Replication of a positive-strand RNA virus involves an RNA-protein complex consisting of viral genomic RNA, host RNA(s), virus-encoded proteins, and host proteins. Dissecting out individual components of the replication complex will help decode the mechanism of viral replication. 5′ and 3′ UTRs in positive-strand RNA viruses play essential regulatory roles in virus replication.

Global dysregulation of RNA splicing and imbalanced sphingolipid metabolism has emerged as promoters of cancer cell transformation. Here, we present specific signature of alternative splicing (AS) events of sphingolipid genes for each breast cancer subtype from the TCGA-BRCA dataset. We show that ceramide synthase 2 (CERS2) undergoes a unique cassette exon event specifically in Luminal B subtype tumors. We validated this exon 8 skipping event in Luminal B cancer cells compared to normal epithelial cells, and in patient-derived tumor tissues compared to matched normal tissues. Differential AS-based survival analysis shows that this AS event of CERS2 is a poor prognostic factor for Luminal B patients. As Exon 8 corresponds to catalytic Lag1p domain, overexpression of AS transcript of CERS2 in Luminal B cancer cells leads to a reduction in the level of very-long-chain ceramides compared to overexpression of protein-coding (PC) transcript of CERS2. We further demonstrate that this AS event-mediated decrease of very-long-chain ceramides leads to enhanced cancer cell proliferation and migration. Therefore, our results show subtype-specific AS of sphingolipid genes as a regulatory mechanism that deregulates sphingolipids like ceramides in breast tumors, and can be explored further as a suitable therapeutic target.

Transcription factor p53 has been shown to aggregate into cytoplasmic/nuclear inclusions, compromising its native tumor suppressive functions. Recently, p53 is shown to form amyloids, which play a role in conferring cancerous properties to cells leading to tumorigenesis. However, the exact pathways involved in p53 amyloid-mediated cellular transformations are unknown. Here, using an in cellulo model of full-length p53 amyloid formation, we demonstrate the mechanism of loss of p53 tumor-suppressive function with concomitant oncogenic gain-of functions. Global gene expression profiling of cells suggests that p53 amyloid formation dysregulates the genes associated with cell cycle, proliferation, apoptosis, senescence along with major signaling pathways. This is further supported by the proteome analysis, showing a significant alteration in levels of p53 target proteins and enhanced metabolism, which enables the survival of cells. Our data indicate that specifically targeting the key molecules in pathways affected by p53 amyloid formation such as cyclin-dependent kinase-1, leads to loss of oncogenic phenotype and induces apoptosis of cells. Overall, our work establishes the mechanism of the transformation of cells due to p53 amyloids leading to cancer pathogenesis.

α-Synuclein is a natively unfolded protein and its deposition in the Lewy body and Lewy neurites in the substantia nigra region of the brain is linked to Parkinson's disease (PD). The molecular mechanisms of α-synuclein aggregation and its clearance have not been well understood. Until now, several strategies have been designed to inhibit α-synuclein aggregation and related cytotoxicity. Polyphenols, small molecules, synthetic peptides, and peptide-derived molecules have been considered as potential candidates that inhibit α-synuclein oligomerization and its fibrillation, and a few of them are in clinical trials. We have identified a polyphenolic compound ellagic acid (EA) that inhibits α-synuclein aggregation. Our results demonstrated that EA inhibits primary nucleation, seeded aggregation, and membrane-induced aggregation. The cytotoxicity of α-synuclein oligomers and fibers treated with EA has been investigated and we found that EA treated oligomers and fibrils showed reduced cytotoxicity. Additionally, we also observed inhibition of membrane binding of αsynuclein by EA in SH-SY5Y cells. In conclusion, the present study suggests that small molecules such as ellagic acid have antiamyloidogenic properties and may have therapeutic potential for Parkinson's disease and other proteinopathies.

Nutrient surplus and consequent free fatty acid accumulation in the liver cause hepatosteatosis. The exposure of free fatty acids to cultured hepatocyte and hepatocellular carcinoma cell lines induces cellular stress, organelle adaptation, and subsequent cell death. Despite many studies, the mechanism associated with lipotoxicity and subsequent cell death still remains poorly understood. Here, we have used the proteomics approach to circumvent the mechanism for lipotoxicity using hepatocellular carcinoma cells as a model. Our quantitative proteomics data revealed that ectopic lipids accumulation in cells severely affects the ubiquitin-proteasomal system. The palmitic acid (PA) partially lowered the expression of deubiquitinating enzyme USP7 which subsequently destabilizes p53 and promotes mitotic entry of cells. Our global phosphoproteomics analysis also provides strong evidence of an altered cell cycle checkpoint proteins’ expression that abrogates early G2/M checkpoints recovery with damaged DNA and induced mitotic catastrophe leading to hepatocyte death. We observe that palmitic acid prefers apoptosis-inducing factor (AIF) mediated cell death by depolarizing mitochondria and translocating AIF to the nucleus. In summary, the present study provides evidence of PA-induced hepatocellular death mediated by deubiquitinase USP7 downregulation and subsequent mitotic catastrophe.