Host lncRNAs recognize invading bacteria and aid against infection by associating with polysomes

Knap, Primoz (2017) Host lncRNAs recognize invading bacteria and aid against infection by associating with polysomes. PhD thesis, University of Trento.

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Abstract

BACKGROUND: Infections mediated by pathogens, such as bacteria, often interfere with the host process of protein synthesis. This process is the most energy consuming in cells, that is why cells fine-tune it to conserve energy and respond quickly to stress. Thus, translation should be tightly regulated upon bacterial infection, yet the picture is still sketchy at best. Surprisingly, lncRNAs have recently been found to associate with ribosomes and polysomes; however, to date no research exists that addresses their role in translation regulation upon bacterial infection. AIM: The key question I wanted to answer during my PhD is whether host produced lncRNAs rewire the cell’s translation upon infection, inducing pathogen-specific and virulent factor-specific translational controls to cope with the infection. EXPERIMENTAL APPROACHES: To address the abovementioned aim, I used human colon epithelial cells (Caco-2) and Listeria monocytogenes as a host-pathogen model to explore the host cell’s response to infection at the translational level. By using a WT and a strain deficient for the expression of the main virulent factor Listeriolysin O (LLO-deficient (∆LLO) strain). Taking advantage of these strains, I explored whether the pore forming toxin, LLO, is able to trigger a host toxin-specific translational controls. To address this question, I massively employed polysome profiling, a classical approach to study translation and Next Generation Sequencing (NGS) to monitored changes in the transcriptome and the translatome upon infection at early time-points after infection and studying the possible function and mechanism of two lncRNAs that I found to be over-expressed upon infection. RESULTS: I showed that infection with either bacterial strain induced strong translational defects especially upon infection with WT Listeria. Cells responded to the infection by expressing numerous lncRNA and uploading them on polysomes. By comparing the transcriptome and the translatome of cells infected with either WT and ∆LLO I focused on two lincRNAs. The first, AC016831.1, is strongly upregulated upon infection with both strains of Listeria and exclusively associates with small active polysomes. In fact, my results show strong evidence that AC016831.1 is in fact actively associated with translating ribosomes and bioinformatics analyses of co-expressed genes showed its involvement in the innate immune response. The second, MIR181A1HG displayed a Listeria-specific and LLO-specific upregulation upon infection. I demonstrated that it is strongly associated with inactive stalled small polysomes. Importantly, its expression upon infection exerted a protective role against bacterial replication in host cells. Considering the obtained results, I propose that MIR181A1HG is acting as a ribosome sponge, decreasing the number of available ribosomes, ultimately leading to translation down-regulation. This role may help cells to keep the overall protein production rate at a low pace during infection, allowing the host to properly activate the innate immune system and fight-off the pathogen. In this research, using polysome profiling, I demonstrated for the first time that lncRNAs play a role in the host-pathogen crosstalk by rewiring translation. Evidence shows they might be even producing peptides, challenging their non-coding status and paving the way for understanding the possible role of short peptides in controlling bacterial infections.

Item Type:Doctoral Thesis (PhD)
Doctoral School:Biomolecular Sciences
PhD Cycle:29
Subjects:Area 05 - Scienze biologiche
Repository Staff approval on:20 Dec 2017 08:50

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