Preprint Article Version 1 This version is not peer-reviewed

MicroRNA-mRNA Co-sequencing Identifies Transcriptional and Post-transcriptional Regulatory Networks Underlying Muscle Wasting in Cancer Cachexia

Version 1 : Received: 30 August 2019 / Approved: 1 September 2019 / Online: 1 September 2019 (08:37:12 CEST)

How to cite: Javier Fernandez, G.; Henrique Ferreira, J.; José Vechetti-Júnior, I.; Nazario de Moraes, L.; Santiloni Cury, S.; Paccielli Freire, P.; Gutiérrez, J.; Ferretti, R.; Dal-Pai-Silva, M.; Regina Rogatto, S.; Francisco Carvalho, R. MicroRNA-mRNA Co-sequencing Identifies Transcriptional and Post-transcriptional Regulatory Networks Underlying Muscle Wasting in Cancer Cachexia. Preprints 2019, 2019090004 (doi: 10.20944/preprints201909.0004.v1). Javier Fernandez, G.; Henrique Ferreira, J.; José Vechetti-Júnior, I.; Nazario de Moraes, L.; Santiloni Cury, S.; Paccielli Freire, P.; Gutiérrez, J.; Ferretti, R.; Dal-Pai-Silva, M.; Regina Rogatto, S.; Francisco Carvalho, R. MicroRNA-mRNA Co-sequencing Identifies Transcriptional and Post-transcriptional Regulatory Networks Underlying Muscle Wasting in Cancer Cachexia. Preprints 2019, 2019090004 (doi: 10.20944/preprints201909.0004.v1).

Abstract

Cachexia is a complex metabolic syndrome characterized by loss of skeletal muscle, leading to a significant weight loss that impacts patient morbidity and mortality. Given the complexity of gene regulatory networks that control gene expression, our objective was to perform an integrative mRNA and miRNA profiling to identify genetic programs that capture essential mechanistic details that promote muscle atrophy in cancer cachexia. Here, we used RNA sequencing to analyze miRNAs and mRNAs expression profiles in tibialis anterior (TA) muscles of the Lewis lung carcinoma model of cancer cachexia. In addition, we compared these findings with RNA-Seq data from C2C12 myotubes treated in vitro with the cachectic factors tumor necrosis factor-alpha (TNF-α) and interferon-gamma (IFN-γ). Extracellular matrix (ECM) alterations were validated by picrosirius staining, western blot, and fractal dimension analyses. We found 1,008 mRNAs and 18 miRNAs differentially expressed in cachectic mice. This set of genes was associated with the ECM, proteolysis, and inflammatory response. Enrichment analysis of transcriptional factor binding sites revealed activation of the atrophy-related transcriptional factors: NF-κB, Stat3, AP-1, and FoxO. Furthermore, the integration of mRNA and miRNA expression profiles identified post-transcriptional regulation by miRNAs of genes involved in ECM organization, cell migration, transcription factors binding, ion transport, and FoxO signaling pathway. C2C12 myotubes treated with TNF-α and IFN-γ similarly down-regulate subsets of ECM genes, including collagens. Our integrative analysis of miRNA-mRNA co-profiles comprehensive characterized regulatory relationships of molecular pathways and revealed miRNAs targeting ECM-associated genes in cancer cachexia. We also confirmed in C2C12 myotubes that changes in ECM-associated genes are dependent on inflammatory signaling of the cytokines TNF-α and IFN-γ.

Subject Areas

Lewis lung cancer; miRNAs; transcription factors; extracellular matrix; cancer cachexia

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