Embryonic expression of the tumor-associated PAX3-FKHR fusion protein interferes with the developmental functions of Pax3 (original) (raw)
Related papers
Genes & Development, 2004
To investigate the role of the translocation-associated gene Pax3:Fkhr in alveolar rhabdomyosarcomas, we generated a Cre-mediated conditional knock-in of Pax3:Fkhr into the mouse Pax3 locus. Exploring embryonic tumor cell origins, we replaced a Pax3 allele with Pax3:Fkhr throughout its expression domain, causing dominant-negative effects on Pax3 and paradoxical activation of the Pax3 target gene, c-Met. Ectopic neuroprogenitor cell proliferation also occurs. In contrast, activation later in embryogenesis in cells that express Pax7 results in viable animals with a postnatal growth defect and a moderately decreased Pax7+ muscle satellite cell pool, phenocopying Pax7 deficiency but remarkably not leading to tumors.
The hybrid PAX3-FKHR fusion protein of alveolar rhabdomyosarcoma transforms fibroblasts in culture
Proceedings of the National Academy of Sciences, 1996
Pediatric alveolar rhabdomyosarcoma is characterized by a chromosomal translocation that fuses parts of the PAX3 and FKHIR genes. PAX3 codes for a transcriptional regulator that controls developmental programs, and FKHR codes for a forkhead-winged helix protein, also a likely transcription factor. The PAX3-FKHR fusion product retains the DNA binding domains of the PAX3 protein and the putative activator domain of the FKHR protein. The PAX3-FKHR protein has been shown to function as a transcriptional activator. Using the RCAS retroviral vector, we have introduced the PAX3-FKHR gene into chicken embryo fibroblasts. Expression of the PAX3-FKHR protein in these cells leads to transformation: the cells become enlarged, grow tightly packed and in multiple layers, and acquire the ability for anchorage-independent growth. This cellular transformation in vitro will facilitate studies on the mechanism of PAX3-FKHR-induced oncogenesis.
transcriptional activator than PAX3. rhabdomyosarcomas is a more potent t(2;13) translocation in alveolar The PAX3-FKHR fusion protein created by the http://mcb.asm.org/content/15/3/1522 Updated information and services can be found at: These include: CONTENT ALERTS more» cite this article), Receive: RSS Feeds, eTOCs, free email alerts (when new articles http://journals.asm.org/site/misc/reprints.xhtml Information about commercial reprint orders: http://journals.asm.org/site/subscriptions/ To subscribe to to another ASM Journal go to: on February 21, 2014 by guest Alveolar rhabdomyosarcomas are pediatric solid tumors with a hallmark cytogenetic abnormality: translocation of chromosomes 2 and 13 [t(2;13) (q35;q14)]. The genes on each chromosome involved in this translocation have been identified as the transcription factor-encoding genes PAX3 and FKHR. The NH 2 -terminal paired box and homeodomain DNA-binding domains of PAX3 are fused in frame to COOH-terminal regions of the chromosome 13-derived FKHR gene, a novel member of the forkhead DNA-binding domain family.
Genes, Chromosomes and Cancer, 2008
Rhabdomyosarcoma is a family of myogenic soft tissue tumors subdivided into two main subtypes: alveolar (ARMS) and embryonal (ERMS). ARMS is characterized by a frequent 2;13 chromosomal translocation that creates a PAX3-FKHR fusion transcription factor. To identify downstream targets of PAX3-FKHR, we introduced an inducible form of PAX3-FKHR into human RD ERMS cells. Microarray analysis identified 39 genes (29 upregulated and 10 downregulated) that are modulated by PAX3-FKHR in RD cells and differentially expressed between ERMS and PAX3-FKHR-positive ARMS tumors. Functional annotation demonstrated that genes involved in regulation of transcription and development, particularly neurogenesis, are represented in this group. MYCN was one notable neural-related transcription factor-encoding gene identified in this set, and its regulation by PAX3-FKHR was further confirmed at the RNA and protein levels. The findings of cycloheximide inhibition and time-course studies are consistent with the hypothesis that the PAX3-FKHR protein acts directly on the MYCN gene at the transcriptional level. Functional studies established that MYCN cooperates with PAX3-FKHR to enhance oncogenic activity. In conclusion, we identified a selected set of biologically relevant genes modulated by PAX3-FKHR, and demonstrated that PAX3-FKHR contributes to the expression of MYCN and in turn MYCN collaborates with PAX3-FKHR in tumorigenesis. V V C 2008 Wiley-Liss, Inc. studies suggest a gain of function with deregulated expression of downstream targets, which participate in pathways related to proliferation, survival, differentiation, and migration (Barr, 2001). A few genes have been defined as possible PAX3-FKHR targets, including MET, PDGFRA, BCL2L1, CXCR4, and TFAP2B (Epstein et al., 1998;
Identification of target genes of PAX3-FOXO1 in alveolar rhabdomyosarcoma
Oncology Reports, 2013
Background: The majority of alveolar rhabdomyosarcoma (ARMS) are distinguished through the paired box 3−forkhead box protein O1 (PAX3−FOXO1) fusion oncoprotein, being generated by a 2;13 chromosomal translocation. This fusion-positive ARMS is the most clinically difficult type of rhabdomyosarcoma. The present study characterized four genes [gremlin 1 (GREM1), deathassociated protein kinase-1 (DAPK1), myogenic differentiation-1 (MYOD1), and hairy/enhancer-of-split related with YRPW motif-1 (HEY1)] as targets of PAX3−FOXO1. Materials and Methods: The expression of the four genes, PAX3−FOXO1, and v-myc myelocytomatosis viral-related oncogene, neuroblastoma-derived (avian) (MYCN) was determined in various ARMS cell models and primary tumors. The roles of PAX3−FOXO1 and MYCN expression were examined. Results: Pulse-chase and cycloheximide experiments suggest that GREM1, DAPK1, and MYOD1 are directly regulated by PAX3−FOXO1. PAX3−FOXO1 appears to indirectly down-regulate HEY1 by up-regulating MYCN. Data reveal that the growthsuppressive activity of high PAX3−FOXO1 expression is closely-associated with up-regulation of the GREM1 and DAPK1 tumor-suppressor genes. Conclusion: This study characterized four downstream targets of PAX3−FOXO1 that contribute to the biological activities of growth suppression and myogenic differentiation. Rhabdomyosarcoma is the most prominent type of pediatric soft tissue sarcoma, being associated with the skeletal muscle lineage. Among pediatric rhabdomyosarcoma, there are two major subtypes based on their histological appearance: embryonal rhabdomyosarcoma (ERMS) and alveolar rhabdomyosarcoma (ARMS) (1). ARMS has a higher frequency of metastases at the initial diagnosis than ERMS, commonly conferring a poorer prognosis than ERMS (2, 3). A common characteristic of ERMS is a loss of heterozygosity at 11p15, however ERMS has not been reported to exhibit a diagnostic genetic alteration. In contrast, chromosomal translocation is frequently observed for ARMS (4, 5). The translocation t(2;13)(q35;q14) generating the paired box 3−forkhead box protein O1 (PAX3−FOXO1) gene fusion was found to occur in 55% of ARMS cases, while the translocation t(1;13)(q36;q14) generating the paired box 7−forkhead box protein O1 (PAX7−FOXO1) gene fusion occurred in 22% of cases, and 23% of ARMS were fusionnegative (2). The 2;13 translocation present in ARMS is characterized by an overexpression of PAX3−FOXO1 relative to wild-type PAX3 (6). Although PAX3−FOXO1-positive ARMS is the most clinically intractable fusion subtype of pediatric rhabdomyosarcoma, it can be difficult to histologically classify rhabdomyosarcoma into ARMS and ERMS in some cases (7), and no specific drugs are available for treating a specific fusion or histological subtype (1, 2, 7, 8). Therefore, there is a strong incentive to elucidate target genes of PAX3−FOXO1, which has the utility of being a therapeutic target and marker for the purpose of diagnosis and treatment of rhabdomyosarcoma. Several recent studies have utilized the gene expression profiles to classify rhabdomyosarcoma or identify target genes of only PAX3−FOXO1, or both PAX3−FOXO1 and PAX7−FOXO1, however, more research is needed to validate the biological function of the genes in ARMS development (9-15). Recently Ahn and co-authors identified 34 potential target genes and validated the four genes, gremlin-1 (GREM1), death-associated protein kinase-1 (DAPK1), myogenic differentiation (MYOD1), and hairy/enhancerof-split related with YRPW motif-1 (HEY1), as target genes of PAX3−FOXO1 in ARMS by analyzing gene 2029
Cancer research, 1998
The 2;13 chromosomal translocation in alveolar rhabdomyosarcoma generates the chimeric protein PAX3-FKHR, which is a powerful transcriptional activator. We hypothesize that PAX3-FKHR regulates downstream effector genes involved in rhabdomyosarcoma tumorigenesis. We evaluated alterations in expression of MET and neural cell adhesion molecule that were proposed previously as downstream targets of wild-type PAX3. We used a myogenic tumor cell culture system and rhabdomyosarcoma tumor specimens to assess candidate gene expression in relationship to various PAX3-FKHR expression levels. We demonstrate that the expression of MET, but not neural cell adhesion molecule, correlates significantly with PAX3-FKHR expression. These findings indicate that MET, which encodes a receptor involved in growth and motility signaling, is a downstream target of PAX3-FKHR in alveolar rhabdomyosarcoma.
A novel PAX3 rearrangement in embryonal rhabdomyosarcoma
Cancer Genetics and Cytogenetics, 2009
Rhabdomyosarcoma is the most common soft tissue tumor seen in children and young adults, and it can be classified into 2 major histological subtypes, alveolar and embryonal. In the alveolar subtype, 2 recurrent chromosomal translocations, t(2;13)(q35;q14) and its variant t(1;13)(p36;q14), have been identified as the specific cytogenetic abnormalities. These translocations produce the PAX3-FOXO1 and PAX7-FOXO1 fusion genes, respectively. In the embryonal subtype, however, no recurrent chromosomal abnormalities have been identified. In this study, we analyzed the complex chromosomal translocation in one case with embryonal rhabdomyosarcoma by means of spectral karyotyping (SKY) and identified a novel translocation involving chromosome band 2q35, which is the locus of PAX3 gene. Furthermore, we identified the novel PAX3 rearrangement using fluorescence in situ hybridization (FISH) analysis. Additional identification of the partner gene may help disclose the molecular mechanism of the development of this embryonal subtype.
Proceedings of the National Academy of Sciences, 1996
The t(2;13) translocation of alveolar rhabdomyosarcoma results in tumor-specific expression of a chimeric transcription factor containing the N-terminal DNA-binding domain of PAX3 and the C-terminal transactivation domain of FKHR. Here we have tested the hypothesis that PAX3-FKHR gains function relative to PAX3 as a consequence of switching PAX3 and FKHR transactivation domains, which were previously shown to have similar potency but distinct structural motifs. In transient cotransfection assays with human expression constructs, we have demonstrated the increased ability of PAX3-FKHR to activate transcription of a reporter gene located downstream of multimerized e5, PRS-9, or CD19 DNA-binding sites in three cell lines. For example, PAX3-FKHR was 100-fold more potent than PAX3 as an activator binding to e5 sites in NIH 3T3 cells. To compare transactivation potency independent of PAX3-specific DNA binding, we tested GAL4 fusions of full-length PAX3 and PAX3-FKHR or their respective C-terminal transactivation domains on a reporter with GAL4 DNA-binding sites. In this context, full-length PAX3-FKHR was also much more potent than PAX3. Additionally, the activity of each full-length protein was decreased relative to its C-terminal domain, demonstrating that N-terminal sequences are inhibitory. By deletion analysis, we mapped a bipartite cis-acting inhibitory domain to the same subregions within the DNA-binding domains of both PAX3 and PAX3-FKHR. We have shown, however, that the structurally distinct transactivation domains of PAX3 and PAX3-FKHR differ 10- to 100-fold in their susceptibility to inhibition, thus elucidating a mechanism by which PAX3 gains enhanced function during oncogenesis.