Dataset Information

Isoform-resolved Correlation Analysis Between mRNA Abundance Dynamics and Protein Level Degradation

ABSTRACT: We employed an optimized, sensitive DIA-MS method on a high-resolution Orbitrap platform and re-measured the proteomic samples used in a previous study, in which the heterogeneity of HeLa cells across different research labs was analyzed by a multi-omic approach. Using the same MS sample sets of all HeLa strains we achieved higher sensitivity compared to our previous SWATH-MS results that were acquired on a TripleTOF instrument (5600 model). To benefit from the significantly improved sensitivity of this single-shot DIA-MS for both proteomic and protein turnover analysis, we analyzed the samples originating from six HeLa Kyoto strains and six HeLa CCL2 strains to profile both total protein-level abundances and the proxy protein turnover rates (Kloss, by pulsed SILAC labeling) across cell lines. Our results emphasized the importance of relative mRNA-protein turnover rate correlation analysis. The dataset demonstrate that specific biological processes, cellular organelles, subunits of organelles, and individual protein isoforms may have distinctive degradation rate and the corresponding buffering or concerting protein turnover control across cancer cell lines.

INSTRUMENT(S): Orbitrap Fusion Lumos

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Epithelial Cell, Cell Culture

DISEASE(S): Cervix Carcinoma

SUBMITTER: Barbora Salovska

LAB HEAD: Yansheng Liu

PROVIDER: PXD014847 | Pride | 2020-03-11

REPOSITORIES: Pride

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Isoform-resolved correlation analysis between mRNA abundance regulation and protein level degradation.

Salovska Barbora B Zhu Hongwen H Gandhi Tejas T Frank Max M Li Wenxue W Rosenberger George G Wu Chongde C Germain Pierre-Luc PL Zhou Hu H Hodny Zdenek Z Reiter Lukas L Liu Yansheng Y

Molecular systems biology 20200301 3

Profiling of biological relationships between different molecular layers dissects regulatory mechanisms that ultimately determine cellular function. To thoroughly assess the role of protein post-translational turnover, we devised a strategy combining pulse stable isotope-labeled amino acids in cells (pSILAC), data-independent acquisition mass spectrometry (DIA-MS), and a novel data analysis framework that resolves protein degradation rate on the level of mRNA alternative splicing isoforms and is ...[more]

PMID: 32175694

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Project description:Background: Skeletal muscle crucially depends on motor innervation, and, when damaged, on the resident muscle stem cells (MuSCs). However, the role and function of MuSCs in the context of denervation remains poorly understood. Methods: Alterations of MuSCs and their myofiber niche after denervation were investigated in a surgery-based mouse model of unilateral sciatic nerve transection. FACS-isolated MuSCs were subjected to RNA-sequencing and mass spectrometry for the analysis of intrinsic changes after denervation and in vivo assays, such as Cardiotoxin-induced muscle injury or MuSC transplantation, were performed to assess MuSC functions after denervation. Bioinformatic and histological analyses were conducted to further examine MuSCs and their myofiber niche after denervation. Results: Muscle cross section analysis revealed a significant increase in Pax7 (p-value= 0.0441), Pax7/Ki67 (p-value= 0.0023), MyoD (p-value= 0.0016) and Myog (p-value= 0.0057) positive cells after denervation, illustrating a break of quiescence and commitment to the myogenic lineage. An Omics approach showed profound intrinsic alterations on the mRNA (2613 differentially expressed genes, p-value <0.05) and protein (1096 differentially abundant proteins, q-value <0.05) level of MuSCs 21 days after denervation. Skeletal muscle injury together with denervation surgery caused deregulated regeneration, indicated by the reduced number of proliferating MuSCs and sustained high levels of developmental myosin heavy chain (Sham: 1 % vs DEN: 40 % of all myofibers), at 21 days post-surgery. In a transplantation assay, MuSCs from a denervated host were still able to engraft and fuse to form new myofibers, irrespective of the innervation status of the recipient muscle. Analysis of myofibers revealed not only massive changes in the expression profile (10492 differentially expressed genes, p-value <0.05) after denervation, but it was also shown that secretion of Opn and Tgfb1 from denervated myofibers was increased 30-fold and 6000-fold, respectively. Bioinformatic analyses indicated strong upregulation of gene expression of the transcription factor Junb in MuSCs from denervated muscles (log2 fold change = 3.27). Of interest, Tgfb1 recombinant protein was able to induce Junb gene expression in vitro, demonstrating that myofiber-secreted ligands can induce gene expression changes in MuSCs, which might result in the phenotypes observed after denervation. Conclusion: Skeletal muscle denervation is altering myofiber secretion, causing MuSC activation and profound intrinsic changes, leading to reduced regenerative capacity. As MuSCs possess a remarkable regenerative potential, they might represent a promising target for novel treatment options for neuromuscular disorders and peripheral nerve injuries.

Dataset Information

Isoform-resolved Correlation Analysis Between mRNA Abundance Dynamics and Protein Level Degradation

Publications

Isoform-resolved correlation analysis between mRNA abundance regulation and protein level degradation.

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