The serum- and glucocorticoid-regulated kinase (SGK1) controls cell transformation and tumor progression. SGK1 affects mitotic stability by regulating the expression of RANBP1/RAN. Here, we demonstrate that SGK1 fluctuations indirectly modify the maturation of pre-miRNAs, by modulating the equilibrium of the RAN/RANBP1/RANGAP1 axis, the main regulator of nucleo-cytoplasmic transport. The levels of pre-miRNAs and mature miRNAs were assessed by qRT-PCR, in total extracts and after differential nuclear/cytoplasmic extraction. RANBP1 expression is the limiting step in the regulation of SGK1-SP1 dependent nuclear export. These results were validated in unrelated tumor models and primary human fibroblasts and corroborated in tumor-engrafted nude mice. The levels of pri-miRNAs, DROSHA, DICER and the compartmental distribution of XPO5 were documented. Experiments using RANGTP conformational antibodies confirmed that SGK1, through RANBP1, decreases the level of the GTP-bound state of RAN. This novel mechanism may play a role in the epigenomic regulation of cell physiology and fate.
Pancreatic ductal adenocarcinoma (PDAC) is traditionally associated with thrombocytosis/hypercoagulation and novel insights on platelet-PDAC “dangerous liaisons” are warranted. Here we performed an integrative omics study investigating the biological processes of mRNAs and expressed miRNAs, as well as proteins in PDAC blood platelets, using benign disease as a reference for inflammatory noise. Gene ontology mining revealed enrichment of RNA splicing, mRNA processing and translation initiation in miRNAs and proteins but depletion in RNA transcripts. Remarkably, correlation analyses revealed a negative regulation on SPARC transcription by isomiRs involved in cancer signaling, suggesting a specific ”education” in PDAC platelets. Platelets of benign patients were enriched for non-templated additions of G nucleotides (#ntaG) miRNAs, while PDAC presented length variation on 3′ (lv3p) as the most frequent modification on miRNAs. Additionally, we provided an actionable repertoire of PDAC and benign platelet-ome to be exploited for future studies. In conclusion, our data show that platelets change their biological repertoire in patients with PDAC, through dysregulation of miRNAs and splicing factors, supporting the presence of de novo protein machinery that can “educate” the platelet. These novel findings could be further exploited for innovative liquid biopsies platforms as well as possible therapeutic targets.
Purpose
Despite extensive biological and clinical studies, including comprehensive genomic and transcriptomic profiling efforts, pancreatic ductal adenocarcinoma (PDAC) remains a devastating disease, with a poor survival and limited therapeutic options. The goal of this study was to assess co-expressed PDAC proteins and their associations with biological pathways and clinical parameters.
Methods
Correlation network analysis is emerging as a powerful approach to infer tumor biology from omics data and to prioritize candidate genes as biomarkers or drug targets. In this study, we applied a weighted gene co-expression network analysis (WGCNA) to the proteome of 20 surgically resected PDAC specimens (PXD015744) and confirmed its clinical value in 82 independent primary cases.
Results
Using WGCNA, we obtained twelve co-expressed clusters with a distinct biology. Notably, we found that one module enriched for metabolic processes and epithelial-mesenchymal-transition (EMT) was significantly associated with overall survival (p = 0.01) and disease-free survival (p = 0.03). The prognostic value of three proteins (SPTBN1, KHSRP and PYGL) belonging to this module was confirmed using immunohistochemistry in a cohort of 82 independent resected patients. Risk score evaluation of the prognostic signature confirmed its association with overall survival in multivariate analyses. Finally, immunofluorescence analysis confirmed co-expression of SPTBN1 and KHSRP in Hs766t PDAC cells.
Conclusions
Our WGCNA analysis revealed a PDAC module enriched for metabolic and EMT-associated processes. In addition, we found that three of the proteins involved were associated with PDAC survival.
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