Dataset Information

Identification of pannexin 1-regulated genes, interactome, and pathways in rhabdomyosarcoma and its tumor inhibitory interaction with AHNAK.

ABSTRACT: Rhabdomyosarcoma (RMS), the most common soft tissue sarcoma in children, is an aggressive cancer with a poor prognosis. Despite current management, the 5-year survival rate for patients with metastatic RMS is ∼30%; underscoring the need to develop better treatment strategies. We have recently reported that pannexin 1 (PANX1) levels are downregulated in RMS and that restoring its expression inhibits RMS progression. Here, we have surveyed and characterized the molecular changes induced by PANX1 re-expression in RMS. We cataloged transcriptomic changes in this context by RNA sequencing. At the protein level, we unveiled PANX1 interactors using BioID, complemented by co-immunoprecipitation coupled to high-performance liquid chromatography/electrospray ionization tandem mass spectrometry performed in PANX1-enriched fractions. Using these data, we generated searchable public databases for the PANX1 interactome and changes to the RMS transcriptome occurring when PANX1 expression is restored. STRING network analyses revealed a PANX1 interactome involving plasma membrane and cytoskeleton-associated proteins including the previously undescribed interactor AHNAK. Indeed, AHNAK knockdown abrogated the PANX1-mediated reduction in RMS cell viability and migration. Using these unbiased approaches, we bring insight to the mechanisms by which PANX1 inhibits RMS progression, identifying the cell migration protein AHNAK as a key modifier of PANX1-mediated changes in RMS malignant properties.

SUBMITTER: Xiang X

PROVIDER: S-EPMC7946643 | biostudies-literature | 2021 Mar

REPOSITORIES: biostudies-literature

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Publications

Identification of pannexin 1-regulated genes, interactome, and pathways in rhabdomyosarcoma and its tumor inhibitory interaction with AHNAK.

Xiang Xiao X Langlois Stéphanie S St-Pierre Marie-Eve ME Blinder Anna A Charron Philippe P Graber Tyson E TE Fowler Stephanie L SL Baird Stephen D SD Bennett Steffany A L SAL Alain Tommy T Cowan Kyle N KN

Oncogene 20210209 10

Rhabdomyosarcoma (RMS), the most common soft tissue sarcoma in children, is an aggressive cancer with a poor prognosis. Despite current management, the 5-year survival rate for patients with metastatic RMS is ∼30%; underscoring the need to develop better treatment strategies. We have recently reported that pannexin 1 (PANX1) levels are downregulated in RMS and that restoring its expression inhibits RMS progression. Here, we have surveyed and characterized the molecular changes induced by PANX1 r ...[more]

PMID: 33564071

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Project description:We have recently reported the down-regulation of pannexin 1 (PANX1) levels in rhabdomyosarcoma (RMS) and that restoration of its expression inhibits the progression of embryonal (eRMS) and alveolar (aRMS) rhabdomyosarcoma in vitro and in vivo. While we have shown that this PANX1-mediated inhibition of RMS progression involves reduction of cell proliferation and migration, as well as induction of apoptosis, the underlining molecular mechanisms at play remained to be elucidated. In the present study, using RNA sequencing (RNA-seq) combined with DAVID analysis, we identified the PANX1 interactome in RMS cells and showed that the Gene Ontology (GO) terms such as apoptosis and migration, and KEGG pathways such as MAPK and Rap1 Signaling pathways were amongst the most highly enriched processes in Rh30 (aRMS) cells expressing ectopic PANX1. We then used a proximity-dependent labeling approach (BioID) to screen for the PANX1 interactors in RMS cells, which was complemented by co-immunoprecipitation (co-IP) coupled to high-performance liquid chromatography/electrospray ionization tandem mass spectrometry (HPLC-ESI-MS/MS) performed using Myc-PANX1-enriched subcellular fractions. STRING protein network analyses of the overlapping hits from these two approaches revealed a PANX1 interactome with plasma membrane and cytoskeleton associated proteins from which ACTB and AHNAK were further validated by co-IP using whole cell lysates followed by Western blotting analysis. Using this new and comprehensive information, we have generated the first online PANX1 transcriptome and interactome databases that allow searching, in a user-friendly manner, for PANX1-regulated genes and PANX1 interactors. Moreover, AHNAK knockdown in PANX1-expressing Rh18 (eRMS) and Rh30 (aRMS) significantly abrogated the PANX1-mediated reduction in cell viability and migration, as well as the PANX1-mediated sensitivity to anoikis. Using a combination of genome-wide unbiased approaches, this study is thus the first to identify the PANX1 transcriptome and interactome, providing the research community with online searchable databases to access the PANX1-regulated genes and PANX1 interacting partners. We have shown that PANX1 interacts with AHNAK and that this interaction is critical to the PANX1-mediated inhibition of RMS cell viability and migration, and induction of apoptosis; thus demonstrating how this new information can be used to identify key members of the PANX1 signaling pathway and their relevance to PANX1 function.

Dataset Information

Identification of pannexin 1-regulated genes, interactome, and pathways in rhabdomyosarcoma and its tumor inhibitory interaction with AHNAK.

Publications

Identification of pannexin 1-regulated genes, interactome, and pathways in rhabdomyosarcoma and its tumor inhibitory interaction with AHNAK.

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