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M van Lohuizen

Publications and source records attributed to M van Lohuizen.

At least 37 records · Page 2Linked to original sources
PubMed1997

MPc2, a new murine homolog of the Drosophila polycomb protein is a member of the mouse polycomb transcriptional repressor complex.

The evolutionarily conserved polycomb and trithorax-group genes are required to maintain stable expression patterns of homeotic genes and other target genes throughout development. Here, we report the cloning and characterization of a novel mouse polycomb homolog, MPc2, in addition to the previously described M33 polycomb gene. Co-immunoprecipitations and subnuclear co-localization studies show that MPc2 interacts with the mouse polycomb-group oncoprotein Bmi1 and is a new member of the mouse polycomb multiprotein complex. Gal4DB-MPc2 or -M33 fusion proteins mediate a five- to tenfold repression of stably integrated reporter constructs carrying GAL4 binding sites, demonstrating that these proteins are transcriptional repressors. The MPc2 gene is localized on chromosome 11, in close proximity to the classical mouse mutations tail short (Ts) and rabo torcido (Rbt). Ts and Rbt hemizygous mice display anemia and transformations of the axial skeleton reminiscent of phenotypes observed in mice with mutated polycomb or trithorax-group genes, suggesting that MPc2 is a candidate gene for Ts and Rbt.

Amino Acid Sequence↗
PubMed1997

Pertubation of B and T cell development and predisposition to lymphomagenesis in Emu Bmi1 transgenic mice require the Bmi1 RING finger.

Proviral activation of the Bmi1 gene has implicated Bmi1 as a collaborator of c-Myc in lymphomagenesis. To determine the effect of Bmi1 overexpression on hematopoiesis and lymphomagenesis transgenic mice were generated that overexpress different forms of the Bmi1 protein in their lymphoid compartment. Emu Bmi1 transgenic mice, overexpressing the wild type Bmi1 protein showed a perturbed lymphoid development and were highly susceptible to B and T cell lymphomagenesis. Mutational analysis of the Bmi1 protein demonstrated that the conserved N-terminal RING finger and central part of Bmi1 are essential for its oncogenic potential whereas the C-terminal Pro-Ser rich region is not required. We have used provirus tagging in the Emu Bmi1 mice to identify genes that cooperate with Bmi1 in lymphomagenesis. MoMLV infection in Emu Bmi1 transgenic mice accelerated lymphoma development. Proviral activation of the Pim and Myc genes but not the Gfi1 gene were frequently observed in these tumors. These results demonstrate that Bmi1 is a potent oncogene and suggest that it plays an important role in early lymphoid development.

Animals↗
PubMed1997

Identification of Bmi1-interacting proteins as constituents of a multimeric mammalian polycomb complex.

The Bmi1 gene has been identified as a mouse Polycomb group (Pc-G) gene implicated in the regulation of Hox gene expression. Here we describe the characterization of a Bmi binding protein Mph1, which shares similarity to Drosophila polyhomeotic. Coimmunoprecipitation experiments indicate that Bmi1 and Mph1, as well as the Mel18 and M33 proteins described previously, are constituents of a multimeric protein complex in mouse embryos and human cells. A central domain of Bmi1 interacts with the carboxyl terminus of Mph1, whereas a conserved alpha-helical domain in the Mph1 protein is required for its homodimerization. Transgenic mice overexpressing various mutant Bmi1 proteins demonstrate that the central domain of Bmil is required for the induction of anterior transformations of the axial skeleton. Bmi1, M33, and Mph1 show an overlapping speckled distribution in interphase nuclei. These data provide molecular evidence for the existence of a mammalian Polycomb complex.

Amino Acid Sequence↗
PubMed1997

Identification and characterization of interactions between the vertebrate polycomb-group protein BMI1 and human homologs of polyhomeotic.

In Drosophila melanogaster, the Polycomb-group (PcG) genes have been identified as repressors of gene expression. They are part of a cellular memory system that is responsible for the stable transmission of gene activity to progeny cells. PcG proteins form a large multimeric, chromatin-associated protein complex, but the identity of its components is largely unknown. Here, we identify two human proteins, HPH1 and HPH2, that are associated with the vertebrate PcG protein BMI1. HPH1 and HPH2 coimmunoprecipitate and cofractionate with each other and with BMI1. They also colocalize with BMI1 in interphase nuclei of U-2 OS human osteosarcoma and SW480 human colorectal adenocarcinoma cells. HPH1 and HPH2 have little sequence homology with each other, except in two highly conserved domains, designated homology domains I and II. They share these homology domains I and II with the Drosophila PcG protein Polyhomeotic (Ph), and we, therefore, have named the novel proteins HPH1 and HPH2. HPH1, HPH2, and BMI1 show distinct, although overlapping expression patterns in different tissues and cell lines. Two-hybrid analysis shows that homology domain II of HPH1 interacts with both homology domains I and II of HPH2. In contrast, homology domain I of HPH1 interacts only with homology domain II of HPH2, but not with homology domain I of HPH2. Furthermore, BMI1 does not interact with the individual homology domains. Instead, both intact homology domains I and II need to be present for interactions with BMI1. These data demonstrate the involvement of homology domains I and II in protein-protein interactions and indicate that HPH1 and HPH2 are able to heterodimerize.

Amino Acid Sequence↗
PubMed1996

Functional analysis of the interaction between the small GTP binding protein Cdc42 and the Ste20 protein kinase in yeast.

STE20 encodes a protein kinase related to mammalian p65Pak which functions in several signal transduction pathways in yeast, including those involved in pseudohyphal and invasive growth, as well as mating. In addition, Ste20 plays an essential role in cells lacking Cla4, a kinase with significant homology to Ste20. It is not clear how the activity of Ste20 is regulated in response to these different signals in vivo, but it has been demonstrated recently that binding of the small GTP binding protein Cdc42 is able to activate Ste20 in vitro. Here we show that Ste20 functionally interacts with Cdc42 in a GTP-dependent manner in vivo: Ste20 mutants that can no longer bind Cdc42 were unable to restore growth of ste20 cla4 mutant cells. They were also defective for pseudohyphal growth and agar invasion, and displayed reduced mating efficiency when mated with themselves. Surprisingly, however, the kinase activity of such Ste20 mutants was normal when assayed in vitro. Furthermore, these alleles were able to fully activate the MAP kinase pathway triggered by mating pheromones in vivo, suggesting that binding of Cdc42 and Ste20 was not required to activate Ste20. Wild-type Ste20 protein was visualized as a crescent at emerging buds during vegetative growth and at shmoo tips in cells arrested with alpha-factor. In contrast, a Ste20 mutant protein unable to bind Cdc42 was found diffusely throughout the cytoplasm, suggesting that Cdc42 is required to localize Ste20 properly in vivo.

Amino Acid Sequence↗
PubMed1995

Transformation of axial skeleton due to overexpression of bmi-1 in transgenic mice.

The oncogene bmi-1, which was originally found to be involved in B- and T-cell lymphoma formation encodes a protein with a domain of homology to the Drosophila protein Posterior sex combs (Psc) and its relative Suppressor 2 of Zeste (Su(z)2) (refs 4 and 5). Psc is a member of the Polycomb-group gene family, which is required to maintain the repression of homeotic genes that regulate the identities of Drosophila segments. The possibility that bmi-1 may play a similar role in vertebrates was suggested by our previous finding that mice lacking the bmi-1 gene show posterior transformations of the axial skeleton. Here we report that transgenic mice overexpressing Bmi-1 protein show the opposite phenotype, namely a dose-dependent anterior transformation of vertebral identity. The anterior expression boundary of the Hoxc-5 gene is shifted in the posterior direction, indicating that Bmi-1 is involved in the repression of Hox genes. We propose that Bmi-1 is a member of a vertebrate Polycomb complex that regulates segmental identity by repressing Hox genes throughout development.

Animals↗
PubMed1994

Posterior transformation, neurological abnormalities, and severe hematopoietic defects in mice with a targeted deletion of the bmi-1 proto-oncogene.

The bmi-1 proto-oncogene has been implicated in B-cell lymphomagenesis in E mu-myc transgenic mice. Distinct domains of the Bmi-1 protein are highly conserved within the Drosophila protein Posterior Sex Combs, a member of the Polycomb group involved in maintaining stable repression of homeotic genes during development. We have inactivated the bmi-1 gene in the germ line of mice by homologous recombination in ES cells. Null mutant mice display three phenotypic alterations: (1) a progressive decrease in the number of hematopoietic cells and an impaired proliferative response of these cells to mitogens; (2) neurological abnormalities manifested by an ataxic gait and sporadic seizures; and (3) posterior transformation, in most cases along the complete anteroposterior axis of the skeleton. The observations indicate that Mbi-1 plays an important role in morphogenesis during embryonic development and in hematopoiesis throughout pre- and postnatal life. Furthermore, these data provide the first evidence of functional conservation of a mammalian Polycomb group homolog.

Animals↗
PubMed1993

Characterization and chromosomal localization of the human proto-oncogene BMI-1.

The proto-oncogene bmi-1 is frequently activated by Moloney murine leukemia proviral insertions in E mu-myc transgenic mice1,2. Using a mouse bmi-1 cDNA probe a transcript of 3.3 kb was detected on Northern blots of human Burkitt's lymphoma cell lines. We have isolated and sequenced cDNA clones from a human erythroleukemia cell line (K562) derived cDNA library, using different mouse bmi-1 cDNA fragments as a probe. Analysis of genomic BMI-1 sequences reveals a gene structure which is very similar to that of the mouse, consisting of at least 10 exons. The human cDNA is 3203 bp in length and shows 86% identity to the mouse nucleotide sequence. The open reading frame encodes a protein of 326 amino acids which shares 98% identity to the amino acid sequence of mouse bmi-1 protein. In vitro translation experiments show that human cDNA derived RNA translates into a protein with a mobility of 44-46 kD on SDS polyacrylamide gels. Fluorescence in situ hybridization (FISH) on metaphase chromosome spreads located the human BMI-1 gene to the short arm of chromosome 10 (10p13), a region known to be involved in translocations in various leukemias.

Amino Acid Sequence↗
PubMed1991

Sequence similarity between the mammalian bmi-1 proto-oncogene and the Drosophila regulatory genes Psc and Su(z)2.

The bmi-1 proto-oncogene can be activated by Moloney murine leukaemia proviral insertions in E mu-myc transgenic mice. It encodes a highly conserved nuclear protein of 324 amino acids which belongs to a family of proteins containing a putative new zinc-finger. Another closely related member of this family is the mouse protein Mel-18. Here we report on the cloning and characterization of a homologous gene (D-bmi) from Drosophila melanogaster. Our analysis indicates that distinct domains of the mouse Bmi-1 protein, including the putative zinc-finger motif, are highly conserved within the much larger D-Bmi protein. Chromosomal localization and sequence comparison reveal that D-bmi is identical to Posterior Sex Combs (Psc) and indicate that the conserved domains between mouse bmi and Psc are also conserved within Suppressor-2 of Zeste (Su(z)2).

Amino Acid Sequence↗
PubMed1991

Identification of cooperating oncogenes in E mu-myc transgenic mice by provirus tagging.

Mo-MLV infection of E mu-myc transgenic mice results in a dramatic acceleration of pre-B cell lymphomagenesis. We have used provirus tagging to identify genes that cooperate with the E mu-myc transgene in B cell transformation. Here we report on the identification of four loci, pim-1, bmi-1, pal-1, and bla-1, which are occupied by proviruses in 35%, 35%, 28%, and 14% of the tumors, respectively. bmi-1, pal-1, and bla-1 represent novel common proviral insertion sites. The bmi-1 gene encodes a 324 amino acid protein with a predominantly nuclear localization. bmi-1 is highly conserved in evolution and contains several motifs frequently found in transcriptional regulators, including a new putative zinc finger motif. No genes have yet been assigned to pal-1 and bla-1. The distribution of proviruses over the four common insertion sites suggests that provirus tagging can be used not only to identify the cooperating oncogenes but also to assign these genes to distinct complementation groups in tumorigenesis.

Amino Acid Sequence↗
PubMed1991

Mice bearing the E mu-myc and E mu-pim-1 transgenes develop pre-B-cell leukemia prenatally.

Previously, it has been shown that E mu-pim-1 transgenic mice are predisposed to T-cell lymphomas, whereas E mu-myc transgenic mice are predisposed to pre-B-cell lymphomas. Here we show that double-transgenic E mu-myc E mu-pim-1 mice exhibit pre-B-cell leukemia in utero. Upon transplantation into recipient mice, embryo-derived double-transgenic leukemic cells frequently progressed to highly malignant monoclonal tumors, indicating that additional (epi)genetic events had occurred during the progression of the disease.

Animals↗
PubMed1989

Very high frequency of lymphoma induction by a chemical carcinogen in pim-1 transgenic mice.

Infection of mice with Moloney murine leukaemia virus (MuLV) induces T-cell lymphomas after an average latency period of 150 days. In these lymphomas the MuLV DNA is frequently integrated into the mouse chromosomal DNA in the vicinity of the pim-1 oncogene. Transgenic mice overexpressing the pim-1 oncogene are predisposed to develop T-cell lymphomas, but only to the extent that approximately 10% of the mice develop a lymphoma within 240 days. When these mice are infected with MuLV, lymphomas develop in all mice in only 50-60 days. In these lymphomas MuLV DNA is integrated near either the c-myc or N-myc gene, suggesting that pim-1 and myc synergize in lymphomagenesis. To determine whether this system has a more general application, we have now tested the susceptibility of pim-1 transgenic mice to N-ethyl-N-nitrosourea (ENU), a chemical carcinogen. With a single low dose of ENU, nearly all pim-1 transgenic mice, but only 15% of non-transgenic mice, develop T-cell lymphomas within 200 days. All ENU-induced lymphomas in both pim-1 transgenic and non-transgenic mice express high levels of c-myc messenger RNA, supporting the notion that pim-1 and c-myc synergize in lymphoma induction. We propose that pim-1 transgenic mice could be used to test the oncogenic potential of other chemical compounds.

Animals↗
PubMed1989

Predisposition to lymphomagenesis in pim-1 transgenic mice: cooperation with c-myc and N-myc in murine leukemia virus-induced tumors.

Transgenic mice bearing the pim-1 gene supplemented with an upstream immunoglobulin enhancer and a downstream murine leukemia virus long terminal repeat express pim-1 mRNA at high levels in both B and T cells. Between 5% and 10% of the pim-1 transgenic mice develop clonal T cell lymphomas before 7 months of age, whereas none of the age-matched control mice do, providing direct evidence for the oncogenic potential of pim-1. Histological examination and FACS analysis revealed no abnormalities in hematopoietic tissues of disease-free pim-1 transgenic mice. When newborn pim-1 transgenic mice are infected with MuLV, T cell lymphomas develop much faster (latency 7-8 weeks) than in nontransgenic mice (latency 22 weeks). In all these T cell lymphomas either c-myc or N-myc was activated by proviral insertion, suggesting strong cooperation between pim-1 and myc in lymphomagenesis.

Animals↗
PubMed1989

Transgenic mice as a means to study synergism between oncogenes.

Transgenic mice present a useful model to study the mechanisms underlying malignant transformation. (i) They can provide information on the oncogenic potential of genes as a function of tissue context. (ii) They allow the analysis of the primary effects of an oncogene on proliferation and differentiation before secondary mutations have occurred. (iii) Crossings between transgenic mice carrying different oncogenes can reveal their capacity to cooperate in transformation. (iv) Transgenic mice bearing a particular oncogene can be used to search for (new) (anti)oncogenes that synergize with the transgene. The non-acute transforming murine leukemia viruses (MuLV) appear very useful for this purpose. This has become clear from our studies with pim-I and c-myc transgenic mice. MuLV dramatically accelerates T-cell lymphomagenesis in transgenic mice overexpressing the pim-I oncogene in their lymphoid compartment. In all tumors induced by MuLV in pim-I transgenic mice, either the c-myc or the N-myc gene was activated by proviral insertion. Similarly, MuLV infection of transgenic mice overexpressing the c-myc gene in their B-cell compartment resulted in the acceleration of pre-B-cell lymphomagenesis. A significant fraction of the resulting pre-B-cell tumors showed proviral activation of pim-I. This shows that pim-I and myc synergize efficiently in both B- and T-cell lymphomagenesis. pim-I transgenic mice are also highly sensitive to tumor induction by N-ethyl-N-nitrosourea (ENU) and therefore represent an excellent in vivo model system to test the oncogenic potential of chemical compounds.

Animals↗
PubMed1989

N-myc is frequently activated by proviral insertion in MuLV-induced T cell lymphomas.

We report a new common proviral insertion site in murine leukemia virus-induced T cell lymphomas to be N-myc. Proviral activation of N-myc was found in 35% of independently induced primary tumors. The vast majority of the proviral insertions occur within a small segment of the 3'-untranslated region of the N-myc gene, directly downstream of the protein-encoding domain. This results in an increased level of expression of a truncated N-myc mRNA. Together with the previously shown c-myc activation we now find involvement of myc genes in greater than 75% of the primary T cell lymphomas induced by Moloney murine leukemia virus in C57BL10 and BALB/c mice, and show for the first time that N-myc can be over-expressed by a mechanism other than gene amplification.

Animals↗
PubMed1986

Construction of a retroviral cDNA version of the int-1 mammary oncogene and its expression in vitro.

The int-1 mammary oncogene is activated by proviruses of the Mouse Mammary Tumor Virus in many different mammary tumors. We have inserted a genomic fragment containing the protein-encoding domain of the gene into the retroviral shuttle vector pZIPneoSV(X)1. After one round of virus replication we recovered recombinant proviral DNA containing a correctly spliced copy of int-1. In vitro transcription of this cDNA version of int-1 using SP6 polymerase and translation in a reticulocyte lysate yielded a protein of approximately 37,000 daltons. High expression of int-1 in NIH-3T3 cells infected with recombinant virus did not lead to morphological transformation.

Animals↗