Mary Cutler - Academia.edu (original) (raw)

Papers by Mary Cutler

Research paper thumbnail of Call for Nominations: APF Gold Medal Award For Life Achievement in the Practice of Psychology

Research paper thumbnail of Rsu1 contributes to cell adhesion and spreading in MCF10A cells via effects on P38 map kinase signaling

Http Dx Doi Org 10 4161 19336918 2014 972775, Oct 23, 2014

The ILK, PINCH, Parvin (IPP) complex regulates adhesion and migration via binding of ILK to β1 in... more The ILK, PINCH, Parvin (IPP) complex regulates adhesion and migration via binding of ILK to β1 integrin and α−parvin thus linking focal adhesions to actin cytoskeleton. ILK also binds the adaptor protein PINCH which connects signaling proteins including Rsu1 to the complex. A recent study of Rsu1 and PINCH1 in non-transformed MCF10A human mammary epithelial cells revealed that the siRNA-mediated depletion of either Rsu1 or PINCH1 decreased the number of focal adhesions (FAs) and altered the distribution and localization of FA proteins. This correlated with reduced adhesion, failure to spread or migrate in response to EGF and a loss of actin stress fibers and caveolae. The depletion of Rsu1 caused significant reduction in PINCH1 implying that Rsu1 may function in part by regulating levels of PINCH1. However, Rsu1, but not PINCH1, was required for EGF-induced activation of p38 Map kinase and ATF2 phosphorylation, suggesting a Rsu1 function independent from the IPP complex. Reconstitution of Rsu1-depleted cells with a Rsu1 mutant (N92D) that does not bind to PINCH1 failed to restore FAs or migration but did promote IPP-independent spreading and constitutive as well as EGF-induced p38 activation. In this commentary we discuss p38 activity in adhesion and how Rsu1 expression may be linked to Map kinase kinase (MKK) activation and detachment-induced stress kinase signaling.

Research paper thumbnail of Long terminal repeat enhancement of v- mos transforming activity: Identification of essential regions

Journal of Virology

The transforming efficiency of recombinant DNA clones containing the Moloney sarcoma virus v-mos ... more The transforming efficiency of recombinant DNA clones containing the Moloney sarcoma virus v-mos sequence was enhanced by introducing the Moloney sarcoma virus long terminal repeat (LTR) in either the 5' or 3' position relative to v-mos. We analyzed the polyadenylated RNA expressed in cells transformed by these recombinant DNA clones and examined the structural integrity of integrated copies of the DNA. In each case, we demonstrated the presence of v-mos containing RNA transcripts in the polyadenylated RNA and showed that these RNA transcripts are consistent with the structure of the transfected DNA. The analysis of DNA from these transformed cells showed that the relative positions of the v-mos and LTR sequences within the transfected DNA were conserved in the integrated DNA copies. These results demonstrate that a single LTR can successfully enhance the transforming activity of v-mos from either a 5' or a 3' relative position. The results from the transfection analysis of recombinant clones containing only portions of the LTR introduced 3' to v-mos demonstrate that the essential region of the LTR responsible for the enhancement of transformation is a region within the unique 3' sequences of the LTR containing the 73-base-pair tandem repeat sequence.

Research paper thumbnail of Properties of the mouse mos proto-oncogene locus

Research paper thumbnail of Food Service Management Companies and the Nutritional Quality of Lunches Offered and Consumed. Washington, DC: Mathematica Policy Research

Research paper thumbnail of Ciardiello F, McGeady ML, Kim N, Basolo F, Hynes N, Langton BC et al.. Transforming growth factor-alpha expression is enhanced in human mammary epithelial cells transformed by an activated c-Ha-ras protooncogene but not by the c-neu protooncogene, and overexpression of the transforming growth fac...

Cell growth & differentiation: the molecular biology journal of the American Association for Cancer Research

ABSTRACT

Research paper thumbnail of Isolation of a new class of ‘flat’ revertants from ras-transformed NIH3T3 cells using cis-4-hydroxy-L-proline

Oncogene

A new class of nontransformed revertant cells has been isolated from the ras-transformed cell lin... more A new class of nontransformed revertant cells has been isolated from the ras-transformed cell line DT using cis-4-hydroxy-L-proline (CHP) as a selective agent. The new revertants, CHP 9CJ and CHP CB4, each contain two copies of the v-Ki-ras gene, elevated levels of phosphorylated p21ras protein, and rescuable transforming virus, indicating that the revertant phenotype observed in these cells does not result from inactivation of v-Ki-ras or inhibition of its expression. Both CHP 9CJ and CHP CB4 revertants show a greatly reduced ability to form colonies in soft agar and to produce tumors in syngeneic mice. CHP 9CJ cells are resistant to retransformation by ras and by additional oncogenes that do not encode tyrosine kinases. A comparison of oncogene resistance patterns in these CHP-derived revertants with those from our original ouabain-derived revertants fos C-11 and F-2 indicates that oncogenes may be divided into four general groups. Oncogenes that encode proteins structurally related to p21ras comprise the first group. The second group contains only tyrosine kinase-encoding oncogenes. The third group is composed of 'nuclear', e.g. fos, and 'cytoplasmic' serine-threonine-encoding oncogenes such as mos and raf. The fourth group contains the oncogenes sis and fms.

Research paper thumbnail of Development of a retroviral vector for inducible expression of transforming growth factor β1

Journal of Virology

A retroviral vector system for the expression of exogenous genes under the control of an inducibl... more A retroviral vector system for the expression of exogenous genes under the control of an inducible promoter was developed. By utilizing this system, the cDNA for human transforming growth factor 01 (TGF-I1) was inserted into a retroviral vector under the control of an internal mouse metallothionein promoter and introduced via infection into normal rat kidney fibroblasts (NRK-49F) and epithelial cells (NRK-52E), Chinese hamster ovary cells (CHO), and the human monocytic cell line U937. Control of TGF-131 expression, achieved by Cd2+ induction of vector-encoded TGF-,l1 mRNA, was cel line specific and resulted in a concomitant increase in neutralizable TGF-pl production by the cells. Autocrine stimulation of vector-containing ceUls by vector-encoded TGF-01 was detected by an increase in soft-agar colony formation of NRK-49F infectants compared with that of the control ceUls. In addition, the use of a second internal promoter in a retroviral vector of similar design allowed isolation of stable infectants from a cel line (CHO) in which the viral long terminal repeat does not function efficiently.

Research paper thumbnail of The Ras suppressor RSU-1 localizes to 10p13 and its expression in the U251 glioblastoma cell line correlates with a decrease in growth rate and tumorigenic potential

Research paper thumbnail of Infection with a TGF-α retroviral vector transforms mouse mammary epithelial cells but not normal fibroblasts

Research paper thumbnail of Control of Mammary Differentiation by Ras-Dependent Signal Transduction Pathways

Public reporting burden for this collection of information is estimated to average 1 hour per res... more Public reporting burden for this collection of information is estimated to average 1 hour per response, including the time for reviewing instructions, searching existing data sources, gathering and maintaining the data needed, and completing and reviewing this collection of Information. Send comments regarding this burden estimate or any other aspect of this collection of information, including suggestions for reducing this burden to Washington Headquarters Services, Directorate for Information Operations and Reports, 1215 Jefferson Davis Highway, Suite 1204, Arlington, VA 22202-4302, and to the Office of Management and Budget, Paperwork Reduction Project (0704-0188), Washington, DC 20503

Research paper thumbnail of Morrison, B. and Mary Lou Cutler. “The Contribution of Adhesion Signaling to Lactogenesis.”

Research paper thumbnail of InVitro Methylation ofSpecific Regions oftheCloned MoloneySarcoma VirusGenomeInhibits ItsTransforming Activity

Thetransforming activity ofcloned Moloney sarcoma virus (MSV)proviral DNA was inhibited byinvitro... more Thetransforming activity ofcloned Moloney sarcoma virus (MSV)proviral DNA was inhibited byinvitro methylation oftheDNA atcytosine residues, using HpaII andHhaImethylases before transfection into NIH3T3cells. Theinhibition oftransforming activity duetoHpaIImethylation was reversed bytreatment of thetransfected cells with5-azacytidine, a specific inhibitor ofmethylation. Analysis ofthegenomic DNA fromthetransformed cells whichresulted fromthe transfection ofmethylated MSV DNA revealed thattheintegrated MSV proviral DNA was sensitive toHpaIIdigestion inallcell lines examined, suggesting that loss ofmethyl groups was necessaryfortransformation. Whencells were infected withMoloneymurineleukemia virus atvarious timesafter transfection with methylated MSV DNA,theamountoftransforming virus produced indicated that theloss ofmethyl groupsoccurred within 24h.Methylation ofMSV DNA atHhaI sites was asinhibitory totransforming activity asmethylation atHpaIIsites. In addition, methylation atbothHp...

Research paper thumbnail of Abstract 2616: Rsu1 regulates MCF10A mammary epithelial cell migration and acini formation by binding to the IPP complex but it promotes spreading and Map kinase activation independent of IPP

Research paper thumbnail of Increased expression of the Ras suppressor Rsu-1 enhances Erk-2 activation and inhibits Jun kinase activation

Molecular and cellular biology, 1996

Studies were undertaken to determine the effect of the Ras suppressor Rsu-1 on Ras signal transdu... more Studies were undertaken to determine the effect of the Ras suppressor Rsu-1 on Ras signal transduction pathways in two different cell backgrounds. An expression vector containing the mouse rsu-1 cDNA under the control of a mouse mammary tumor virus promoter was introduced into NIH 3T3 cells and the pheochromocytoma cell line PC12. Cell lines developed in the NIH 3T3 background expressed p33rsu-1 at approximately twice the normal endogenous level. However, PC12 cell clones which expressed p33rsu-1 at an increased level in a regulatable fashion in response to dexamethasone were isolated. Analysis of proteins involved in regulation of Ras and responsive to Ras signal transduction revealed similar changes in the two cell backgrounds in the presence of elevated p33rsu-1. There was an increase in the level of SOS, the guanine nucleotide exchange factor, and an increase in the percentage of GTP-bound Ras. In addition, there was an increase in the amount of p120 Ras-specific GTPase-activati...

Research paper thumbnail of In vitro methylation of specific regions of the cloned Moloney sarcoma virus genome inhibits its transforming activity

Molecular and cellular biology, 1983

The transforming activity of cloned Moloney sarcoma virus (MSV) proviral DNA was inhibited by in ... more The transforming activity of cloned Moloney sarcoma virus (MSV) proviral DNA was inhibited by in vitro methylation of the DNA at cytosine residues, using HpaII and HhaI methylases before transfection into NIH 3T3 cells. The inhibition of transforming activity due to HpaII methylation was reversed by treatment of the transfected cells with 5-azacytidine, a specific inhibitor of methylation. Analysis of the genomic DNA from the transformed cells which resulted from the transfection of methylated MSV DNA revealed that the integrated MSV proviral DNA was sensitive to HpaII digestion in all cell lines examined, suggesting that loss of methyl groups was necessary for transformation. When cells were infected with Moloney murine leukemia virus at various times after transfection with methylated MSV DNA, the amount of transforming virus produced indicated that the loss of methyl groups occurred within 24 h. Methylation of MSV DNA at HhaI sites was as inhibitory to transforming activity as me...

Research paper thumbnail of Isolation of rsp-1, a novel cDNA capable of suppressing v-Ras transformation

Molecular and cellular biology, 1992

Using an expression cloning assay, we have isolated a novel cDNA, referred to as rsp-1, which sup... more Using an expression cloning assay, we have isolated a novel cDNA, referred to as rsp-1, which suppresses the v-Ras-transformed phenotype. When introduced into NIH 3T3 fibroblasts under the control of a metallothionein promoter, rsp-1 confers resistance to v-Ras, but not to v-Mos or v-Src, and inhibits growth of the cells. The rsp-1 cDNA contains a 831-bp open reading frame encoding a 277-amino-acid leucine-rich protein. The rsp-1 cDNA exhibits no significant homology to sequences in the DNA data bases. However, searches of the protein data bases revealed that it contains a series of leucine-based repeats which are homologous to the leucine repeats found in the regulatory region of the yeast adenylyl cyclase. rsp-1 specific RNA is detectable in a wide variety of cell lines and tissues, and the gene is conserved among eukaryotic species. These data suggest that rsp-1 plays a role in Ras signal transduction.

Research paper thumbnail of Global Expression Profiling Reveals a Role for CTGF/CCN2 in Lactogenic Differentiation of Mouse Mammary Epithelial Cells

CCN Proteins in Health and Disease, 2010

Mammary epithelial cells go through a series of developmental changes during pregnancy and lactat... more Mammary epithelial cells go through a series of developmental changes during pregnancy and lactation including proliferation, differentiation, secretion and apoptosis. HC11 mouse mammary epithelial cells, which undergo lactogen-induced differentiation in cell culture, were used to follow the changes in gene expression during this process. The expression profiles of over 20,000 genes were compared in HC11 cells undergoing lactogenic differentiation to non-differentiated cells using DNA microarray analysis. Greater than two fold changes were detected in 998 genes in the differentiated cells versus growth controls. Several genes including CTGF/CCN2 exhibited greater than five-fold increase. Validation of the gene expression pattern for more than twenty genes was performed. The results indicate the involvement of numerous genes and pathways in the differentiation of mouse mammary epithelial cells in culture and they identify genetic pathways associated with specific transcriptional regulation. In addition, the expression of a subset of genes regulated by lactogenic differentiation in HC11 cells, including CTGF/CCN2 and osteopontin, was examined in mouse mammary glands revealing expression during pregnancy and lactation that declined during involution of the glands. To probe the mechanism by which epidermal growth factor (EGF), a known inhibitor of lactogenic differentiation in HC11 cells, blocks lactogenesis, the HC11 cells stimulated with lactogenic hormone in the presence of EGF were profiled. This data revealed EGF regulation of a specific subset of genes including important cell cycle regulators. The studies confirm the value of expression profiling in defining gene transcription associated with differentiation of mammary epithelial cells.

Research paper thumbnail of Rsu1 contributes to regulation of cell adhesion and spreading by PINCH1-dependent and - independent mechanisms

Journal of Cell Communication and Signaling, 2013

Cell adhesion and migration are complex processes that require integrin activation, the formation... more Cell adhesion and migration are complex processes that require integrin activation, the formation and dissolution of focal adhesion (FAs), and linkage of actin cytoskeleton to the FAs. The IPP (ILK, PINCH, Parvin) complex regulates FA formation via binding of the adaptor protein ILK to β1 integrin, PINCH and parvin. The signaling protein Rsu1 is linked to the complex via binding PINCH1. The role of Rsu1 and PINCH1 in adhesion and migration was examined in non-transformed mammary epithelial cells. Confocal microscopy revealed that the depletion of either Rsu1 or PINCH1 by siRNA in MCF10A cells decreased the number of focal adhesions and altered the distribution and localization of β1 integrin, vinculin, talin and paxillin without affecting the levels of FA protein expression. This correlated with reduced adhesion, failure to spread or migrate in response to EGF and a loss of actin stress fibers and caveolae. In addition, constitutive phosphorylation of actin regulatory proteins occurred in the absence of PINCH1. The depletion of Rsu1 caused significant reduction in PINCH1 implying that Rsu1 may function by regulating levels of PINCH1. However, while both Rsu1-or PINCH1-depleted cells retained the ability to activate adhesion signaling in response to EGF stimulation, only Rsu1 was required for EGFinduced p38 Map Kinase phosphorylation and ATF2 activation, suggesting an Rsu1 function independent from the IPP complex. Reconstitution of Rsu1-depleted cells with an Rsu1 mutant that does not bind to PINCH1 failed to restore FAs or migration but did promote spreading and constitutive p38 activation. These data show that Rsu1-PINCH1 association with ILK and the IPP complex is required for regulation of adhesion and migration but that Rsu1 has a critical role in linking integrin-induced adhesion to activation of p38 Map kinase signaling and cell spreading. Moreover, it suggests that Rsu1 may regulate p38 signaling from the IPP complex affecting other functions including survival.

Research paper thumbnail of Role for chondroitin sulfate glycosaminoglycan in NEDD9-mediated breast cancer cell growth

Experimental Cell Research, 2015

There are lines of evidence demonstrating that NEDD9 (Cas-L, HEF-1) plays a key role in the devel... more There are lines of evidence demonstrating that NEDD9 (Cas-L, HEF-1) plays a key role in the development, progression, and metastasis of breast cancer cells. We previously reported that NEDD9 plays a critical role for promoting migration and growth of MDA-MB-231. In order to further characterize the mechanisms of NEDD9-mediated cancer migration and growth, stable cells overexpressing NEDD9 were generated using HCC38 as a parental cell line which expresses low level of endogenous NEDD9. Microarray studies demonstrated that core proteins of CD44 and Serglycin were markedly upregulated in HCC38(NEDD9) cells compared to HCC38(Vector) cells, while those of Syndecan-1, Syndecan-2, and Versican were downregulated in HCC38(NEDD9). Importantly, enzymes generating chondroitin sulfate glycosaminoglycans (CS) such as CHST11, CHST15, and CSGALNACT1 were upregulated in HCC38(NEDD9) compared to HCC38(Vector). Immunofluorescence studies using specific antibody, GD3G7, confirmed the enhanced expression of CS-E subunit in HCC38(NEDD9). Immunoprecipitation and western blotting analysis demonstrated that CS-E was attached to CD44 core protein. We demonstrated that removing CS by chondroitinase ABC significantly inhibited anchorage-independent colony formation of HCC38(NEDD9) in methylcellulose. Importantly, the fact that GD3G7 significantly inhibited colony formation of HCC38(NEDD9) cells suggests that CS-E subunit plays a key role in this process. Furthermore, treatment of HCC38(NEDD9) cells with chondroitinase ABC or GD3G7 significantly inhibited mammosphere formation. Exogenous addition of CS-E enhanced colony formation and mammosphere formation of HCC38 parental and HCC38(Vector) cells. These results suggest that NEDD9 regulates the synthesis and expression of tumor associated glycocalyx structures including CS-E, which plays a key role in promoting and regulating breast cancer progression and metastasis and possibly stem cell phenotypes.

Research paper thumbnail of Call for Nominations: APF Gold Medal Award For Life Achievement in the Practice of Psychology

Research paper thumbnail of Rsu1 contributes to cell adhesion and spreading in MCF10A cells via effects on P38 map kinase signaling

Http Dx Doi Org 10 4161 19336918 2014 972775, Oct 23, 2014

The ILK, PINCH, Parvin (IPP) complex regulates adhesion and migration via binding of ILK to β1 in... more The ILK, PINCH, Parvin (IPP) complex regulates adhesion and migration via binding of ILK to β1 integrin and α−parvin thus linking focal adhesions to actin cytoskeleton. ILK also binds the adaptor protein PINCH which connects signaling proteins including Rsu1 to the complex. A recent study of Rsu1 and PINCH1 in non-transformed MCF10A human mammary epithelial cells revealed that the siRNA-mediated depletion of either Rsu1 or PINCH1 decreased the number of focal adhesions (FAs) and altered the distribution and localization of FA proteins. This correlated with reduced adhesion, failure to spread or migrate in response to EGF and a loss of actin stress fibers and caveolae. The depletion of Rsu1 caused significant reduction in PINCH1 implying that Rsu1 may function in part by regulating levels of PINCH1. However, Rsu1, but not PINCH1, was required for EGF-induced activation of p38 Map kinase and ATF2 phosphorylation, suggesting a Rsu1 function independent from the IPP complex. Reconstitution of Rsu1-depleted cells with a Rsu1 mutant (N92D) that does not bind to PINCH1 failed to restore FAs or migration but did promote IPP-independent spreading and constitutive as well as EGF-induced p38 activation. In this commentary we discuss p38 activity in adhesion and how Rsu1 expression may be linked to Map kinase kinase (MKK) activation and detachment-induced stress kinase signaling.

Research paper thumbnail of Long terminal repeat enhancement of v- mos transforming activity: Identification of essential regions

Journal of Virology

The transforming efficiency of recombinant DNA clones containing the Moloney sarcoma virus v-mos ... more The transforming efficiency of recombinant DNA clones containing the Moloney sarcoma virus v-mos sequence was enhanced by introducing the Moloney sarcoma virus long terminal repeat (LTR) in either the 5' or 3' position relative to v-mos. We analyzed the polyadenylated RNA expressed in cells transformed by these recombinant DNA clones and examined the structural integrity of integrated copies of the DNA. In each case, we demonstrated the presence of v-mos containing RNA transcripts in the polyadenylated RNA and showed that these RNA transcripts are consistent with the structure of the transfected DNA. The analysis of DNA from these transformed cells showed that the relative positions of the v-mos and LTR sequences within the transfected DNA were conserved in the integrated DNA copies. These results demonstrate that a single LTR can successfully enhance the transforming activity of v-mos from either a 5' or a 3' relative position. The results from the transfection analysis of recombinant clones containing only portions of the LTR introduced 3' to v-mos demonstrate that the essential region of the LTR responsible for the enhancement of transformation is a region within the unique 3' sequences of the LTR containing the 73-base-pair tandem repeat sequence.

Research paper thumbnail of Properties of the mouse mos proto-oncogene locus

Research paper thumbnail of Food Service Management Companies and the Nutritional Quality of Lunches Offered and Consumed. Washington, DC: Mathematica Policy Research

Research paper thumbnail of Ciardiello F, McGeady ML, Kim N, Basolo F, Hynes N, Langton BC et al.. Transforming growth factor-alpha expression is enhanced in human mammary epithelial cells transformed by an activated c-Ha-ras protooncogene but not by the c-neu protooncogene, and overexpression of the transforming growth fac...

Cell growth & differentiation: the molecular biology journal of the American Association for Cancer Research

ABSTRACT

Research paper thumbnail of Isolation of a new class of ‘flat’ revertants from ras-transformed NIH3T3 cells using cis-4-hydroxy-L-proline

Oncogene

A new class of nontransformed revertant cells has been isolated from the ras-transformed cell lin... more A new class of nontransformed revertant cells has been isolated from the ras-transformed cell line DT using cis-4-hydroxy-L-proline (CHP) as a selective agent. The new revertants, CHP 9CJ and CHP CB4, each contain two copies of the v-Ki-ras gene, elevated levels of phosphorylated p21ras protein, and rescuable transforming virus, indicating that the revertant phenotype observed in these cells does not result from inactivation of v-Ki-ras or inhibition of its expression. Both CHP 9CJ and CHP CB4 revertants show a greatly reduced ability to form colonies in soft agar and to produce tumors in syngeneic mice. CHP 9CJ cells are resistant to retransformation by ras and by additional oncogenes that do not encode tyrosine kinases. A comparison of oncogene resistance patterns in these CHP-derived revertants with those from our original ouabain-derived revertants fos C-11 and F-2 indicates that oncogenes may be divided into four general groups. Oncogenes that encode proteins structurally related to p21ras comprise the first group. The second group contains only tyrosine kinase-encoding oncogenes. The third group is composed of 'nuclear', e.g. fos, and 'cytoplasmic' serine-threonine-encoding oncogenes such as mos and raf. The fourth group contains the oncogenes sis and fms.

Research paper thumbnail of Development of a retroviral vector for inducible expression of transforming growth factor β1

Journal of Virology

A retroviral vector system for the expression of exogenous genes under the control of an inducibl... more A retroviral vector system for the expression of exogenous genes under the control of an inducible promoter was developed. By utilizing this system, the cDNA for human transforming growth factor 01 (TGF-I1) was inserted into a retroviral vector under the control of an internal mouse metallothionein promoter and introduced via infection into normal rat kidney fibroblasts (NRK-49F) and epithelial cells (NRK-52E), Chinese hamster ovary cells (CHO), and the human monocytic cell line U937. Control of TGF-131 expression, achieved by Cd2+ induction of vector-encoded TGF-,l1 mRNA, was cel line specific and resulted in a concomitant increase in neutralizable TGF-pl production by the cells. Autocrine stimulation of vector-containing ceUls by vector-encoded TGF-01 was detected by an increase in soft-agar colony formation of NRK-49F infectants compared with that of the control ceUls. In addition, the use of a second internal promoter in a retroviral vector of similar design allowed isolation of stable infectants from a cel line (CHO) in which the viral long terminal repeat does not function efficiently.

Research paper thumbnail of The Ras suppressor RSU-1 localizes to 10p13 and its expression in the U251 glioblastoma cell line correlates with a decrease in growth rate and tumorigenic potential

Research paper thumbnail of Infection with a TGF-α retroviral vector transforms mouse mammary epithelial cells but not normal fibroblasts

Research paper thumbnail of Control of Mammary Differentiation by Ras-Dependent Signal Transduction Pathways

Public reporting burden for this collection of information is estimated to average 1 hour per res... more Public reporting burden for this collection of information is estimated to average 1 hour per response, including the time for reviewing instructions, searching existing data sources, gathering and maintaining the data needed, and completing and reviewing this collection of Information. Send comments regarding this burden estimate or any other aspect of this collection of information, including suggestions for reducing this burden to Washington Headquarters Services, Directorate for Information Operations and Reports, 1215 Jefferson Davis Highway, Suite 1204, Arlington, VA 22202-4302, and to the Office of Management and Budget, Paperwork Reduction Project (0704-0188), Washington, DC 20503

Research paper thumbnail of Morrison, B. and Mary Lou Cutler. “The Contribution of Adhesion Signaling to Lactogenesis.”

Research paper thumbnail of InVitro Methylation ofSpecific Regions oftheCloned MoloneySarcoma VirusGenomeInhibits ItsTransforming Activity

Thetransforming activity ofcloned Moloney sarcoma virus (MSV)proviral DNA was inhibited byinvitro... more Thetransforming activity ofcloned Moloney sarcoma virus (MSV)proviral DNA was inhibited byinvitro methylation oftheDNA atcytosine residues, using HpaII andHhaImethylases before transfection into NIH3T3cells. Theinhibition oftransforming activity duetoHpaIImethylation was reversed bytreatment of thetransfected cells with5-azacytidine, a specific inhibitor ofmethylation. Analysis ofthegenomic DNA fromthetransformed cells whichresulted fromthe transfection ofmethylated MSV DNA revealed thattheintegrated MSV proviral DNA was sensitive toHpaIIdigestion inallcell lines examined, suggesting that loss ofmethyl groups was necessaryfortransformation. Whencells were infected withMoloneymurineleukemia virus atvarious timesafter transfection with methylated MSV DNA,theamountoftransforming virus produced indicated that theloss ofmethyl groupsoccurred within 24h.Methylation ofMSV DNA atHhaI sites was asinhibitory totransforming activity asmethylation atHpaIIsites. In addition, methylation atbothHp...

Research paper thumbnail of Abstract 2616: Rsu1 regulates MCF10A mammary epithelial cell migration and acini formation by binding to the IPP complex but it promotes spreading and Map kinase activation independent of IPP

Research paper thumbnail of Increased expression of the Ras suppressor Rsu-1 enhances Erk-2 activation and inhibits Jun kinase activation

Molecular and cellular biology, 1996

Studies were undertaken to determine the effect of the Ras suppressor Rsu-1 on Ras signal transdu... more Studies were undertaken to determine the effect of the Ras suppressor Rsu-1 on Ras signal transduction pathways in two different cell backgrounds. An expression vector containing the mouse rsu-1 cDNA under the control of a mouse mammary tumor virus promoter was introduced into NIH 3T3 cells and the pheochromocytoma cell line PC12. Cell lines developed in the NIH 3T3 background expressed p33rsu-1 at approximately twice the normal endogenous level. However, PC12 cell clones which expressed p33rsu-1 at an increased level in a regulatable fashion in response to dexamethasone were isolated. Analysis of proteins involved in regulation of Ras and responsive to Ras signal transduction revealed similar changes in the two cell backgrounds in the presence of elevated p33rsu-1. There was an increase in the level of SOS, the guanine nucleotide exchange factor, and an increase in the percentage of GTP-bound Ras. In addition, there was an increase in the amount of p120 Ras-specific GTPase-activati...

Research paper thumbnail of In vitro methylation of specific regions of the cloned Moloney sarcoma virus genome inhibits its transforming activity

Molecular and cellular biology, 1983

The transforming activity of cloned Moloney sarcoma virus (MSV) proviral DNA was inhibited by in ... more The transforming activity of cloned Moloney sarcoma virus (MSV) proviral DNA was inhibited by in vitro methylation of the DNA at cytosine residues, using HpaII and HhaI methylases before transfection into NIH 3T3 cells. The inhibition of transforming activity due to HpaII methylation was reversed by treatment of the transfected cells with 5-azacytidine, a specific inhibitor of methylation. Analysis of the genomic DNA from the transformed cells which resulted from the transfection of methylated MSV DNA revealed that the integrated MSV proviral DNA was sensitive to HpaII digestion in all cell lines examined, suggesting that loss of methyl groups was necessary for transformation. When cells were infected with Moloney murine leukemia virus at various times after transfection with methylated MSV DNA, the amount of transforming virus produced indicated that the loss of methyl groups occurred within 24 h. Methylation of MSV DNA at HhaI sites was as inhibitory to transforming activity as me...

Research paper thumbnail of Isolation of rsp-1, a novel cDNA capable of suppressing v-Ras transformation

Molecular and cellular biology, 1992

Using an expression cloning assay, we have isolated a novel cDNA, referred to as rsp-1, which sup... more Using an expression cloning assay, we have isolated a novel cDNA, referred to as rsp-1, which suppresses the v-Ras-transformed phenotype. When introduced into NIH 3T3 fibroblasts under the control of a metallothionein promoter, rsp-1 confers resistance to v-Ras, but not to v-Mos or v-Src, and inhibits growth of the cells. The rsp-1 cDNA contains a 831-bp open reading frame encoding a 277-amino-acid leucine-rich protein. The rsp-1 cDNA exhibits no significant homology to sequences in the DNA data bases. However, searches of the protein data bases revealed that it contains a series of leucine-based repeats which are homologous to the leucine repeats found in the regulatory region of the yeast adenylyl cyclase. rsp-1 specific RNA is detectable in a wide variety of cell lines and tissues, and the gene is conserved among eukaryotic species. These data suggest that rsp-1 plays a role in Ras signal transduction.

Research paper thumbnail of Global Expression Profiling Reveals a Role for CTGF/CCN2 in Lactogenic Differentiation of Mouse Mammary Epithelial Cells

CCN Proteins in Health and Disease, 2010

Mammary epithelial cells go through a series of developmental changes during pregnancy and lactat... more Mammary epithelial cells go through a series of developmental changes during pregnancy and lactation including proliferation, differentiation, secretion and apoptosis. HC11 mouse mammary epithelial cells, which undergo lactogen-induced differentiation in cell culture, were used to follow the changes in gene expression during this process. The expression profiles of over 20,000 genes were compared in HC11 cells undergoing lactogenic differentiation to non-differentiated cells using DNA microarray analysis. Greater than two fold changes were detected in 998 genes in the differentiated cells versus growth controls. Several genes including CTGF/CCN2 exhibited greater than five-fold increase. Validation of the gene expression pattern for more than twenty genes was performed. The results indicate the involvement of numerous genes and pathways in the differentiation of mouse mammary epithelial cells in culture and they identify genetic pathways associated with specific transcriptional regulation. In addition, the expression of a subset of genes regulated by lactogenic differentiation in HC11 cells, including CTGF/CCN2 and osteopontin, was examined in mouse mammary glands revealing expression during pregnancy and lactation that declined during involution of the glands. To probe the mechanism by which epidermal growth factor (EGF), a known inhibitor of lactogenic differentiation in HC11 cells, blocks lactogenesis, the HC11 cells stimulated with lactogenic hormone in the presence of EGF were profiled. This data revealed EGF regulation of a specific subset of genes including important cell cycle regulators. The studies confirm the value of expression profiling in defining gene transcription associated with differentiation of mammary epithelial cells.

Research paper thumbnail of Rsu1 contributes to regulation of cell adhesion and spreading by PINCH1-dependent and - independent mechanisms

Journal of Cell Communication and Signaling, 2013

Cell adhesion and migration are complex processes that require integrin activation, the formation... more Cell adhesion and migration are complex processes that require integrin activation, the formation and dissolution of focal adhesion (FAs), and linkage of actin cytoskeleton to the FAs. The IPP (ILK, PINCH, Parvin) complex regulates FA formation via binding of the adaptor protein ILK to β1 integrin, PINCH and parvin. The signaling protein Rsu1 is linked to the complex via binding PINCH1. The role of Rsu1 and PINCH1 in adhesion and migration was examined in non-transformed mammary epithelial cells. Confocal microscopy revealed that the depletion of either Rsu1 or PINCH1 by siRNA in MCF10A cells decreased the number of focal adhesions and altered the distribution and localization of β1 integrin, vinculin, talin and paxillin without affecting the levels of FA protein expression. This correlated with reduced adhesion, failure to spread or migrate in response to EGF and a loss of actin stress fibers and caveolae. In addition, constitutive phosphorylation of actin regulatory proteins occurred in the absence of PINCH1. The depletion of Rsu1 caused significant reduction in PINCH1 implying that Rsu1 may function by regulating levels of PINCH1. However, while both Rsu1-or PINCH1-depleted cells retained the ability to activate adhesion signaling in response to EGF stimulation, only Rsu1 was required for EGFinduced p38 Map Kinase phosphorylation and ATF2 activation, suggesting an Rsu1 function independent from the IPP complex. Reconstitution of Rsu1-depleted cells with an Rsu1 mutant that does not bind to PINCH1 failed to restore FAs or migration but did promote spreading and constitutive p38 activation. These data show that Rsu1-PINCH1 association with ILK and the IPP complex is required for regulation of adhesion and migration but that Rsu1 has a critical role in linking integrin-induced adhesion to activation of p38 Map kinase signaling and cell spreading. Moreover, it suggests that Rsu1 may regulate p38 signaling from the IPP complex affecting other functions including survival.

Research paper thumbnail of Role for chondroitin sulfate glycosaminoglycan in NEDD9-mediated breast cancer cell growth

Experimental Cell Research, 2015

There are lines of evidence demonstrating that NEDD9 (Cas-L, HEF-1) plays a key role in the devel... more There are lines of evidence demonstrating that NEDD9 (Cas-L, HEF-1) plays a key role in the development, progression, and metastasis of breast cancer cells. We previously reported that NEDD9 plays a critical role for promoting migration and growth of MDA-MB-231. In order to further characterize the mechanisms of NEDD9-mediated cancer migration and growth, stable cells overexpressing NEDD9 were generated using HCC38 as a parental cell line which expresses low level of endogenous NEDD9. Microarray studies demonstrated that core proteins of CD44 and Serglycin were markedly upregulated in HCC38(NEDD9) cells compared to HCC38(Vector) cells, while those of Syndecan-1, Syndecan-2, and Versican were downregulated in HCC38(NEDD9). Importantly, enzymes generating chondroitin sulfate glycosaminoglycans (CS) such as CHST11, CHST15, and CSGALNACT1 were upregulated in HCC38(NEDD9) compared to HCC38(Vector). Immunofluorescence studies using specific antibody, GD3G7, confirmed the enhanced expression of CS-E subunit in HCC38(NEDD9). Immunoprecipitation and western blotting analysis demonstrated that CS-E was attached to CD44 core protein. We demonstrated that removing CS by chondroitinase ABC significantly inhibited anchorage-independent colony formation of HCC38(NEDD9) in methylcellulose. Importantly, the fact that GD3G7 significantly inhibited colony formation of HCC38(NEDD9) cells suggests that CS-E subunit plays a key role in this process. Furthermore, treatment of HCC38(NEDD9) cells with chondroitinase ABC or GD3G7 significantly inhibited mammosphere formation. Exogenous addition of CS-E enhanced colony formation and mammosphere formation of HCC38 parental and HCC38(Vector) cells. These results suggest that NEDD9 regulates the synthesis and expression of tumor associated glycocalyx structures including CS-E, which plays a key role in promoting and regulating breast cancer progression and metastasis and possibly stem cell phenotypes.