scholarly journals RING1 Interacts with Multiple Polycomb-Group Proteins and Displays Tumorigenic Activity

1999 ◽  
Vol 19 (1) ◽  
pp. 57-68 ◽  
Author(s):  
David P. E. Satijn ◽  
Arie P. Otte
Keyword(s):  
Gene Expression ◽  
Protein Interactions ◽  
Mammalian Cells ◽  
Protein Complexes ◽  
Cellular Transformation ◽  
Polycomb Group ◽  
Expression Levels ◽  
Self Association

ABSTRACT Polycomb-group (PcG) proteins form large multimeric protein complexes that are involved in maintaining the transcriptionally repressive state of genes. Previously, we reported that RING1 interacts with vertebrate Polycomb (Pc) homologs and is associated with or is part of a human PcG complex. However, very little is known about the role of RING1 as a component of the PcG complex. Here we undertake a detailed characterization of RING1 protein-protein interactions. By using directed two-hybrid and in vitro protein-protein analyses, we demonstrate that RING1, besides interacting with the human Pc homolog HPC2, can also interact with itself and with the vertebrate PcG protein BMI1. Distinct domains in the RING1 protein are involved in the self-association and in the interaction with BMI1. Further, we find that the BMI1 protein can also interact with itself. To better understand the role of RING1 in regulating gene expression, we overexpressed the protein in mammalian cells and analyzed differences in gene expression levels. This analysis shows that overexpression of RING1 strongly represses En-2, a mammalian homolog of the well-characterized Drosophila PcG target geneengrailed. Furthermore, RING1 overexpression results in enhanced expression of the proto-oncogenes c-jun and c-fos. The changes in expression levels of these proto-oncogenes are accompanied by cellular transformation, as judged by anchorage-independent growth and the induction of tumors in athymic mice. Our data demonstrate that RING1 interacts with multiple human PcG proteins, indicating an important role for RING1 in the PcG complex. Further, deregulation of RING1 expression leads to oncogenic transformation by deregulation of the expression levels of certain oncogenes.

scholarly journals DNA Methylation and Chromatin: Role(s) of Methyl-CpG-Binding Protein ZBTB38

2018 ◽  
Vol 11 ◽  
pp. 251686571881111 ◽  
Author(s):  
Maud de Dieuleveult ◽  
Benoit Miotto
Keyword(s):  
Gene Expression ◽  
Dna Methylation ◽  
Transcription Factors ◽  
Dna Sequences ◽  
Mammalian Cells ◽  
Expression Patterns ◽  
Control Of Gene Expression ◽  
Methylated Dna

DNA methylation plays an essential role in the control of gene expression during early stages of development as well as in disease. Although many transcription factors are sensitive to this modification of the DNA, we still do not clearly understand how it contributes to the establishment of proper gene expression patterns. We discuss here the recent findings regarding the biological and molecular function(s) of the transcription factor ZBTB38 that binds methylated DNA sequences in vitro and in cells. We speculate how these findings may help understand the role of DNA methylation and DNA methylation–sensitive transcription factors in mammalian cells.

Expression of Midkine(MDK) and Sensitivity to Anti-Leukemic Drugs in Pediatric Acute Lymphoblastic Leukemia (ALL).

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 1432-1432
Author(s):  
Weihua Zeng ◽  
I-Ming Chen ◽  
Justin Tibbitts ◽  
Huining Kang ◽  
Kerem Ar ◽  
...  
Keyword(s):  
Gene Expression ◽  
Lymphoblastic Leukemia ◽  
Mrna Levels ◽  
Healthy Controls ◽  
Expression Levels ◽  
Whitney Test ◽  
Mann Whitney Test ◽  
Reh Cells

Abstract Midkine (MDK), a heparin-binding growth factor, is highly expressed in pediatric ALL, neuroblastomas, and various solid tumors. Previous gene expression profiling studies conducted in our laboratory on a cohort of 220 children with B precursor ALL treated on Children’s Oncology Group (COG) clinical trials have shown that higher levels of MDK expression in leukemic blasts are strongly associated with continuous complete remission (CCR) and that MDK is a powerful prognostic predictor of outcome (Wilcoxon ranksum test, p=0.000001), particularly in patients with central nervous system (CNS) involvement (Wilcoxon ranksum test, p =0.009). To explore the role of MDK in hematopoiesis and leukemogenesis and to further explore MDK as a potential novel target for ALL therapy, we examined MDK mRNA expression in three stages of normal B cell development in healthy controls and compared it to expression levels in leukemic blasts from children with B-ALL. Quantitative RT-PCR assays showed that MDK mRNA levels in CD19+ cells from peripheral blood mononuclear cells (PBMNC) were significantly higher than those in CD34+ or CD19+ cells from bone marrow (BM) (Mann Whitney test, p=0.0025 and 0.0018, respectively). ALL leukemic blasts samples expressed significantly higher levels of MDK than those in CD19+ cell in the BM of healthy controls (Mann Whitney test, p=0.0043). Similar results were obtained by ELISA detection of MDK protein in plasma of ALL patients and healthy controls. We have also investigated the relationship of MDK expression and the chemo-sensitivity of ALL cell lines. Using a lentiviral system, we have successfully over-expressed or knocked down MDK gene expression in the REH cell line containing the t(12;21). Unexpectedly, MTS assays revealed that MDK expression is inversely associated with sensitivity to L-asparaginase and doxorubicin, but not to vincristine and cytarabine, in REH cells. Higher levels of MDK were associated with chemo-resistance. Finally, we tested the effects of MDK expression on proliferation and apoptosis in REH cells and determined that although MDK expression was not associated with REH cell proliferation in vitro, it protected REH cells from doxorubicin-induced apoptotic cell death. Taken together, these results suggest that expression levels of MDK in ALL is inversely associated with in vitro drug sensitivity to specific anti-leukemic agents; consistent with observations in solid tumor models. It has been suggested that 4β1- and 6β1-integrins bind directly to MDK and are involved in MK-dependent cell migration. Further studies of the role of MDK signaling in vivo in the complex marrow/adhesion microenvironment and the effects on ALL survival and chemo-sensitivity and resistance are in progress.

scholarly journals Efficient and Specific Targeting of Polycomb Group Proteins Requires Cooperative Interaction between Grainyhead and Pleiohomeotic

2006 ◽  
Vol 26 (4) ◽  
pp. 1434-1444 ◽  
Author(s):  
András Blastyák ◽  
Rakesh K. Mishra ◽  
Francois Karch ◽  
Henrik Gyurkovics
Keyword(s):  
Dna Binding ◽  
Binding Proteins ◽  
Protein Complexes ◽  
Dna Binding Proteins ◽  
Polycomb Group ◽  
Cooperative Interaction ◽  
Protein Protein Interaction ◽  
Specific Targeting

ABSTRACT Specific targeting of the protein complexes formed by the Polycomb group of proteins is critically required to maintain the inactive state of a group of developmentally regulated genes. Although the role of DNA binding proteins in this process has been well established, it is still not understood how these proteins target the Polycomb complexes specifically to their response elements. Here we show that the grainyhead gene, which encodes a DNA binding protein, interacts with one such Polycomb response element of the bithorax complex. Grainyhead binds to this element in vitro. Moreover, grainyhead interacts genetically with pleiohomeotic in a transgene-based, pairing-dependent silencing assay. Grainyhead also interacts with Pleiohomeotic in vitro, which facilitates the binding of both proteins to their respective target DNAs. Such interactions between two DNA binding proteins could provide the basis for the cooperative assembly of a nucleoprotein complex formed in vitro. Based on these results and the available data, we propose that the role of DNA binding proteins in Polycomb group-dependent silencing could be described by a model very similar to that of an enhanceosome, wherein the unique arrangement of protein-protein interaction modules exposed by the cooperatively interacting DNA binding proteins provides targeting specificity.

Role of small nuclear RNAs in eukaryotic gene expression

2013 ◽  
Vol 54 ◽  
pp. 79-90 ◽  
Author(s):  
Saba Valadkhan ◽  
Lalith S. Gunawardane
Keyword(s):  
Gene Expression ◽  
Protein Interactions ◽  
Rna Stability ◽  
Splice Sites ◽  
Branch Site ◽  
Small Nuclear Rnas ◽  
Eukaryotic Gene Expression ◽  
Eukaryotic Gene ◽  
Rna Biogenesis

Eukaryotic cells contain small, highly abundant, nuclear-localized non-coding RNAs [snRNAs (small nuclear RNAs)] which play important roles in splicing of introns from primary genomic transcripts. Through a combination of RNA–RNA and RNA–protein interactions, two of the snRNPs, U1 and U2, recognize the splice sites and the branch site of introns. A complex remodelling of RNA–RNA and protein-based interactions follows, resulting in the assembly of catalytically competent spliceosomes, in which the snRNAs and their bound proteins play central roles. This process involves formation of extensive base-pairing interactions between U2 and U6, U6 and the 5′ splice site, and U5 and the exonic sequences immediately adjacent to the 5′ and 3′ splice sites. Thus RNA–RNA interactions involving U2, U5 and U6 help position the reacting groups of the first and second steps of splicing. In addition, U6 is also thought to participate in formation of the spliceosomal active site. Furthermore, emerging evidence suggests additional roles for snRNAs in regulation of various aspects of RNA biogenesis, from transcription to polyadenylation and RNA stability. These snRNP-mediated regulatory roles probably serve to ensure the co-ordination of the different processes involved in biogenesis of RNAs and point to the central importance of snRNAs in eukaryotic gene expression.

scholarly journals The role of the PZP domain of AF10 in acute leukemia driven by AF10 translocations

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Brianna J. Klein ◽  
Anagha Deshpande ◽  
Khan L. Cox ◽  
Fan Xuan ◽  
Mohamad Zandian ◽  
...  
Keyword(s):  
Critical Role ◽  
Cellular Transformation ◽  
Acute Leukemias ◽  
Hematopoietic Stem ◽  
Nucleosome Core ◽  
Stem And Progenitor Cells ◽  
Transforming Activity

AbstractChromosomal translocations of the AF10 (or MLLT10) gene are frequently found in acute leukemias. Here, we show that the PZP domain of AF10 (AF10PZP), which is consistently impaired or deleted in leukemogenic AF10 translocations, plays a critical role in blocking malignant transformation. Incorporation of functional AF10PZP into the leukemogenic CALM-AF10 fusion prevents the transforming activity of the fusion in bone marrow-derived hematopoietic stem and progenitor cells in vitro and in vivo and abrogates CALM-AF10-mediated leukemogenesis in vivo. Crystallographic, biochemical and mutagenesis studies reveal that AF10PZP binds to the nucleosome core particle through multivalent contacts with the histone H3 tail and DNA and associates with chromatin in cells, colocalizing with active methylation marks and discriminating against the repressive H3K27me3 mark. AF10PZP promotes nuclear localization of CALM-AF10 and is required for association with chromatin. Our data indicate that the disruption of AF10PZP function in the CALM-AF10 fusion directly leads to transformation, whereas the inclusion of AF10PZP downregulates Hoxa genes and reverses cellular transformation. Our findings highlight the molecular mechanism by which AF10 targets chromatin and suggest a model for the AF10PZP-dependent CALM-AF10-mediated leukemogenesis.

scholarly journals Signaling Pathways Regulating TC21-induced Tumorigenesis

2007 ◽  
Vol 282 (38) ◽  
pp. 27713-27720 ◽  
Author(s):  
Mete Erdogan ◽  
Ambra Pozzi ◽  
Neil Bhowmick ◽  
Harold L Moses ◽  
Roy Zent
Keyword(s):  
P38 Mapk ◽  
Cancer Progression ◽  
Transforming Growth Factor ◽  
Cellular Transformation ◽  
Tumor Formation ◽  
Transformed Cells ◽  
Growth Inhibitory ◽  
Phosphoinositide 3 Kinase

TC21(R-Ras2), a Ras-related GTPase with transforming potential similar to H-, K- and N-Ras, is implicated in the pathogenesis of human cancers. Transforming growth factor β (TGF-β), a cytokine that plays a significant role in modulating tumorigenesis, normally prevents uncontrolled cell proliferation but paradoxically induces proliferation in H-Ras-transformed cancer cells. Although TC21 activates some pathways that mediate cellular transformation by the classical Ras proteins, the mechanisms through which TC21 induces tumor formation and how TGF-β regulates TC21 transformed cells is not known. To better understand the role of TC21 in cancer progression, we overexpressed an activated G23V mutant of TC21 in a nontumorigenic murine mammary epithelial (EpH4) cell line. Mutant TC21-expressing cells were significantly more oncogenic than cells expressing activated G12V H-Ras both in vivo and in vitro. TC21-induced transformation and proliferation required activation of p38 MAPK, mTOR (the mammalian target of rapamycin), and phosphoinositide 3-kinase but not Akt/PKB. Transformation by TC21 rendered EpH4 cells insensitive to the growth inhibitory effects of TGF-β, and the soft agar growth of these cells was increased upon TGF-β stimulation. Despite losing responsiveness to TGF-β-mediated growth inhibition, both Smad-dependent and independent pathways remained intact in TC21-transformed cells. Thus, overexpression of active TC21 in EpH4 cells induces tumorigenicity through the phosphoinositide 3-kinase, p38 MAPK, and mTOR pathways, and these cells lose their sensitivity to the normal growth inhibitory role of TGF-β.

A Proline-Rich Region in the Zeste Protein Essential for Transvection and white Repression by Zeste1

Genetics ◽  
1998 ◽  
Vol 148 (4) ◽  
pp. 1865-1874
Author(s):  
Christina Rosen ◽  
Dale Dorsett ◽  
Joseph Jack
Keyword(s):  
Dna Binding ◽  
Protein Interactions ◽  
Homologous Chromosome ◽  
Protein Complexes ◽  
Dna Binding Protein ◽  
Polycomb Group ◽  
The Self ◽  
Rich Region ◽  
Interaction Domain ◽  
Proline Rich Region

Abstract The DNA-binding protein encoded by the zeste gene of Drosophila activates transcription and mediates interchromosomal interactions such as transvection. The mutant protein encoded by the zeste1 (z1) allele retains the ability to support transvection, but represses white. Similar to transvection, repression requires Zeste-Zeste protein interactions and a second copy of white, either on the homologous chromosome or adjacent on the same chromosome. We characterized two pseudorevertants of z1 (z1-35 and z1-42) and another zeste mutation (z78c) that represses white. The z1 lesion alters a lysine residue located between the N-terminal DNA-binding domain and the C-terminal hydrophobic repeats involved in Zeste self-interactions. The z78c mutation alters a histidine near the site of the z1 lesion. Both z1 pseudorevertants retain the z1 lesion and alter different prolines in a proline-rich region located between the z1 lesion and the self-interaction domain. The pseudorevertants retain the ability to self-interact, but fail to repress white or support transvection at Ultrabithorax. To account for these observations and evidence indicating that Zeste affects gene expression through Polycomb group (Pc-G) protein complexes that epigenetically maintain chromatin states, we suggest that the regions affected by the z1, z78c, and pseudorevertant lesions mediate interactions between Zeste and the maintenance complexes.

Notochord to endoderm signaling is required for pancreas development

Development ◽  
1997 ◽  
Vol 124 (21) ◽  
pp. 4243-4252 ◽  
Author(s):  
S.K. Kim ◽  
M. Hebrok ◽  
D.A. Melton
Keyword(s):  
Gene Expression ◽  
Developmental Stage ◽  
Pancreas Development ◽  
Chick Embryos ◽  
Carboxypeptidase A ◽  
Bud Development ◽  
Dorsal Pancreas ◽  
Stepwise Manner

The role of the notochord in inducing and patterning adjacent neural and mesodermal tissues is well established. We provide evidence that the notochord is also required for one of the earliest known steps in the development of the pancreas, an endodermally derived organ. At a developmental stage in chick embryos when the notochord touches the endoderm, removal of notochord eliminates subsequent expression of several markers of dorsal pancreas bud development, including insulin, glucagon and carboxypeptidase A. Pancreatic gene expression can be initiated and maintained in prepancreatic chick endoderm grown in vitro with notochord. Non-pancreatic endoderm, however, does not express pancreatic genes when recombined with the same notochord. The results suggest that the notochord provides a permissive signal to endoderm to specify pancreatic fate in a stepwise manner.

scholarly journals Postrecruitment Function of Yeast Med6 Protein during the Transcriptional Activation by Mediator Complex

2018 ◽  
Vol 2018 ◽  
pp. 1-9
Author(s):  
Gwang Sik Kim ◽  
Young Chul Lee
Keyword(s):  
Gene Expression ◽  
Reporter Gene ◽  
Transcriptional Activation ◽  
Reporter Gene Expression ◽  
Temperature Sensitive ◽  
Internal Deletion ◽  
Cell Extracts ◽  
Evolutionarily Conserved

Med6 protein (Med6p) is a hallmark component of evolutionarily conserved Mediator complexes, and the genuine role of Med6p in Mediator functions remains elusive. For the functional analysis ofSaccharomyces cerevisiaeMed6p (scMed6p), we generated a series of scMed6p mutants harboring a small internal deletion. Genetic analysis of these mutants revealed that three regions (amino acids 33–42 (Δ2), 125–134 (Δ5), and 157–166 (Δ6)) of scMed6p are required for cell viability and are located at highly conserved regions of Med6 homologs. Notably, the Med6p-Δ2 mutant was barely detectable in whole-cell extracts and purified Mediator, suggesting a loss of Mediator association and concurrent rapid degradation. Consistent with this, the recombinant forms of Med6p having these mutations partially (Δ2) restore or fail (Δ5 and Δ6) to restore in vitro transcriptional defects caused by temperature-sensitivemed6mutation. In an artificial recruitment assay, Mediator containing a LexA-fused wild-type Med6p or Med6p-Δ5 was recruited to thelexAoperator region with TBP and activated reporter gene expression. However, the recruitment of Mediator containing LexA-Med6p-Δ6 tolexAoperator region resulted in neither TBP recruitment nor reporter gene expression. This result demonstrates a pivotal role of Med6p in the postrecruitment function of Mediator, which is essential for transcriptional activation by Mediator.

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