxidative phosphorylation and the presence of a putative ngr1 of Saccharomyces cerevisiae could help explain its lower growth rate. In contrast, the saprotrophic mycelium expresses genes related to the metabolism of hexoses, ammonia, and oxidative phosphorylation, which could explain its faster growth. Antifungal toxins were upregulated and could prevent the colonization by competing fungi. This work significantly contributes to our understanding of the molecular mechanisms of WBD and, to our knowledge, is the first to analyze differential gene expression of the different phases of a hemibiotrophic fungus.

Characterization in vitro of luminal and myoepithelial cells isolated from the human mammary gland by cell sorting

1991 ◽  
Vol 46 (3) ◽  
pp. 209-221 ◽  
Author(s):  
Michael J. O'Hare ◽  
Michael G. Ormerod ◽  
Paul Monaghan ◽  
E. Birgitte Lane ◽  
Barry A. Gusterson
Keyword(s):  
Mammary Gland ◽  
Cell Sorting ◽  
Myoepithelial Cells ◽  
Human Mammary Gland ◽  
Luminal And Myoepithelial Cells

Immunohistochemical investigation into the distribution pattern of myoepithelial cells in the bovine mammary gland

1997 ◽  
Vol 64 (2) ◽  
pp. 197-205 ◽  
Author(s):  
EVA HELLMÉN ◽  
AXEL ISAKSSON
Keyword(s):  
Smooth Muscle ◽  
Mammary Gland ◽  
Distribution Pattern ◽  
Basal Cell ◽  
Cell Layer ◽  
Smooth Muscle Actin ◽  
Myoepithelial Cells ◽  
Muscle Actin ◽  
Bovine Mammary Gland ◽  
Basal Cell Layer

The distribution pattern of myoepithelial cells in the bovine mammary gland was investigated by an immunohistochemical technique, using monoclonal antibodies against cytokeratins 5, 6 and 18 and cytokeratins 8 and 14 and against α-smooth-muscle actin filaments. Myoepithelial cells were shown to be present as a continuous basal cell layer in the intralobular ducts, as discontinuous cell rows in the basal cell layer of the interlobular ducts, and as single cells dispersed in the basal cell layer of the quarter cisterns, while they were apparently absent in the teat cisterns. Unlike the case with myoepithelial cells of the human breast, anti-cytokeratin 14 was less specific as a marker of bovine myoepithelial cells than was anti-α-smooth-muscle actin.

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