Dataset Information

Quantitative ubiquitylome analysis and crosstalk with proteome/acetylome analysis identified novel pathways and targets of perifosine treatment in neuroblastoma.

ABSTRACT: Background:Perifosine, is a third generation alkylphospholipid analog which has promising anti-tumor efficacy in clinical trials of refractory/recurrent neuroblastoma (NB). However, perifosine's mechanism of action remains unclear. Previously, we have shown that perifosine changes global proteome and acetylome profiles in NB. Methods:To obtain a more comprehensive understanding of the perifosine mechanism, we performed a quantitative assessment of the lysine ubiquitylome in SK-N-AS NB cells using SILAC labeling, affinity enrichment and high-resolution liquid chromatography combined with mass spectrometry analysis. To analyse the data of ubiquitylome, we performed enrichment analysis with gene ontology (GO), the Encyclopedia of Genes and Genomes (KEGG) pathway, ubiquitylated lysine motif, protein complex and protein domain. Protein-protein interaction was conducted to explore the crosstalk between ubiquitylome and previous global proteome/acetylome. Co-immunoprecipitation and western blotting were used to validate the results of the ubiquitylome analysis. Results:Altogether, 3,935 sites and 1,658 proteins were quantified. These quantified ubiquitylated proteins participated in various cellular processes such as binding, catalytic activity, biological regulation, metabolic process and signaling pathways involving non-homologous end-joining, steroid biosynthesis and Ras signaling pathway. Ubiquitylome and proteome presented negative connection. We identified 607 sites which were modified with both ubiquitination and acetylation. We selected 14 proteins carrying differentially quantified lysine ubiquitination and acetylation sites at the threshold of 1.5 folds as potential targets. These proteins were enriched in activities associated with ribosome, cell cycle and metabolism. Conclusions:Our study extends our understanding of the spectrum of novel targets that are differentially ubiquitinated after perifosine treatment of NB tumor cells.

SUBMITTER: Jiang M

PROVIDER: S-EPMC6370305 | biostudies-literature | 2018 Dec

REPOSITORIES: biostudies-literature

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Quantitative ubiquitylome analysis and crosstalk with proteome/acetylome analysis identified novel pathways and targets of perifosine treatment in neuroblastoma.

Jiang Min M Hua Zhongyan Z Dong Yudi Y Liu Zhihui Z Thiele Carol J CJ Li Zhijie Z

Translational cancer research 20181201 6

<h4>Background</h4>Perifosine, is a third generation alkylphospholipid analog which has promising anti-tumor efficacy in clinical trials of refractory/recurrent neuroblastoma (NB). However, perifosine's mechanism of action remains unclear. Previously, we have shown that perifosine changes global proteome and acetylome profiles in NB.<h4>Methods</h4>To obtain a more comprehensive understanding of the perifosine mechanism, we performed a quantitative assessment of the lysine ubiquitylome in SK-N-A ...[more]

PMID: 30761266

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Project description:Hepatitis B virus (HBV) infection remains a major health issue worldwide and the leading cause of cirrhosis and hepatocellular carcinoma (HCC). It has been reported previously that HBV invasion can extensively alter transcriptome, the proteome of exosomes and host cell lipid rafts. The impact of HBV on host proteins through regulating their global post-translational modifications (PTMs), however, is not well studied. Viruses have been reported to exploit cellular processes by enhancing or inhibiting the ubiquitination of specific substrates. Nevertheless, host cell physiology in terms of global proteome and ubiquitylome has not been addressed yet. Here by using HBV-integrated HepG2.2.15 model cell line we first report that HBV significantly modify the host global ubiquitylome. As currently the most widely used HBV cell culture model, HepG2.2.15 can be cultivated for multiple generations for protein labeling, and can replicate HBV, express HBV proteins and secrete complete HBV Dane particles, which makes it a suitable cell line for ubiquitylome analysis to study HBV replication, hepatocyte immune response and HBV-related HCC progression. Our previous experimental results showed that the total ubiquitination level of HepG2.2.15 cell line was significantly higher than that of the corresponding parental HepG2 cell line. By performing a Ubiscan quantification analysis based on stable isotope labeling of amino acids in cell culture (SILAC) of HepG2.2.15 and HepG2 cell lines, we identified a total of 7188 proteins and the protein levels of nearly 19% of them were changed over 2-folds. We further identified 3798 ubiquitinated Lys sites in 1476 host proteins with altered ubiquitination in response to HBV. Our results also showed that the global proteome and ubiquitylome were negatively correlated, indicating that ubiquitination might be involved in the degradation of host proteins upon HBV integration. We first demonstrated the ubiquitination change of VAMP3, VAMP8, DNAJB6, RAB8A, LYN, VDAC2, OTULIN, SLC1A4, SLC1A5, HGS and TOLLIP. In addition, we described 5 novel host factors SLC1A4, SLC1A5, EIF4A1, TOLLIP and BRCC36 that efficiently reduced the amounts of secreted HBsAg and HBeAg. Overall, the HBV-mediated host proteome and ubiquitylome change we reported will provide a valuable resource for further investigation of HBV pathogenesis and host-virus interaction networks.

Dataset Information

Quantitative ubiquitylome analysis and crosstalk with proteome/acetylome analysis identified novel pathways and targets of perifosine treatment in neuroblastoma.

Publications

Quantitative ubiquitylome analysis and crosstalk with proteome/acetylome analysis identified novel pathways and targets of perifosine treatment in neuroblastoma.

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