scholarly journals NuMA Influences Higher Order Chromatin Organization in Human Mammary Epithelium

2007 ◽  
Vol 18 (2) ◽  
pp. 348-361 ◽  
Author(s):  
Patricia C. Abad ◽  
Jason Lewis ◽  
I. Saira Mian ◽  
David W. Knowles ◽  
Jennifer Sturgis ◽  
...  
Keyword(s):  
Mammary Epithelial Cells ◽  
Sequence Similarity ◽  
Three Dimensional ◽  
Coiled Coil ◽  
Mammary Epithelium ◽  
Chromatin Organization ◽  
Mammary Epithelial ◽  
Cell Phenotype ◽  
Deoxyribonuclease I ◽  
C Terminus

The coiled-coil protein NuMA is an important contributor to mitotic spindle formation and stabilization. A potential role for NuMA in nuclear organization or gene regulation is suggested by the observations that its pattern of nuclear distribution depends upon cell phenotype and that it interacts and/or colocalizes with transcription factors. To date, the precise contribution of NuMA to nuclear function remains unclear. Previously, we observed that antibody-induced alteration of NuMA distribution in growth-arrested and differentiated mammary epithelial structures (acini) in three-dimensional culture triggers the loss of acinar differentiation. Here, we show that in mammary epithelial cells, NuMA is present in both the nuclear matrix and chromatin compartments. Expression of a portion of the C terminus of NuMA that shares sequence similarity with the chromatin regulator HPC2 is sufficient to inhibit acinar differentiation and results in the redistribution of NuMA, chromatin markers acetyl-H4 and H4K20m, and regions of deoxyribonuclease I-sensitive chromatin compared with control cells. Short-term alteration of NuMA distribution with anti-NuMA C-terminus antibodies in live acinar cells indicates that changes in NuMA and chromatin organization precede loss of acinar differentiation. These findings suggest that NuMA has a role in mammary epithelial differentiation by influencing the organization of chromatin.

scholarly journals ErbB4 Splice Variants Cyt1 and Cyt2 Differ by 16 Amino Acids and Exert Opposing Effects on the Mammary Epithelium In Vivo

2009 ◽  
Vol 29 (18) ◽  
pp. 4935-4948 ◽  
Author(s):  
Rebecca S. Muraoka-Cook ◽  
Melissa A. Sandahl ◽  
Karen E. Strunk ◽  
Leah C. Miraglia ◽  
Carty Husted ◽  
...  
Keyword(s):  
Amino Acids ◽  
Mammary Epithelial Cells ◽  
Splice Variants ◽  
Three Dimensional ◽  
Mammary Epithelium ◽  
Mammary Epithelial ◽  
Lumen Formation ◽  
Transgenic Expression ◽  
Opposing Effects

ABSTRACT Data concerning the prognostic value of ErbB4 in breast cancer and effects on cell growth have varied in published reports, perhaps due to the unknown signaling consequences of expression of the intracellular proteolytic ErbB4 s80HER4 fragment or due to differing signaling capabilities of alternatively spliced ErbB4 isoforms. One isoform (Cyt1) contains a 16-residue intracellular sequence that is absent from the other (Cyt2). We expressed s80Cyt1 and s80Cyt2 in HC11 mammary epithelial cells, finding diametrically opposed effects on the growth and organization of colonies in three-dimensional matrices. Whereas expression of s80Cyt1 decreased growth and increased the rate of three-dimensional lumen formation, that of s80Cyt2 increased proliferation without promoting lumen formation. These results were recapitulated in vivo, using doxycycline-inducible, mouse breast-transgenic expression of s80Cyt1 amd s80Cyt2. Expression of s80Cyt1 decreased growth of the mammary ductal epithelium, caused precocious STAT5a activation and lactogenic differentiation, and increased cell surface E-cadherin levels. Remarkably, ductal growth inhibition by s80Cyt1 occurred simultaneously with lobuloalveolar growth that was unimpeded by s80Cyt1, suggesting that the response to ErbB4 may be influenced by the epithelial subtype. In contrast, expression of s80Cyt2 caused epithelial hyperplasia, increased Wnt and nuclear β-catenin expression, and elevated expression of c-myc and cyclin D1 in the mammary epithelium. These results demonstrate that the Cyt1 and Cyt2 ErbB4 isoforms, differing by only 16 amino acids, exhibit markedly opposing effects on mammary epithelium growth and differentiation.

Quantitative Model-Based Image Analysis of NuMa Distribution Links Nuclear Organization with Cell Phenotype

2001 ◽  
Vol 7 (S2) ◽  
pp. 578-579
Author(s):  
David W. Knowles ◽  
Sophie A. Lelièvre ◽  
Carlos Ortiz de Solόrzano ◽  
Stephen J. Lockett ◽  
Mina J. Bissell ◽  
...  
Keyword(s):  
Image Analysis ◽  
Mammary Epithelial Cells ◽  
Nuclear Organization ◽  
Critical Role ◽  
Quantitative Model ◽  
Mammary Epithelial ◽  
Cell Phenotype ◽  
Proliferating Cells ◽  
Mitotic Apparatus ◽  
Model Based

The extracellular matrix (ECM) plays a critical role in directing cell behaviour and morphogenesis by regulating gene expression and nuclear organization. Using non-malignant (S1) human mammary epithelial cells (HMECs), it was previously shown that ECM-induced morphogenesis is accompanied by the redistribution of nuclear mitotic apparatus (NuMA) protein from a diffuse pattern in proliferating cells, to a multi-focal pattern as HMECs growth arrested and completed morphogenesis . A process taking 10 to 14 days.To further investigate the link between NuMA distribution and the growth stage of HMECs, we have investigated the distribution of NuMA in non-malignant S1 cells and their malignant, T4, counter-part using a novel model-based image analysis technique. This technique, based on a multi-scale Gaussian blur analysis (Figure 1), quantifies the size of punctate features in an image. Cells were cultured in the presence and absence of a reconstituted basement membrane (rBM) and imaged in 3D using confocal microscopy, for fluorescently labeled monoclonal antibodies to NuMA (fαNuMA) and fluorescently labeled total DNA.

scholarly journals Fine-tuning of Epithelial EGFR signals Supports Coordinated Mammary Gland Development

2020 ◽  
Author(s):  
Alexandr Samocha ◽  
Hanna M. Doh ◽  
Vaishnavi Sitarama ◽  
Quy H. Nguyen ◽  
Oghenekevwe Gbenedio ◽  
...  
Keyword(s):  
Mammary Gland ◽  
Mammary Epithelial Cells ◽  
Mammary Epithelium ◽  
Gene Signature ◽  
Mammary Epithelial ◽  
Fine Tuning ◽  
Basal Cells ◽  
Stem And Progenitor Cells

SummaryDuring puberty, robust morphogenesis occurs in the mammary gland; stem- and progenitor-cells develop into mature basal- and luminal-cells to form the ductal tree. The receptor signals that govern this process in mammary epithelial cells (MECs) are incompletely understood. The EGFR has been implicated and here we focused on EGFR’s downstream pathway component Rasgrp1. We find that Rasgrp1 dampens EGF-triggered signals in MECs. Biochemically and in vitro, Rasgrp1 perturbation results in increased EGFR-Ras-PI3K-AKT and mTORC1-S6 kinase signals, increased EGF-induced proliferation, and aberrant branching-capacity in 3D cultures. However, in vivo, Rasgrp1 perturbation results in delayed ductal tree maturation with shortened branches and reduced cellularity. Rasgrp1-deficient MEC organoids revealed lower frequencies of basal cells, the compartment that incorporates stem cells. Molecularly, EGF effectively counteracts Wnt signal-driven stem cell gene signature in organoids. Collectively, these studies demonstrate the need for fine-tuning of EGFR signals to properly instruct mammary epithelium during puberty.

Effects of dietary fat on the growth of normal, preneoplastic and neoplastic mammary epithelial cells in vivo and in vitro

1985 ◽  
Vol 75 (1) ◽  
pp. 269-278 ◽  
Author(s):  
C.A. Carrington ◽  
H.L. Hosick
Keyword(s):  
Mammary Epithelial Cells ◽  
Saturated Fat ◽  
Mammary Epithelium ◽  
Mammary Epithelial ◽  
Dietary Fats ◽  
Polyunsaturated Fat ◽  
Fat Pad ◽  
Mammary Fat Pad

In order to determine: (1) whether there is a growth-regulating interaction between the mammary fat pad and mammary epithelium; (2) whether this interaction could be modified by dietary fats; and (3) whether these effects could be demonstrated in vitro, the following experiments were performed. Virgin Balb/c mice had the left inguinal mammary fat pad cleared of epithelium and were then maintained on one of four fully defined diets. These diets contained the following proportions of fat by weight: 5% or 10% mixed fats; 20% saturated fat plus cholesterol; or 20% polyunsaturated fat. To test for effects in vivo, animals received subcutaneous injections into the cleared fat pad of tumorigenic mammary cells (WAZ-2T(+SA) or WAZ-2T(-SA)) or preneoplastic mammary cells (CL-S1). Dietary fat had little effect on the latent period of tumour formation, but a low-fat diet increased the invasive/metastatic potential of both tumorigenic cell lines. A high-saturated-fat diet inhibited the growth of normal and preneoplastic epithelium in vivo. To test for effects in vitro, CL-S1 cells were co-cultured with explants of cleared mammary fat pad embedded within collagen gels. CL-S1 cells co-cultured with adipose explants obtained from mice fed on a diet containing 20% polyunsaturated fat showed a threefold increase in incorporation of [3H]thymidine into trichloroacetic acid-precipitable material. These results imply that dietary fats may affect the growth of mammary epithelium in two ways: the inhibition of growth caused by the high-saturated-fat diet may be due to systemic effects as it was not apparent in vitro; the increase in growth seen in vitro and caused by a high-polyunsaturated-fat diet is due to a direct interaction between the mammary fat pad and mammary epithelial cells. This interaction may be masked by systemic effects in vivo.

scholarly journals Conditional Deletion of β-Catenin in Mammary Epithelial Cells of Ron Receptor, Mst1r, Overexpressing Mice Alters Mammary Tumorigenesis

Endocrinology ◽  
2012 ◽  
Vol 153 (6) ◽  
pp. 2735-2746 ◽  
Author(s):  
Purnima K. Wagh ◽  
Glendon M. Zinser ◽  
Jerilyn K. Gray ◽  
Archana Shrestha ◽  
Susan E. Waltz
Keyword(s):  
Epithelial Cells ◽  
Transgenic Mice ◽  
Target Genes ◽  
Mammary Epithelial Cells ◽  
Mammary Tumorigenesis ◽  
Mammary Epithelium ◽  
Mammary Tumors ◽  
Mammary Epithelial ◽  
Conditional Deletion ◽  
Ron Receptor

The Ron receptor tyrosine kinase (macrophage stimulating 1 receptor) is overexpressed in approximately 50% of human breast cancers. Transgenic mice overexpressing Ron in the mammary epithelium [mouse mammary tumor virus driven (MMTV)-Ron expressing mice] develop mammary tumors that exhibit up-regulation of β-catenin and β-catenin target genes. β-Catenin has been shown to be a mediator of mammary tumorigenesis in various breast cancer models, including downstream of Ron. However, the in vivo impact of a conditional loss of β-catenin downstream of Ron receptor overexpression on the onset, growth, turnover, and metastasis of mammary tumors has not been addressed. To determine the significance of β-catenin in the context of Ron overexpression, we conditionally deleted β-catenin in mammary epithelial cells of MMTV-Ron mice. Conditional deletion of β-catenin in the mammary epithelium, through the use of whey acidic protein (WAP)-Cre transgenic mice, significantly delayed the onset of mammary hyperplastic nodules, the presence of palpable mammary tumors, and ultimately decreased liver metastasis. β-Catenin loss in this model was also associated with decreased expression of cyclin D1. In total, these studies support an important role for β-catenin downstream of Ron receptor signaling during the development of mammary tumorigenesis.

Abstract 3035: Cripto/GRP78 signaling promotes the stem cell phenotype in normal and neoplastic mammary epithelial cells

2014 ◽  
Author(s):  
Benjamin T. Spike ◽  
Jonathan A. Kelber ◽  
Evan Booker ◽  
Madhuri Kalathur ◽  
Rose Rodewald ◽  
...  
Keyword(s):  
Stem Cell ◽  
Epithelial Cells ◽  
Mammary Epithelial Cells ◽  
Mammary Epithelial ◽  
Cell Phenotype ◽  
Stem Cell Phenotype

scholarly journals MMTV Env encodes an ITAM responsible for transformation of mammary epithelial cells in three-dimensional culture

2005 ◽  
Vol 201 (3) ◽  
pp. 431-439 ◽  
Author(s):  
Elad Katz ◽  
Mohamed H. Lareef ◽  
John C. Rassa ◽  
Shannon M. Grande ◽  
Leslie B. King ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Tyrosine Kinases ◽  
Mammary Epithelial Cells ◽  
Cell Transformation ◽  
Three Dimensional ◽  
Soft Agar ◽  
Mammary Epithelial ◽  
Matrigel Invasion ◽  
Tumor Virus

Expression of immunoreceptor tyrosine-based activation motif (ITAM)-containing signaling proteins is normally restricted to hematopoietic tissues. The basal activity of ITAM-containing proteins is mediated through negative regulation by coreceptors restricted to hematopoietic tissues. We have identified an ITAM signaling domain encoded within the env gene of murine mammary tumor virus (MMTV). Three-dimensional structures derived in vitro from murine cells stably transfected with MMTV env display a depolarized morphology in comparison with control mammary epithelial cells. This effect is abolished by Y>F substitution within the Env ITAM, as well as inhibitors of Syk and Src protein tyrosine kinases. Env-expressing cells bear hallmarks of cell transformation such as sensitivity to apoptosis induced by tumor necrosis factor (TNF)–related apoptosis-inducing ligand (TRAIL) or TNFα, as well as down-regulation of E-cadherin and Keratin-18. Human normal mammary epithelial cells expressing MMTV Env also develop transformed phenotype, as typified by growth in soft agar and Matrigel invasion. These disruptions are abrogated by Y>F substitutions. We conclude that ITAM-dependent signals are generated through MMTV Env and trigger early hallmarks of transformation of mouse and human mammary epithelial cells. Therefore, these data suggest a heretofore unappreciated potential mechanism for the initiation of breast cancer and identify MMTV Env and ITAM-containing proteins in human breast tumors as probable oncoproteins.

scholarly journals A Role for Site-Specific Phosphorylation of Mouse Progesterone Receptor at Serine 191 in Vivo

2014 ◽  
Vol 28 (12) ◽  
pp. 2025-2037 ◽  
Author(s):  
Sandra L. Grimm ◽  
Robert D. Ward ◽  
Alison E. Obr ◽  
Heather L. Franco ◽  
Rodrigo Fernandez-Valdivia ◽  
...  
Keyword(s):  
Mammary Epithelial Cells ◽  
Phosphorylation Site ◽  
Three Dimensional ◽  
Progesterone Receptors ◽  
Nuclear Factor Κb ◽  
Mammary Epithelial ◽  
Nuclear Factor ◽  
Wild Type ◽  
Receptor Activator

Progesterone receptors (PRs) are phosphorylated on multiple sites, and a variety of roles for phosphorylation have been suggested by cell-based studies. Previous studies using PR-null mice have shown that PR plays an important role in female fertility, regulation of uterine growth, the uterine decidualization response, and proliferation as well as ductal side-branching and alveologenesis in the mammary gland. To study the role of PR phosphorylation in vivo, a mouse was engineered with homozygous replacement of PR with a PR serine-to-alanine mutation at amino acid 191. No overt phenotypes were observed in the mammary glands or uteri of PR S191A treated with progesterone (P4). In contrast, although PR S191A mice were fertile, litters were 19% smaller than wild type and the estrous cycle was lengthened slightly. Moreover, P4-dependent gene regulation in primary mammary epithelial cells (MECs) was altered in a gene-selective manner. MECs derived from wild type and PR S191A mice were grown in a three-dimensional culture. Both formed acinar structures that were morphologically similar, and proliferation was stimulated equally by P4. However, P4 induction of receptor activator of nuclear factor-κB ligand and calcitonin was selectively reduced in S191A cultures. These differences were confirmed in freshly isolated MECs. Chromatin immunoprecipitation analysis showed that the binding of S191A PR to some of the receptor activator of nuclear factor-κB ligand enhancers and a calcitonin enhancer was substantially reduced. Thus, the elimination of a single phosphorylation site is sufficient to modulate PR activity in vivo. PR contains many phosphorylation sites, and the coordinate regulation of multiple sites is a potential mechanism for selective modulation of PR function.

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