Whole Genome DNA Methylation Analysis of Active Pulmonary Tuberculosis Disease Identifies Novel Epigenotypes: PARP9/miR-505/RASGRP4/GNG12 Gene Methylation and Clinical Phenotypes

Abstract

We hypothesized that DNA methylation patterns may contribute to the development of active pulmonary tuberculosis (TB). Illumina's DNA methylation 450 K assay was used to identify differentially methylated loci (DML) in a discovery cohort of 12 active pulmonary TB patients and 6 healthy subjects (HS). DNA methylation levels were validated in an independent cohort of 64 TB patients and 24 HS. Microarray analysis identified 1028 DMLs in TB patients versus HS, and 3747 DMLs in TB patients after versus before anti-TB treatment, while autophagy was the most enriched signaling pathway. In the validation cohort, <i>PARP9</i> and <i>miR505</i> genes were hypomethylated in the TB patients versus HS, while <i>RASGRP4</i> and <i>GNG12</i> genes were hypermethylated, with the former two further hypomethylated in those with delayed sputum conversion, systemic symptoms, or far advanced lesions. <i>MRPS18B</i> and <i>RPTOR</i> genes were hypomethylated in TB patients with pleural involvement. <i>RASGRP4</i> gene hypermethylation and RPTOR gene down-regulation were associated with high mycobacterial burden. TB patients with <i>WIPI2</i>/<i>GNG12</i> hypermethylation or <i>MRPS18B</i>/<i>FOXO3</i> hypomethylation had lower one-year survival. In vitro ESAT6 and CFP10 stimuli of THP-1 cells resulted in DNA de-methylation changes of the <i>PARP9</i>, <i>RASGRP4</i>, <i>WIPI2</i>, and <i>FOXO3</i> genes. In conclusions, aberrant DNA methylation over the PARP9/miR505/RASGRP4/GNG12 genes may contribute to the development of active pulmonary TB disease and its clinical phenotypes, while aberrant DNA methylation over the <i>WIPI2</i>/<i>GNG12</i>/<i>MARPS18B</i>/<i>FOXO3</i> genes may constitute a determinant of long-term outcomes.