Oncogene database

4094

MAF

v-maf avian musculoaponeurotic fibrosarcoma oncogene homolog;MAF;v-maf avian musculoaponeurotic fibrosarcoma oncogene homolog

Avian musculoaponeurotic fibrosarcoma (MAF) protooncogene|T lymphocyte c-maf long form|c-maf proto-oncogene|proto-oncogene c-Maf|transcription factor Maf

16q22-q23

protein-coding

Maf oncoprotein is a basic-leucine zipper (bZip) type of transcriptional activator. Since many transcription factors are known to form functional complexes, we searched for proteins that interact with the DNA-binding domain of Maf using the phage display method and identified two homeodomain-containing proteins, Hoxd12 and MHox/Prx1/Phox1/Pmx1. Studies with mutants of Hox and Maf proteins showed that they associate through their DNA-binding domains; the homeodomain of Hox and the bZip domain of Maf, respectively. Reflecting the high similarity of the bZip domain, ALL other Maf family members tested (c-/v-Maf, MafB, MafK, MafF, and MafG) also associated with the Hox proteins. Pax6, whose homeodomain is relatively similar to MHox, also could interact with Maf. However, two other bZip oncoproteins, Fos and Jun, failed to associate with the Hox proteins, while a distantly related Hox family member, Meis1, could not interact with Maf. Through interactions with the bZip domain, the Hox proteins inhibited the DNA binding activity of Maf, whereas the binding of Hox proteins to their recognition sequences was not abrogated by Maf. We further showed that coexpression of the Hox proteins repressed transcriptional activation and transforming activity of Maf. These results suggested that the interaction of a set of Hox proteins with Maf family members may interfere not only with their oncogenicity but also with their physiological roles.

Mechanisms underlying the pathophysiology of minimal change nephrotic syndrome (MCNS), the most frequent of glomerular diseases in children, remain elusive, although recent arguments suggest that T cell dysfunction may be involved in the pathogenesis of this disease. Recently, we reported that activated T cells of these patients display a down-regulation of IL-12R beta2 chain, suggesting an early commitment toward Th2 phenotype. In this study, we show that the short form of the proto-oncogene c-maf, a known activator of the IL-4 gene, is highly induced in MCNS T cells during relapse, where it translocates to the nuclear compartment and binds to the DNA responsive element. Unexpectedly, the nuclear localization of c-maf did not promote the IL-4 gene transcription in relapse. Using several approaches, we show in this study that RelA blunts IL-4 induction in T cells during the relapse in these patients. We demonstrate that the ex vivo inhibition of proteasome activity in T cells from relapse, which blocks NF-kappaB activity, strongly increases the IL-4 mRNA levels. Overexpression of c-maf in T cells induces a high level of IL-4 promoter-driven luciferase activity. In contrast, coexpression of c-maf with NF-kappaB RelA/p50, or RelA, but not p50, inhibits the c-maf-dependent IL-4 promoter activity. Finally, we demonstrated that, in T cell overexpressing RelA and c-maf, RelA expelled c-maf from its DNA binding site on IL-4 gene promoter, which results in active inhibition of IL-4 gene transcription. Altogether, these results suggest that the involvement of c-maf in Th2 commitment in MCNS operates through IL-4-independent mechanisms.

Gene expression profiling studies reported by Hurt et al. in this issue of cancer Cell reveal an unexpectedly frequent overexpression of c-maf in multiple myeloma and identify molecular targets of c-maf transactivation. The results define c-maf as a prototype of a class of oncogenes that not only stimulate cell cycle progression, but also promote pathological interactions between tumor and stroma cells.

The oncogene c-maf is translocated in approximately 5%-10% of multiple myelomas. Unexpectedly, we observed c-maf expression in myeloma cell lines lacking c-maf translocations and in 50% of multiple myeloma bone marrow samples. By gene expression profiling, we identified three c-maf target genes: cyclin D2, integrin beta7, and CCR1. c-maf transactivated the cyclin D2 promoter and enhanced myeloma proliferation, whereas dominant inhibition of c-maf blocked tumor formation in immunodeficient mice. c-maf-driven expression of integrin beta7 enhanced myeloma adhesion to bone marrow stroma and increased production of VEGF. We propose that c-maf transforms plasma cells by stimulating cell cycle progression and by altering bone marrow stromal interactions. The frequent overexpression of c-maf in myeloma makes it an attractive target for therapeutic intervention.

A novel avian transforming retrovirus was isolated from a chicken musculoaponeurotic fibrosarcoma. This virus (called AS42) induces tumors histopathologically indistinguishable from the original sarcoma after a long latent period when inoculated into newborn chickens. AS42 also exhibits a weak transforming activity when infected into chicken embryo fibroblasts (CEF). This virus is replication-defective and associated with a helper virus of subgroup A (called ASAV). An AS42-specific protein of about 100 kDa was immunoprecipitated from lysates of AS42-transformed CEF with antiserum directed against avian retrovirus virion proteins. Molecular analysis of the genomic structure of the AS42 virus has revealed that this 100-kDa protein represents a novel oncogene, v-maf of cellular origin, which is fused with a part of the viral gag gene (Nishizawa et al., Proc. Natl. Acad. Sci. USA 86, 7711-7715, 1989). Interestingly, some size variation was observed among the gag-maf fusion proteins found in individual clones of transformed CEF. Consistent with this observation, Southern blot analyses and nucleotide sequence determination of several independent isolates of proviral DNA indicated that this virus segregates multiple forms of deletion mutants, probably through homologous recombinations among the repetitive sequences present within the v-maf coding region.

Single bp mutations in the RET proto-oncogene can cause multiple endocrine neoplasia type 2 syndromes. The conventional approach for genotyping RET mutations is sequencing the exons. A closed-tube RET genotyping assay using a saturating DNA dye, unlabeled probes, and amplicon high-resolution melting analysis was developed. The method required two sequential polymerase chain reaction stages, a primary and secondary assay. The primary assay analyzed RET exons 10, 11, 13, 14, and 16 with a total of seven reactions using eight unlabeled probes. The primary assay genotyped wild-type exons, a common exon 13 polymorphism, and an exon 16 mutation, whereas other RET sequence variation was detected. The primary unlabeled probe data limited the possible genotypes for the detected RET sequence variation, which permitted genotyping in a secondary assay with only two to five reactions. Six probes were designed with the masking technique and masked selected sequence variations to allow unambiguous analysis of other mutations elsewhere under the probe. After this two-stage RET genotyping assay, less than 0.2% of exons tested would require sequencing for genotype. A blinded study generated from five wild type and 29 available RET sequence variation samples was 100% concordant with sequencing. Amplicon high-resolution melting analysis with unlabeled probes and the masking technique is a fast, accurate method for genotyping the >50 RET sequence variations.

Angiogenic growth factors induce the transcription of the cell surface peptidase CD13/APN in activated endothelial cells of the tumor vasculature. Inhibition of CD13/APN abrogates endothelial invasion and morphogenesis in vitro and tumor growth in vivo suggesting a critical functional role for CD13 in angiogenesis. Experiments to identify the transcription factors responsible for this regulation demonstrated that exogenous expression of the proto-oncogene c-Maf, but not other bZip family members tested, potently activates transcription from a critical regulatory region of the CD13 proximal promoter between -115 and -70 bp which is highly conserved among mammalian species. Using promoter mutation, EMSA and ChIP analyses we established that both endogenous and recombinant c-Maf directly interact with an atypical Maf response element contained within this active promoter region via its basic DNA/leucine zipper domain. However full activity of c-Maf requires the amino-terminal transactivation domain, and site-directed mutation of putative phosphorylation sites within the transactivation domain (serines 15 and 70) shows that these sites behave in a dramatic cell type-specific manner. Therefore, this atypical response element predicts a broader range of c-Maf target genes than previously appreciated and thus impacts its regulation of multiple myeloma as well as endothelial cell function and angiogenesis.

The tick-borne protozoan parasite Theileria annulata causes a debilitating disease of cattle called Tropical Theileriosis. The parasite predominantly invades bovine macrophages (m phi) and induces host cell transformation by a mechanism that has not been fully elucidated. Infection is associated with loss of characteristic m phi functions and phenotypic markers, indicative of host cell de-differentiation. We have investigated the effect of T. annulata infection on the expression of the m phi differentiation marker c-maf. The up-regulation of c-maf mRNA levels observed during bovine monocyte differentiation to m phi was suppressed by T. annulata infection. Furthermore, mRNA levels for c-maf and the closely related transcription factor mafB were significantly lower in established T. annulata-infected cell-lines than in bovine monocyte-derived m phi. Treatment of T. annulata-infected cells with the theileriacidal drug buparvaquone induced up-regulation of c-maf and mafB, which correlated with altered expression of down-stream target genes, e.g. up-regulation of integrin B7 and down-regulation of IL12A. Furthermore, T. annulata infection is associated with the suppression of the transcription factors, Pu.1 and RUNX1, and colony stimulating factor 1 receptor (CSF1R) which are also involved in the regulation of monocyte/m phi differentiation. We believe these results provide the first direct evidence that T. annulata modulates the host m phi differentiation state, which may diminish the defence capabilities of the infected cell and/or promote cell proliferation. Musculoaponeurotic fibrosarcoma oncogene (MAF) transcription factors play an important role in cell proliferation, differentiation and survival; therefore, regulation of these genes may be a major mechanism employed by T. annulata to survive within the infected m phi.

Modulating aberrant transcription of oncogenes is a relatively unexplored opportunity in cancer therapeutics. In approximately 10% of multiple myelomas, the initiating oncogenic event is translocation of musculoaponeurotic fibrosarcoma oncogene homolog (MAF), a transcriptional activator of key target genes, including cyclinD2. Our prior work showed that MAF is up-regulated in an additional 30% of multiple myeloma cases. The present study describes a common mechanism inducing MAF transcription in both instances. The second mode of MAF transcription occurred in myelomas with multiple myeloma SET domain (MMSET) translocation. MMSET knockdown decreased MAF transcription and cell viability. A small-molecule screen found an inhibitor of mitogen-activated protein kinase kinase (MEK), which activates extracellular signal-regulated kinase (ERK)-MAP kinases, reduced MAF mRNA in cells representing MMSET or MAF subgroups. ERK activates transcription of FOS, part of the AP-1 transcription factor. By chromatin immunoprecipitation, FOS bound the MAF promoter, and MEK inhibition decreased this interaction. MEK inhibition selectively induced apoptosis in MAF-expressing myelomas, and FOS inactivation was similarly toxic. Reexpression of MAF rescued cells from death induced by MMSET depletion, MEK inhibition, or FOS inactivation. The data presented herein demonstrate that the MEK-ERK pathway regulates MAF transcription, providing molecular rationale for clinical evaluation of MEK inhibitors in MAF-expressing myeloma.

Protein methylation pathways comprise methionine adenosyltransferase (MAT), which produces S-adenosylmethionine (SAM) and SAM-dependent substrate-specific methyltransferases. However, the function of MAT in the nucleus is largely unknown. MafK represses or activates expression of heme oxygenase-1 (HO-1) gene, depending on its heterodimer partners. Proteomics analysis of MafK revealed its interaction with MATIIalpha, a MAT isozyme. MATIIalpha was localized in nuclei and found to form a dense network with chromatin-related proteins including Swi/Snf and NuRD complexes. MATIIalpha was recruited to Maf recognition element (MARE) at HO-1 gene. When MATIIalpha was knocked down in murine hepatoma cell line, expression of HO-1 was derepressed at both basal and induced levels. The catalytic activity of MATIIalpha, as well as its interacting factors such as MATIIbeta, BAF53a, CHD4, and PARP1, was required for HO-1 repression. MATII serves as a transcriptional corepressor of MafK by interacting with chromatin regulators and supplying SAM for methyltransferases.

Most colorectal cancers (CRCs) containing activated BRAF (BRAF[V600E]) have a CpG island methylator phenotype (CIMP) characterized by aberrant hypermethylation of many genes, including the mismatch repair gene MLH1. MLH1 silencing results in microsatellite instability and a hypermutable phenotype. Through an RNAi screen, here we identify the transcriptional repressor MAFG as the pivotal factor required for MLH1 silencing and CIMP in CRCs containing BRAF(V600E). In BRAF-positive human CRC cell lines and tumors, MAFG is bound at the promoters of MLH1 and other CIMP genes, and recruits a corepressor complex that includes its heterodimeric partner BACH1, the chromatin remodeling factor CHD8, and the DNA methyltransferase DNMT3B, resulting in hypermethylation and transcriptional silencing. BRAF(V600E) increases BRAF/MEK/ERK signaling resulting in phosphorylation and elevated levels of MAFG, which drives DNA binding. Analysis of transcriptionally silenced CIMP genes in KRAS-positive CRCs indicates that different oncoproteins direct the assembly of distinct repressor complexes on common promoters.

BACKGROUND: There are currently no biomarkers for early breast cancer patient populations at risk of bone metastasis. Identification of mediators of bone metastasis could be of clinical interest. METHODS: A de novo unbiased screening approach based on selection of highly bone metastatic breast cancer cells in vivo was used to determine copy number aberrations (CNAs) associated with bone metastasis. The CNAs associated with bone metastasis were examined in independent primary breast cancer datasets with annotated clinical follow-up. The MAF gene encoded within the CNA associated with bone metastasis was subjected to gain and loss of function validation in breast cancer cells (MCF7, T47D, ZR-75, and 4T1), its downstream mechanism validated, and tested in clinical samples. A multivariable Cox cause-specific hazard model with competing events (death) was used to test the association between 16q23 or MAF and bone metastasis. ALL statistical tests were two-sided. RESULTS: 16q23 gain CNA encoding the transcription factor MAF mediates breast cancer bone metastasis through the control of PTHrP. 16q23 gain (hazard ratio (HR) for bone metastasis = 14.5, 95% confidence interval (CI) = 6.4 to 32.9, P < .001) as well as MAF overexpression (HR for bone metastasis = 2.5, 95% CI = 1.7 to 3.8, P < .001) in primary breast tumors were specifically associated with risk of metastasis to bone but not to other organs. CONCLUSIONS: These results suggest that MAF is a mediator of breast cancer bone metastasis. 16q23 gain or MAF protein overexpression in tumors may help to select patients at risk of bone relapse.

L7 is expressed in ALL adult cerebellar Purkinje cells, although during development it appears in a stereotyped spatial and temporal pattern that is manifested as parasagittal domains of neurons. mutations of the L7 promoter in transgenic mice have established that these domains represent functional compartments of Purkinje neurons. Therefore, it is hoped that by defining the transcriptional control of the L7 gene insights into the mechanisms that control functional fate and organization in the nervous system can be gained. Fragments of the L7 promoter were introduced into a selectable reporter gene in Saccharomyces cerevisiae, and these strains were used to select for cerebellar cDNAs encoding proteins that can bind to, and activate transcription from, these elements. This assay identified the c-Maf proto-oncogene as activating transcription from two sites in the L7 promoter. We did a functional domain analysis of vertebrate c-Maf based upon transcriptional activation in S. cerevisiae and showed the requirement for a transactivation domain, leucine zipper, and DNA-binding region in c-Maf. The c-Maf interaction site was mapped to the sequence G/TGG/CNG/TNCT CAGNN in the L7 promoter, which represents an atypical 12-O-tetradecanoate-13-acetate-responsive element-type Maf-responsive element. However, neither Fos nor Jun, either alone or in combination with each other or c-Maf, altered transcription from this element. In contrast, a Maf-related protein, Nrl, completely mimicked c-Maf actions. These data suggest that Maf may interact with additional basic-zipper proteins that determine a subtype of Maf-responsive element binding.

The ras oncogene family has been implicated in tumor resistance to ionizing radiotherapy. Using the gene-transfer model, we show here that ras expression may also affect cell responses to chemical inducers of oxidative stress. Studies involving human osteosarcoma subclones, which vary in their levels of EJras expression, revealed a tight correlation between the amounts of ras-encoded mRNA and p21 produced, and the degree of resistance to doxorubicin or hydrogen peroxide. Differences in response could not be explained by increased activity of anti-oxidant enzymes such as superoxide dismutase, glutathione reductase, glutathione S-transferase or glutathione peroxidase. Moreover, there were no significant differences in glutathione levels. Although the resistant cells had elevated levels of gamma-glutamyl-transferase mRNA indicative of an increased rate of glutathione turnover, this elevation was not specific for ras-transfected cell lines. Lovastatin, an inhibitor of protein isoprenylation critical for p21ras membrane association and function, restored the sensitivity of ras-transformed cells to doxorubicin and hydrogen peroxide. The data indicate that pharmacological agents affecting ras expression may enhance responses of some human tumors to free-radical-mediated chemotherapies.

Erythroid transcription factor NF-E2 is a tissue-restricted heterodimeric protein which recognizes an extended AP-1 motif [(T/C)TGCTGA(C/G)TCA(T/C)] found in the upstream locus control regions of the alpha- and beta-globin gene clusters. A cDNA clone encoding a cell-type-specific subunit of NF-E2, designated p45 NF-E2, has previously been characterized and shown to encode a basic-leucine zipper DNA-binding protein. Here we describe protein purification and cloning of cDNA that encodes the second basic-leucine zipper subunit of the native NF-E2 heterodimer. This polypeptide, designated p18, is widely expressed. It displays extensive homology to the v-maf oncogene product and a human retinal-specific protein, NRL. Unusual features in the basic region shared by v-Maf, NRL, and p18 place them in a distinct subfamily of AP-1-like proteins.

The v-maf oncogene of the avian musculoaponeurotic fibrosarcoma virus, AS42, encodes a nuclear protein which contains a characteristic b-Zip domain. By screening a chicken embryo fibroblast (CEF) cDNA library under moderately stringent hybridization conditions, we picked up a series of cDNA clones for a novel maf-related gene which we named mafK. We also identified another maf-related gene named mafF by screening a chicken genomic library using a mafK probe. Structural analyses suggested that the mafK and mafF genes consist of three exons. The exon-intron structures of the two genes resemble each other, but differ from that of the chicken c-maf gene. As compared to the c-Maf protein, the proteins encoded by the mafK and the mafF genes are rather small in size and lack the regions corresponding to the amino terminal acidic domain present in the c-Maf protein. On the other hand, the structures of the b-Zip domain are well conserved among these Maf-related proteins. When overexpressed by using an avian retroviral vector, the two maf-related genes did not induce morphological transformation of CEF cells but induced colony formation in soft agar with very low efficiencies. With a specific antibody, the MafK protein was detected predominantly in the nuclei of the cells infected with the virus which carries the mafK gene. Tissue distributions of these three maf-family genes are different from one another, probably reflecting their different functions in vivo.

The v-maf oncogene, identified as the transforming gene of the avian retrovirus AS42, encodes a protein containing a b-Zip motif. From this structural feature, the v-Maf protein was expected to form a dimer and function as a nuclear DNA-binding protein. In this study, we demonstrate that this protein indeed localizes predominantly in the nucleus and forms a homodimer through its leucine zipper structure. To delineate the structural requirement for the transforming activity, we constructed and characterized a panel of v-maf mutants harboring various deletions or point mutations. A region of about 100 amino acid residues located near its carboxyl terminus, which contains the b-Zip motif, was found to be essential for the basal transforming activity of v-Maf on chicken embryo fibroblasts. On the other hand, the amino-terminal two-thirds of the v-Maf protein seems to play a role in potentiating the transforming activity of v-Maf. It was also found that the c-maf proto-oncogene, without any structural modification in its protein-coding region, could transform cells as efficiently as could the v-maf oncogene when transduced by a retroviral vector. Thus, it is probably deregulated expression that makes the v-maf gene oncogenic. In addition, we discovered one point mutation, altering the structure of the b-Zip domain, which further enhances the transforming activity of the v-maf oncogene. Such mutant will be useful in exploring the mechanism of action of the Maf protein.

The v-maf oncogene encodes a nuclear bZip protein which specifically recognizes relatively long palindromic sequences related to an AP-1 site. In this study, we investigated the relationship of transactivation and transformation activity of Maf. The amino-terminal two thirds of the molecule were dispensable for its DNA-binding activity but conferred its transactivation potential. Transactivation activities of a set of deletion mutants correlated well with their cell transforming abilities. However, a point mutant associated with enhanced oncogenic activity was not more effective in transactivation than the wild type, suggesting that some other function(s) of Maf is also important for its transforming ability. We also examined the effect of other bZip proteins on the transactivation activity of Maf. Three small Maf family proteins (MafK, MafF and MafG), which are missing the transactivation domain of v-Maf, competitively inhibited transactivation by Maf. Co-expression of Jun or Fos also affected the transactivation potential of Maf by forming Maf/Jun or Maf/Fos heterodimers of distinct DNA-binding specificities. In addition to these factors, we noticed the presence of a strong endogenous transactivating activity associated with a sequence related to an NF-E2 site rather than the typical AP-1 site in fibroblast cells. These results indicate that AP-1 site-like cis-regulatory elements of eukaryotic genes are regulated by multiple sets of bZip dimers with different DNA-binding and transactivation properties.

The molecular basis for the distinctive cytokine expression of CD4+ T helper 1 (Th1) and T helper 2 (Th2) subsets remains elusive. Here, we report that the proto-oncogene c-maf, a basic region/leucine zipper transcription factor, controls tissue-specific expression of IL-4. c-Maf is expressed in Th2 but not Th1 clones and is induced during normal precursor cell differentiation along a Th2 but not Th1 lineage. c-Maf binds to a c-Maf response element (MARE) in the proximal IL-4 promoter adjacent to a site footprinted by extracts from Th2 but not Th1 clones. Ectopic expression of c-Maf transactivates the IL-4 promoter in Th1 cells, B cells, and nonlymphoid cells, a function that maps to the MARE and Th2-specific footprint. Furthermore, c-Maf acts in synergy with the nuclear factor of activated T cells (NF-ATp) to initiate endogeneous IL-4 production by B cells. Manipulation of c-Maf may alter Th subset ratios in human disease.

To assess the normal function of the c-maf protooncogene product, we examined its tissue distribution during fetal growth and organogenesis of the chicken using an immunohistochemical technique. Nuclei of neuronal cells in the optic lobe and in the outer granular layer of the cerebral cortex were efficiently stained by anti-Maf serum throughout ALL developmental stages examined. In the lung, mesenchymal cells were stained intensely by the antiserum, with the most intense staining around embryonic day 15. During the embryonic period, nuclei of mesenchymal cells in the perichondrial and periosteal tissues as well as intestinal submucosa were also stained specifically by the antiserum. In addition, we detected substantial Maf immunoreactivity in nuclei of renal glomerular and proximal-tubular epithelial cells. These results suggest that c-Maf protein plays some important roles in the development of the central nervous system and tissues of mesodermal origin such as connective and renal tissues.

Dysregulation of oncogenes by translocation to an IgH (14q32) or IgL (kappa, 2p11 or lambda, 22q11) locus is a frequent event in the pathogenesis of B-cell tumors. Translocations involving an IgH locus and a diverse but nonrandom array of chromosomal loci occur in most multiple myeloma (MM) tumors even though the translocations often are not detected by conventional cytogenetic analysis. In a continuing analysis of translocations in 21 MM lines, we show that the novel, karyotypically silent t(14;16)(q32.3;q23) translocation is present in 5 MM lines, with cloned breakpoints from 4 lines dispersed over an approximately 500-kb region centromeric to the c-maf proto-oncogene at 16q23. Another line has a t(16;22)(q23;q11), with the breakpoint telomeric to c-maf, so that the translocation breakpoints in these 6 lines bracket c-maf. Only these 6 lines overexpress c-maf mRNA. As predicted for dysregulation of c-maf by translocation, there is selective expression of one c-maf allele in 2 informative lines with translocations. This is the first human tumor in which the basic zipper c-maf transcription factor is shown to function as an oncogene.

Transcription factors of the Maf proto-oncogene family have been shown to participate in the regulation of several differentiation specific genes. We previously reported that a member(s) of this family is involved in the regulation of the neuroretina specific gene, QR1, through a promoter region, designated the A box, that is closely related to the Maf recognition element (MARE). We undertook an identification of Maf family genes expressed in the quail neuroretina (QNR) and we report the isolation of mafA, a gene encoding a novel member of the large Maf proteins subgroup. expression of this gene is developmentally regulated in the neuroretina. MafA is able to bind to MARE sequence and to heterodimerize with v-Maf, MafB, Jun and Fos, but not with the small MafF and MafK proteins. Accordingly, it is able to transactivate the QR1 promoter A box. We also show that increased expression of mafA induces sustained proliferation of postmitotic QNR cells.