1 Legionella pneumophila is an opportunist pathogen that infects an immune-compromised host and causes 2 inflammatory pneumonia in the lungs termed as legionnaire’s disease. L. pneumophila infects alveolar 3 macrophages within the lungs and using a type IV secretion system, discharges an array of more than 300 4 proteins called effectors that interfere with the cell’s biochemical pathways to prepare a cozy niche for the 5 bacterial replication. Several recent studies also indicate that L. pneumophila infections in primary mouse 6 macrophages and protozoan cells manipulate host transcriptome and result in differential gene expression in the 7 infected host cell to facilitate their survival and evasion of the immune mechanisms. However, the mechanisms 8 involved in gene expression alterations are not clear and undefined. A comprehensive localization analysis of 9 Legionella effectors using GFP-tag revealed that a few of these Legionella effectors translocate into the nucleus 10 of the cell. An effector protein with an objective localization into the nucleus to modulate gene expression of the 11 host is called nucleomodulin. Recent studies indicate that several bacterial pathogens produce nucleomodulins 12 that aid in survival and pathogenesis through transient or long-term epigenetic modifications. In our prior work, 13 we identified five Legionella nucleomodulins with unknown function and the purpose of this study is to elucidate 14 the role of the Legionella nucleomodulins in understanding the pathogenesis of L. pneumophilia. 15 To better understand the effect of Legionella nucleomodulins on the host cells and determine their function 16 during the growth of Legionella pneumophila, we examined the differential gene expression pattern by 17 performing RNA sequence analysis on RNA extracted from mammalian cells expressing Legionella 18 nucleomodulin. The pathways and genes identified through functional analysis of RNA-seq data support 19 epigenetic modifications of host cells through methylation and histone modifications. The first objective of this 20 study is to examine the DNA methylation pattern on macrophages infected with L. pneumophila using Bisulfite 21 sequencing and methylation assays in comparison to uninfected macrophages. The second objective examines 22 interaction of Legionella nucleomodulin with histone and non-histone proteins by using chromatin 23 immunoprecipitation sequencing and co-immunoprecipitation assays. 24 The proposed research aims to detect transcriptional responses of a cell to its environmental influence such 25 as infections and understand how bacterial nucleomodulins mediate regulatory control through epigenetic 26 changes during an infection by intracellular pathogens to trigger altered gene expression patterns in the host cell 27 and manipulate cellular processes. As these mechanisms are ambiguous, this study will contribute to better 28 understanding of bacterial nucleomodulins and their role in bacterial ...