Abstract
BackgroundElucidating molecular mechanisms that are altered during HIV-1 infection may provide a better understanding of the HIV-1 life cycle and how it interacts with infected T-cells. One such mechanism is alternative splicing (AS), which has been studied for HIV-1 itself, but no systematic analysis has yet been performed on infected T-cells. We hypothesized that AS patterns in infected T-cells may illuminate the molecular mechanisms underlying HIV-1 infection and identify candidate molecular markers for specifically targeting infected T-cells.MethodsWe downloaded previously published raw RNA-seq data obtained from HIV-1 infected and non-infected T-cells. We estimated percent spliced in (PSI) levels for each AS exon, then identified differential AS events in the infected cells (FDR < 0.05, PSI difference > 0.1). We performed functional gene set enrichment analysis on the genes with differentially expressed AS exons to identify their functional roles. In addition, we used RT-PCR to validate differential alternative splicing events in cyclin T1 (CCNT1) as a case study.ResultsWe identified 427 candidate genes with differentially expressed AS exons in infected T-cells, including 20 genes related to cell surface, 35 to kinases, and 121 to immune-related genes. In addition, protein-protein interaction analysis identified six essential subnetworks related to the viral life cycle, including Transcriptional regulation by TP53, Class I MHC mediated antigen, G2/M transition, and late phase of HIV life cycle. CCNT1 exon 7 was more frequently skipped in infected T-cells, leading to loss of the key Cyclin_N motif and affecting HIV-1 transcriptional elongation.ConclusionsOur findings may provide new insight into systemic host AS regulation under HIV-1 infection and may provide useful initial candidates for the discovery of new markers for specifically targeting infected T-cells.
Highlights
Elucidating molecular mechanisms that are altered during Human immunodeficiency virus 1 (HIV-1) infection may provide a better understanding of the HIV-1 life cycle and how it interacts with infected T-cells
alternative splicing (AS) genes are enriched in pathways relevant to the HIV-1 life cycle To gain insight into the molecular functions of the 427 AS genes and their roles in the HIV-1 life cycle, we performed gene set enrichment analysis for canonical pathways and Gene Ontology (GO) terms using ConsensusPathDB v32 (CPDB)
These included pathways related to the late phase of HIV-1 infection and transcriptional reactivation of HIV, such as “Late Phase of HIV Life Cycle,” “HIV Transcription Initiation,” “Formation of HIV-1 Elongation Complex Containing HIV-1 Tat,” “Transcriptional Regulation by TP53,” “Clathrin Derived Vesicle Budding,” and “Membrane Trafficking.”
Summary
Elucidating molecular mechanisms that are altered during HIV-1 infection may provide a better understanding of the HIV-1 life cycle and how it interacts with infected T-cells. Current treatments for HIV patients (e.g. highly active antiretroviral therapy, HAART) mainly aim to effectively suppress viral load, recover immunologic function, and prevent HIV from acquiring drug-resistant mutations [1,2,3] Such treatments rely on preventing the infection from progressing to active AIDS, thereby improving survival. Genome-wide gene expression and DNA methylation profiling have successfully demonstrated changes in the molecular patterns of HIV-infected cells [9,10,11,12]. One such change occurs for nuclear factor IX (NFIX), which showed lower histone methylation in Tcells infected with HIV-1, leading to its. This increased NFIX may interact with the core-negative regulatory element (NRE) in the HIV-1 long terminal repeat (LTR), inhibiting the transcription of HIV-1
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