Role of hPHF1 in H3K27 Methylation and Hox Gene Silencing

ABSTRACT Polycomb group (PcG) proteins are required for maintaining the silent state of the homeotic genes and other important developmental regulators. The silencing function of the PcG proteins has been linked to their intrinsic histone modifying enzymatic activities. The EED-EZH2 complex, containing the core subunits EZH2, EED, SUZ12, and RbAp48, functions as a histone H3K27-specific methyltransferase. Here we describe the identification and characterization of a related EED-EZH2 protein complex which is distinguished from the previous complex by the presence of another PcG protein, hPHF1. Consistent with the ability of hPHF1 to stimulate the enzymatic activity of the core EED-EZH2 complex in vitro, manipulation of mPcl1, the mouse counterpart of hPHF1, in NIH 3T3 cells and cells of the mouse male germ cell line GC1spg results in global alteration of H3K27me2 and H3K27me3 levels and Hox gene expression. Small interfering RNA-mediated knockdown of mPcl1 affects association of the Eed-Ezh2 complex with certain Hox genes, such as HoxA10, as well as Hox gene expression concomitant with an alteration on the H3K27me2 levels of the corresponding promoters. Therefore, our results reveal hPHF1 as a component of a novel EED-EZH2 complex and demonstrate its important role in H3K27 methylation and Hox gene silencing.

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Altered cellular proliferation and mesoderm patterning in Polycomb-M33-deficient mice

Development ◽

10.1242/dev.124.3.721 ◽

1997 ◽

Vol 124 (3) ◽

pp. 721-729 ◽

Cited By ~ 22

Author(s):

N. Core ◽

S. Bel ◽

S.J. Gaunt ◽

M. Aurrand-Lions ◽

J. Pearce ◽

...

Keyword(s):

Cellular Proliferation ◽

Hox Genes ◽

Es Cells ◽

Embryonic Stem ◽

Axial Skeleton ◽

Polycomb Group ◽

Homeotic Genes ◽

Loss Of Function ◽

Polycomb Group Genes

In Drosophila, the trithorax-group and the Polycomb-group genes are necessary to maintain the expression of the homeobox genes in the appropriate segments. Loss-of-function mutations in those groups of genes lead to misexpression of the homeotic genes resulting in segmental homeotic transformations. Recently, mouse homologues of the Polycomb-group genes were identified including M33, the murine counterpart of Polycomb. In this report, M33 was targeted in mice by homologous recombination in embryonic stem (ES) cells to assess its function during development. Homozygous M33 (−/−) mice show greatly retarded growth, homeotic transformations of the axial skeleton, sternal and limb malformations and a failure to expand in vitro of several cell types including lymphocytes and fibroblasts. In addition, M33 null mutant mice show an aggravation of the skeletal malformations when treated to RA at embryonic day 7.5, leading to the hypothesis that, during development, the M33 gene might play a role in defining access to retinoic acid response elements localised in the regulatory regions of several Hox genes.

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Regulation of Hox Genes by Polycomb Group Protein EZH2 Plays a Critical Role in Diffuse Large B-Cell Lymphoma Cells.

Blood ◽

10.1182/blood.v110.11.1562.1562 ◽

2007 ◽

Vol 110 (11) ◽

pp. 1562-1562

Author(s):

Irina Velichutina ◽

Ari Melnick

Keyword(s):

Gene Expression ◽

B Cells ◽

Target Genes ◽

Hox Genes ◽

Critical Role ◽

Polycomb Group ◽

Lymphoma Cells ◽

Self Renewal ◽

Hox Gene ◽

Large B Cell

Abstract Coordinated regulation of Hox gene expression during hematopoiesis is epigenetically controlled via chromatin modification by Polycomb group (PcG) and Trithorax (MLL) protein complexes. Whereas the oncogenic potential of certain HOX genes in leukemia has already been defined, little is known about their role in Diffuse Large B-cell Lymphomas (DLBCL). The primary focus of our studies is to determine the contribution of PcG-mediated repression of HOX and other genes to DLBCL pathogenesis. The PcG protein, Ezh2, is vital for maintaining both pluripotency of stem cells and identity of differentiated cells. Ezh2 tri-methylates lysine K27 of histone 3 (H3K27me3), a histone modification associated with gene silencing. Importantly, Ezh2 is frequently overexpressed in DLBCLs suggesting a role for EZH2 in lymphomagenesis. In support to this notion we discovered that Ezh2 is essential for DLBCL cell survival. By depleting Ezh2 level using RNAi, we found that loss of Ezh2 triggers cell cycle arrest and death of DLBCL cells. This finding prompted us to initiate functional studies aimed at uncovering Ezh2 target genes that mediate the observed cellular response in DLBCL cells. We first focused on a potential role of Ezh2 in regulation of HOX genes. We compared and contrasted Ezh2 targets in both normal Germinal Center (GC) B-cells and GC-derived DLBCLs to determine the normal and pathologic function of EZH2. We employed a tiling ChIP-chip approach covering the four human HOX clusters and mapped Ezh2 and H3K27m3 within HOX gene clusters. We further verified gene expression status of a subset of Hox genes by QPCR. These data indicated that Ezh2 and its cognate H3K27m3 mark are present at promoters of HoxC genes in both mature GC B-cells and GC-derived lymphoma cells, thereby driving the HoxC locus silent, suggesting that both rapidly dividing GC cells and GC-derived lymphoma cells require epigenetic silencing of this locus in order to maintain their phenotype. Both Ezh2 and the corresponding H3K27m3 transcription repression mark are absent within the promoter region of HoxA9 gene. HoxA9 promotes stem cell self-renewal and it is aberrantly activated in AML cells. This observation is especially striking as the HoxA9 is embedded into the Ezh2-sealed region in DLBCL cells, suggesting an Ezh2-independent mode of regulation. We are in the process of testing functional significance of this finding for lymphoma pathogenesis. we found that HoxB genes that are differentially expressed in progenitor vs. lineage committed cells are silent in DLBCL cells according to H3K27m3/Ezh2 pattern and gene expression analysis. Intriguingly, the early progenitor specific gene, HoxB3, is uniquely not bound by EZH2 nor H3K27 methylated and was highly expressed in lymphoma cells. This finding underscores a potential functional significance of re-expression of genes that control cell self-renewal in malignances that derive from mature B cells. We also examined transcriptional programming by EZH2 at the genomic level by ChIP-on-chip using NimbleGen 24,000 promoter arrays. EZH2 was bound to ∼1700 promoters in DLBCL cells and a similar number of genes displayed H3K27 methylation. Gain and loss of function studies are underway to identify the contribution of the most likely EZH2 direct targets genes to the DLBCL survival including both HOX genes and other genomic direct target genes. Taken together, our data suggest a critical role for EZH2 mediated epigenetic silencing of HOX and other genes in DLBCL - and implicate aberrant HOX gene expression in DLBCL pathogenesis.

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Comprehensive Analysis Of HOX Gene Expression and DNA Methylation From 189 Primary AMLs Demonstrates Canonical Patterns Associated With Hematopoietic Stem/Progenitors and Recurrent AML Mutations

Blood ◽

10.1182/blood.v122.21.2496.2496 ◽

2013 ◽

Vol 122 (21) ◽

pp. 2496-2496 ◽

Cited By ~ 2

Author(s):

David H Spencer ◽

Margaret A. Young ◽

Jeffery M. Klco ◽

Timothy J. Ley

Keyword(s):

Gene Expression ◽

Stem Cells ◽

Hox Genes ◽

Embryonic Stem ◽

Normal Karyotype ◽

Hematopoietic Stem ◽

Hox Gene ◽

Cd34 Cells ◽

Methylation Patterns

Abstract HOX genes encode a family of homeodomain transcription factors with important roles in hematopoiesis. Expression of HOX genes is also a common feature of acute myeloid leukemia (AML), and functional studies have suggested that HOX-dependent pathways may contribute to leukemogenesis. Although HOX expression is known to correlate with specific AML mutations, the patterns of expression of all 39 HOX genes in primary AML samples, and their relationships with recurrent AML mutations, are incompletely understood. In addition, little is known about the influence of AML mutations on DNA methylation at the HOX loci, and the relationship between HOX gene expression and methylation in AML. In this study, we carried out a combined analysis of gene expression data from microarray and RNA-sequencing platforms and genome-wide DNA array-based methylation from 189 primary AML samples that have been previously characterized by either whole-genome or whole exome sequencing. We also measured expression and methylation using the same platforms from normal bone marrow subsets, including CD34+ cells, promyelocytes, monocytes, neutrophils and lymphocytes, and obtained expression data from CD34+ hematopoietic precursors generated from in vitro differentiation of human embryonic stem cells. Our analysis confirmed previous work on the general patterns of HOX expression in AML. The HOXA and HOXB genes showed variation both within each cluster and across the AMLs, although high level expression was restricted to a subset of these genes, including HOXA3, HOXA5, HOXA7, HOXA9, HOXA10, HOXB2-HOXB4, and HOXB6, as well as HOX cofactor MEIS1; HOXC and HOXD genes were minimally expressed in all of the samples. These observations were orthogonally validated by RNA-seq, and with a targeted Nanostring expression platform. Consistent with previous studies, MLL-positive AML samples (n=11) expressed only HOXA genes and MEIS1. AML samples with CBFB-MYH11 rearrangements (n=12) showed expression of only MEIS1, and HOXB2-HOXB4 at moderate levels; RUNX1-RUNX1T1 (n=7) and PML-RARA (n=19) samples did not detectably express any HOX genes. In AMLs with a normal karyotype (n=85), we observed two distinct patterns; one pattern displayed little or no HOX gene expression (7/85; 8%), and another displayed canonical expression of a specific subset of the HOXA and HOXB genes and MEIS1 (78/85; 92%) with similar relative HOX gene expression levels in all cases. Comparison of this pattern with normal bone marrow revealed the same HOX expression pattern in normal CD34+ cells; additional analysis showed that this pattern was confined to hematopoietic stem/progenitor cells, but was not seen in more mature cells, including other CD34+ subsets, promyelocytes, monocytes and neutrophils. We also measured HOX gene expression in CD34+ hematopoietic precursors generated from in vitro differentiation of human embryonic stem cells, which revealed expression of only MEIS1 and the canonical HOXB genes, suggesting that activation of these genes may represent the earliest events in the HOX pathway of hematopoietic development. Correlation of HOX expression with recurrent AML mutations by gene set enrichment analysis demonstrated a significant association with NPM1 (P<10-4) and DNMT3A (P<10-2) mutations, but not with other recurrent somatic mutations, including FLT3,IDH1/IDH2, and TET2. Methylation at the HOX loci demonstrated patterns that correlated with HOX expression, including hypomethylation at HOX promoters in samples with high expression. However, additional mutation-specific patterns were apparent. For example, NPM1-mutant AMLs demonstrated a distinct methylation pattern that included hypomethylation at the HOXB3 promoter, which was not shared with CBFB-MYH11 cases or other AMLs with HOXB3 expression. In summary, our comprehensive analysis demonstrates canonical expression and methylation patterns at the HOX loci in AML. These patterns correspond to specific recurrent AML mutations, and the dominant pattern in most normal karyotype AMLs mimics the signature of hematopoietic stem cells. This supports previous observations of developmental regulation of HOX genes in hematopoiesis, and implies that this normal stem cell signature is “captured” in the majority of AMLs with normal karyotype. In addition, distinct methylation patterns at HOX loci suggest that multiple regulatory mechanisms are involved in HOX expression in AML. Disclosures: No relevant conflicts of interest to declare.

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Leukemic Pattern Of HOX Gene Expression Is Driven By Genetic Aberrations Through Epigenetic Modifiers

Blood ◽

10.1182/blood.v122.21.2504.2504 ◽

2013 ◽

Vol 122 (21) ◽

pp. 2504-2504

Author(s):

Julia Starkova ◽

Karolina Kramarzova ◽

Karel Fiser ◽

Ester Mejstrikova ◽

Katerina Rejlova ◽

...

Keyword(s):

Gene Expression ◽

Expression Pattern ◽

Hox Genes ◽

De Novo ◽

In Vitro Experiments ◽

Genetic Aberrations ◽

Hox Gene ◽

Epigenetic Modifiers ◽

Fab Classification

Abstract Introduction Homeobox (HOX) genes encode transcription factors crucial in embryogenesis. They are often dysregulated in malignancies including leukemias. The aberrant HOX gene expression and its regulation in leukemic cells is neither completely described nor understood. Aims Our main aim was to determine whether the leukemic HOX gene expression pattern is driven by differentiation stage of hematopoietic cells or determined de novo during the process of malignant transformation. Consequentially, we aimed to study the role epigenetic modifiers in regulation of HOX gene expression in normal and malignant hematopoiesis. Methods The expression pattern of HOX genes (cluster of HOX A and B) and epigenetic modifiers (DNMT1, DNMT3a, DNMT3b, EZH2, BMI-1, MLL, JMJD3, UTX) was assessed by qPCR in 8 FACS-sorted subpopulations of healthy BM representing stages of myeloid differentiation (each sample representing a pool of cells sorted from five individuals). The leukemic expression pattern of these genes was analyzed in diagnostic BM samples of childhood AML patients with typical genotypic and morphological (FAB classification) characteristics (N=46). In vitro experiments were performed using NB4 cell line. Results As expected HOX genes were gradually downregulated during normal differentiation of granulocytic and monocytic lineages (assessed in four consecutive differentiation stages for each lineage). In AML samples, HOX gene expression patterns differed significantly among morphological subtypes. However, HOX gene expression did not correlate among subtypes of AML and their physiologically differentiated counterparts. Interestingly, unsupervised hierarchical clustering (HCA) divided AML patients into four main clusters characterized by the presence of prevalent gene rearrangement (PML-RARa, AML1-ETO, MLL rearrangements and NK-AML). The presence of PML/RARa rearrangement was strongly associated with the lowest expression of both HOXA and HOXB clusters, while the other groups had more variable expression of HOX genes. Moreover, the effect of genetic aberrations on HOX gene expression was clearly apparent within AML M2 and M4 subtypes, where AML1/ETO+ or CBFb/MYH11+ patients had significantly lower expression of HOX genes compared to patients with the same FAB classification but without the rearrangements. The expression pattern of epigenetic modifiers in sorted subpopulations of healthy BM followed their expected role in transcriptional regulation during differentiation. However, there was no relation of this pattern to HOX gene expression. On the contrary, in AML samples, the expression levels of epigenetic modifiers clearly correlated with expression profile of HOX genes. These results were supported by unsupervised HCA based on the expression of epigenetic modifiers that showed upregulation of histon demethylases JMJD3 and UTX together with downregulation of DNMT3b in concordance with high levels of HOX genes. Negative correlation between JMJD3 and DNMT3b expression was observed in all leukemic samples (p=0.03); most apparently in PML/RARa+ patients. Therefore we further studied the impact of genetic aberrations on the epigenetic regulation of HOX gene expression in vitrowith PML-RARa+ cell line. Treatment of NB4 cells with ATRA (8, 24hours, 1uM, 10uM) increased the levels of particular HOX genes (HOXA5, A7, B4, B7; FCA=2.8; 1.7; 4; 4 respectively) as well as JMJD3 (FCA=3) and UTX (FCA=1.6). Concordantly, the expression of DNMT3b (FCA=5) was downregulated. The hypothetical driving effect of PML-RARa on de novo determination of leukemic HOX gene expression is further supported by our Results. PML-RARa+ patients had the lowest HOX gene expression regardless of their FLT3/ITD status – previously shown to upregulate strongly HOX genes expression. Conclusion We conclude that the leukemic expression pattern of HOX genes does not reflect the differentiation stages of malignant cells. Our data also demonstrate different contribution of epigenetic modifiers to the HOX gene expression in healthy and malignant hematopoiesis. Moreover, HCA and expression data together with the results of in vitro experiments suggest that the specific molecular aberrations (as exemplified by PML-RARa) participate in regulation of leukemic HOX gene expression through epigenetic changes. Supported by GACR P304/12/2214, GAUK 568213, 00064203. Disclosures: No relevant conflicts of interest to declare.

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Analysis of Hox gene expression in the chick limb bud

Development ◽

10.1242/dev.122.5.1449 ◽

1996 ◽

Vol 122 (5) ◽

pp. 1449-1466 ◽

Cited By ~ 26

Author(s):

C.E. Nelson ◽

B.A. Morgan ◽

A.C. Burke ◽

E. Laufer ◽

E. DiMambro ◽

...

Keyword(s):

Gene Expression ◽

Cell Proliferation ◽

Sonic Hedgehog ◽

Hind Limb ◽

Hox Genes ◽

Expression Patterns ◽

Limb Bud ◽

Posterior Portion ◽

Hox Gene ◽

Hoxd Gene

The vertebrate Hox genes have been shown to be important for patterning the primary and secondary axes of the developing vertebrate embryo. The function of these genes along the primary axis of the embryo has been generally interpreted in the context of positional specification and homeotic transformation of axial structures. The way in which these genes are expressed and function during the development of the secondary axes, particularly the limb, is less clear. In order to provide a reference for understanding the role of the Hox genes in limb patterning, we isolated clones of 23 Hox genes expressed during limb development, characterized their expression patterns and analyzed their regulation by the signalling centers which pattern the limb. The expression patterns of the Abd-B-related Hoxa and Hoxd genes have previously been partially characterized; however, our study reveals that these genes are expressed in patterns more dynamic and complex than generally appreciated, only transiently approximating simple, concentric, nested domains. Detailed analysis of these patterns suggests that the expression of each of the Hoxa and Hoxd genes is regulated in up to three independent phases. Each of these phases appears to be associated with the specification and patterning of one of the proximodistal segments of the limb (upper arm, lower arm and hand). Interestingly, in the last of these phases, the expression of the Hoxd genes violates the general rule of spatial and temporal colinearity of Hox gene expression with gene order along the chromosome. In contrast to the Abd-B-related Hoxa and Hoxd genes, which are expressed in both the fore and hind limbs, different sets of Hoxc genes are expressed in the two limbs. There is a correlation between the relative position of these genes along the chromosome and the axial level of the limb bud in which they are expressed. The more 3′ genes are expressed in the fore limb bud while the 5′ genes are expressed in the hind limb bud; intermediate genes are transcribed in both limbs. However, there is no clear correlation between the relative position of the genes along the chromosome and their expression domains within the limb. With the exception of Hoxc-11, which is transcribed in a posterior portion of the hind limb, Hoxc gene expression is restricted to the anterior/proximal portion of the limb bud. Importantly, comparison of the distributions of Hoxc-6 RNA and protein products reveals posttranscriptional regulation of this gene, suggesting that caution must be exercised in interpreting the functional significance of the RNA distribution of any of the vertebrate Hox genes. To understand the genesis of the complex patterns of Hox gene expression in the limb bud, we examined the propagation of Hox gene expression relative to cell proliferation. We find that shifts in Hox gene expression cannot be attributed to passive expansion due to cell proliferation. Rather, phase-specific Hox gene expression patterns appear to result from a context-dependent response of the limb mesoderm to Sonic hedgehog. Sonic hedgehog (the patterning signal from the Zone of Polarizing Activity) is known to be able to activate Hoxd gene expression in the limb. Although we find that Sonic hedgehog is capable of initiating and polarizing Hoxd gene expression during both of the latter two phases of Hox gene expression, the specific patterns induced are not determined by the signal, but depend upon the temporal context of the mesoderm receiving the signal. Misexpression of Sonic hedgehog also reveals that Hoxb-9, which is normally excluded from the posterior mesenchyme of the leg, is negatively regulated by Sonic hedgehog and that Hoxc-11, which is expressed in the posterior portion of the leg, is not affected by Sonic hedgehog and hence is not required to pattern the skeletal elements of the lower leg.

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The Role of Histone Demethylases and DNA Methyltransferases in the Transcription Regulation of HOX Genes in PML-RARa+ AML Patients

Blood ◽

10.1182/blood.v128.22.3921.3921 ◽

2016 ◽

Vol 128 (22) ◽

pp. 3921-3921

Author(s):

Katerina Rejlova ◽

Alena Musilova ◽

Martina Slamova ◽

Karel Fiser ◽

Karolina Skvarova Kramarzova ◽

...

Keyword(s):

Gene Expression ◽

Hox Genes ◽

Fold Increase ◽

Dna Methyltransferases ◽

Histone Mark ◽

Gene Promoters ◽

Histone Demethylases ◽

Atra Treatment ◽

Hox Gene

Abstract Homeobox genes (HOX) encode transcription factors that are frequently deregulated in leukemias. Our previous results showed that HOX gene expression differs among genetically characterized subtypes of pediatric acute myeloid leukemia (AML). Specifically, PML-RARa positive AML patients have overall lowest HOX gene expression which positively correlates with expression of histone 3 lysine 27 (H3K27) demethylases - JMJD3 and UTX and negatively with the expression of DNA methyltransferases - DNMT3a and DNMT3b. Interestingly, JMJD3 was already shown to be a direct target of PML-RARa protein (Martens, JH et al, 2010, Cancer Cell). From these findings we postulated a hypothesis that reduced levels of HOX genes in PML-RARa positive AML are a consequence of suppressed expression of histone demethylases resulting in increased H3K27 methylation and/or of elevated levels of DNMTs leading to de novoDNA methylation. We studied the role of histone demethylases and DNMTs in the regulation of HOX gene expression and the effect of treatment in PML-RARa positive cell lines (NB4 and ATRA-resistant clones NB4-LR2 and NB4-MR2). We treated NB4 cell line by all-trans retinoic acid (ATRA; 1uM), which was described to release the differentiation block caused by the presence of PML-RARa and to degrade the fusion protein. We observed that expression of particular HOX genes (HOXA1, HOXA3, HOXA4, HOXA5, HOXA7, HOXB4, HOXB6) measured by qPCR was significantly increased after ATRA treatment. While the level of JMJD3 was significantly increased upon ATRA treatment as well, the expression of UTX did not change. Furthermore, we detected significantly reduced expression of DNMT3b gene. To exclude a non-specific effect of ATRA, independent of PML-RARa, we used resistant clones LR2 and MR2 bearing mutations in retinoic acid-binding domain. HOX gene expression together with JMJD3, UTX and DNMT3b expression did not change upon ATRA treatment. These results confirm the PML-RARa-dependent regulation of HOX genes. To test the role of JMJD3 in the HOX gene expression regulation, we cultured NB4 cells with a specific inhibitor of histone demethylases, GSK-J4 (1 uM, 10 uM), in the presence of ATRA. The co-treatment caused significant decrease in the expression of studied HOX genes (HOXA1, HOXA3, HOXA5, HOXA7, HOXA10, HOXB4, HOXB6) in comparison to ATRA alone which supports the role of JMJD3 in the transcription regulation. Further, we performed chromatin immunoprecipitation (ChIP) to investigate if the changes of HOX gene expression upon ATRA and GSK-J4 treatment would correspond with changes of histone code on HOX gene promoter regions. ATRA treatment caused reduction of repressive histone mark (H3K27me3) on particular HOX gene promoters (HOXA1, HOXA3, HOXA5, HOXA7), by contrast, combinational treatment of ATRA and GSK-J4 reversed this effect. Accordingly, we detected that ATRA/GSK-J4 co-treatment reduced active histone mark H3K4me2. Next we were interested if JMJD3 inhibition would interfere with the differentiation effect of ATRA. As shown previously, ATRA treatment alone caused differentiation of NB4 cell line whereas the combination with GSK-J4 did not reduce the effect. Interestingly, in addition to differentiation it led cells to apoptosis. Combination of drugs (ATRA - 1uM, GSK-J4 - 1, 2, 5uM) increased significantly the percentage of dead cells in comparison to ATRA or GSK treatment alone (GSK-J4 alone vs in combination with ATRA, 1uM - 1.8 fold, 2uM - 2.2 fold, 5 uM - 2.3 fold increase). Next we measured apoptosis in resistant clones LR2 and MR2. In both cases the highest concentration used of GSK-J4 (5uM) in combination with ATRA caused significant increase of dead cells as well (LR2 - 2.1 fold, MR2 - 2.0 fold increase). Our results indicate that JMJD3 is responsible for the regulation of HOX gene expression in PML-RARa positive leukemia since changes of HOX gene expression correspond with histone modifications on the regions of HOX gene promoters. We assume that DNA methylation driven by DNMT3b can also participate in this process. Moreover, our findings demonstrate potential therapeutic implications of GSK-J4 inhibitor in combination with ATRA in patients with acute promyelocytic leukemia who are not responsive to ATRA monotherapy. Supported by P304/12/2214 and GAUK 196616 Disclosures No relevant conflicts of interest to declare.

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A Wnt signaling pathway controls hox gene expression and neuroblast migration in C. elegans

Development ◽

10.1242/dev.126.1.37 ◽

1999 ◽

Vol 126 (1) ◽

pp. 37-49 ◽

Cited By ~ 9

Author(s):

J.N. Maloof ◽

J. Whangbo ◽

J.M. Harris ◽

G.D. Jongeward ◽

C. Kenyon

Keyword(s):

Gene Expression ◽

Wnt Signaling ◽

Wnt Pathway ◽

Hox Genes ◽

Wnt Signaling Pathway ◽

Beta Catenin ◽

Hox Gene ◽

C Elegans ◽

Neuroblast Migration ◽

Pathway Gene

The specification of body pattern along the anteroposterior (A/P) body axis is achieved largely by the actions of conserved clusters of Hox genes. Limiting expression of these genes to localized regional domains and controlling the precise patterns of expression within those domains is critically important for normal patterning. Here we report that egl-20, a C. elegans gene required to activate expression of the Hox gene mab-5 in the migratory neuroblast QL, encodes a member of the Wnt family of secreted glycoproteins. We have found that a second Wnt pathway gene, bar-1, which encodes a beta-catenin/Armadillo-like protein, is also required for activation of mab-5 expression in QL. In addition, we describe the gene pry-1, which is required to limit expression of the Hox genes lin-39, mab-5 and egl-5 to their correct local domains. We find that egl-20, pry-1 and bar-1 all function in a linear genetic pathway with conserved Wnt signaling components, suggesting that a conserved Wnt pathway activates expression of mab-5 in the migratory neuroblast QL. Moreover, we find that members of this Wnt signaling system play a major role in both the general and fine-scale control of Hox gene expression in other cell types along the A/P axis.

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Signalling by FGF8 from the isthmus patterns anterior hindbrain and establishes the anterior limit of Hox gene expression

Development ◽

10.1242/dev.127.1.177 ◽

2000 ◽

Vol 127 (1) ◽

pp. 177-186 ◽

Cited By ~ 12

Author(s):

C. Irving ◽

I. Mason

Keyword(s):

Gene Expression ◽

Hox Genes ◽

Protein A ◽

Anterior Segment ◽

Morphogen Gradient ◽

Current Evidence ◽

Hox Gene ◽

Developmental Fate ◽

Developmental Patterning

Current evidence suggests that the anterior segment of the vertebrate hindbrain, rhombomere 1, gives rise to the entire cerebellum. It is situated where two distinct developmental patterning mechanisms converge: graded signalling from an organising centre (the isthmus) located at the midbrain/hindbrain boundary confronts segmentation of the hindbrain. The unique developmental fate of rhombomere 1 is reflected by it being the only hindbrain segment in which no Hox genes are expressed. In this study we show that ectopic FGF8 protein, a candidate for the isthmic organising activity, is able to induce and repress gene expression within the hindbrain in a manner appropriate to rhombomere 1. Using a heterotopic, heterospecific grafting strategy we demonstrate that rhombomere 1 is able to express Hox genes but that both isthmic tissue and FGF8 inhibit their expression. Inhibition of FGF8 function in vivo shows that it is responsible for defining the anterior limit of Hox gene expression within the developing brain and thereby specifies the extent of the rl territory. Previous studies have suggested that a retinoid morphogen gradient determines the axial limit of expression of individual Hox genes within the hindbrain. We propose a model whereby activation by retinoids is antagonised by inhibition by FGF8 in the anterior hindbrain to set aside the territory from which the cerebellum will develop.

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Induction of HOX genes by HCV infection via impairment of histone H2A monoubiquitination.

Journal of Virology ◽

10.1128/jvi.01784-20 ◽

2020 ◽

pp. JVI.01784-20

Author(s):

Hirotake Kasai ◽

Kazuki Mochizuki ◽

Tomohisa Tanaka ◽

Atsuya Yamashita ◽

Yoshiharu Matsuura ◽

...

Keyword(s):

Hepatitis C ◽

Protein Expression ◽

Hox Genes ◽

Core Protein ◽

Gene Promoter ◽

Hcv Infection ◽

Histone H2a ◽

Hox Gene ◽

The Core ◽

Hcv Core Protein

Hepatitis C virus (HCV) infection causes liver pathologies, including hepatocellular carcinoma (HCC). Homeobox (HOX) gene products regulate embryonic development and are associated with tumorigenesis, although the regulation of HOX genes by HCV infection has not been clarified in detail. We examined the effect of HCV infection on HOX gene expression. In this study, HCV infection induced more than half of the HOX genes and reduced the level of histone H2A monoubiquitination on lysine (K) 119 (H2Aub), which represses HOX gene promoter activity. HCV infection also promoted proteasome-dependent degradation of RNF2, which is an E3 ligase mediating H2A monoubiquitination as a component of polycomb repressive complex 1. Since full-genomic replicon cells but not subgenomic replicon cells exhibited reduced RNF2 and H2Aub levels and induction of HOX genes, we focused on the core protein. Expression of the core protein reduced the amounts of RNF2 and H2Aub and induced HOX genes. Treatment with LY-411575, which can reduce HCV core protein expression via SPP inhibition without affecting other viral proteins, dose-dependently restored the amounts of RNF2 and H2Aub in HCV-infected cells and impaired the induction of HOX genes and production of viral particles but not viral replication. The chromatin immunoprecipitation assay results also indicated infection- and proteasome-dependent reductions in H2Aub located in HOX gene promoters. These results suggest that HCV infection or core protein induces HOX genes by impairing histone H2A monoubiquitination via a reduction in the RNF2 level.Importance Recently sustained virologic response can be achieved by direct acting antiviral therapy in most of hepatitis C patients. Unfortunately, DAA therapy does not completely eliminate a risk of HCC. Several epigenetic factors, including histone modifications, are well known to contribute to HCV-associated HCC. However, the regulation of histone modifications by HCV infection has not been clarified in detail. In this study, our data suggest that HCV infection or HCV core protein expression impairs monoubiquitination of histone H2A K119 in HOX gene promoter via destabilization of RNF2 and then induces HOX genes. Several lines of evidence suggest that the expression of several HOX genes is dysregulated in certain types of tumors. These findings reveal a novel mechanism of HCV-related histone modification and may provide information about new targets for diagnosis and prevention of HCC occurrence.

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Posterior Hox Gene Expression and Differential Androgen Regulation in the Developing and Adult Rat Prostate Lobes

Endocrinology ◽

10.1210/en.2006-1250 ◽

2007 ◽

Vol 148 (3) ◽

pp. 1235-1245 ◽

Cited By ~ 48

Author(s):

Liwei Huang ◽

Yongbing Pu ◽

David Hepps ◽

David Danielpour ◽

Gail S. Prins

Keyword(s):

Gene Expression ◽

Hox Genes ◽

Prostate Gland ◽

Hox Gene ◽

Expression Levels ◽

Adult Rat ◽

Organ Specific ◽

Androgen Regulation ◽

Hox Code ◽

Rat Prostate

Axis positioning and tissue determination during development involve coordinated expression of Hox genes throughout the body. The most posterior Hox gene clusters are involved in prostate organogenesis. In the present study, we characterized and compared the expression profiles of posterior (5′) Hox genes in the separate lobes of the adult rat prostate gland, the coagulating gland, seminal vesicles, and epididymis using quantitative real-time RT-PCR. These genes include Hoxa9–11, Hoxa13, Hoxd13, and Hoxb13. We identified a unique Hox code for each of these organs and propose that this contributes to the organ-specific and prostate lobe-specific identities in the adult rat. Using the ventral prostate (VP) as a model, we characterized the Hox genes expression patterns over time from birth through adulthood. Expression levels of the three Hox13 genes and Hoxa10 were significantly higher in the adult VP compared with the neonatal developing VP suggesting an important role during adult homeostasis. In contrast, Hoxa9 and Hoxa11 levels declined after morphogenesis suggesting a specific developmental role. Overall, the Hoxb13 gene exhibited the most striking temporal and organ-specific differences. Using in situ hybridization and immunohistochemistry, a distinct Hoxb13 anterior-to-posterior expression gradient was observed with the highest expression levels in the VP luminal epithelial cells, moderate levels in the lateral prostate, and low expression in the dorsal prostate. An expression gradient was also observed along the ductal length in all three prostate lobes with strongest expression at the distal tips and limited expression in the proximal ducts. After infection with a lentivirus expressing the Hoxb13 gene, NRP-152 cells cultured under nondifferentiating conditions exhibited robust cytokeratin 8 immunostain indicating that Hoxb13 expression drives luminal cell differentiation in the rat epithelium. Androgen regulation of prostatic Hox gene expression was examined during development in vitro and after castration in the adult rat. In the neonatal VP, all six Hox genes were significantly up-regulated by androgens, whereas none of the genes were affected by testosterone in the lateral prostate. In the adult rat, castration resulted in up-regulation of Hoxa9 and Hoxa13 in the VP and down-regulation of Hoxb13 in the dorsal prostate and lateral prostate. Taken together, we conclude that the prostatic Hox genes reach a destined expression level at specific developmental time points in the prostate gland and possess differential androgenic regulation in a temporal and lobe-specific manner. We suggest that this timely Hox code participates in determining lobe-specific prostatic identity and cellular differentiation.

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