Epigenetic regulation profile in uninfected and TB-HIV infected Patients

Abstract

Devastatingly, both public and private health institutions in sub-Saharan Africa are burdened by the high prevalence of TB-HIV co-infections. Epigenetics involves changes in gene activity, occurring independently of alterations in the underlying DNA sequence. Epigenetic modifications, such as aberrant methylation, acetylation, phosphorylation, and ribosylation, are introduced by HIV and Mtb to evade host immunity, leading to microbial infection. This study aimed to detect the degree of epigenetic modifications in the TB-HIV co-infected cohort and the uninfected control subjects. METHOD: We obtained t issue samples from the lymph nodes of 45 TB-HIV co-infected patients and 45 TB-HIV uninfected individuals which were fixed in formalin and embedded in paraffin between 2017 and 2019 from the Charlotte Maxeke Johannesburg Academic Hospital, Republic of South Africa. TB-HIV co-infection was confirmed using conventional Polymerase chain reaction (PCR) targeting the Mtb rpoB and HIV gag-pol genes. To unravel the genetic identity of the HIV and Mtb strains in the TB-HIV cohort. Sanger sequencing and phylogenetic analysis were performed. Reverse Transcription quantitative PCR (RT-qPCR) was utilized to compare the degree of DNA methylation, histone acetylation, and histone methylation by analyzing the expression of corresponding target genes: DNA Methyltransferase 1(DNMT1), histone acetylation (HAT), Class II transactivator (CIITA), histone lysine methyl transferase (KMT), and SET domain bifurcated histone lysine methyltransferase2 (SETDB2). Subsequently, the H3 modification assay was used to detect and quantify patterns of histone methylation, acetylation and phosphorylation. Finally, the associated between age, gender and race in the observed epigenetic expression as well as H3 modification were assessed using statistical based methods. RESULTS: TB-HIV co-infection was confirmed among the TB-HIV tissue samples through the successful amplification and sequencing of the Mtb rpoB and HIV gag-pol genes, respectively. HIV-1 sequencing predominantly aligned with the intersubtype 07 BC strain, vii while the Mtb sequencing showed high identity with an Mtb strain with an accession number CP085590.1, revealing distinct phylogenetic clustering. Gene expression profiling via RT- qPCR demonstrated significant upregulation of the DNMT1, HAT, CIITA, KMT, and SETDB2 in the TB-HIV co-infected group. Histone modification analysis revealed widespread chromatin remodelling, including increased methylation (H3K4me1, H3K27me3), acetylation (H3K9ac, H3K27ac), and phosphorylation (H3S10ph, H3S28ph). Race and gender specific differences in gene expression and histone modification were observed, with race exerting a stronger influence on the expression of the CIITA and SETDB2 genes in the TB-HIV group. CONCLUSION: The disparities in the expression profiles of our candidate genes between the TB-HIV cohort and the non-TB-HIV cohort highlight the important role played by various TB and HIV strains in regulating the host gene expression landscape. Importantly, our findings reveal the potential of epigenetic modifications to be used as valuable biological markers that may, in the long run be applied for diagnostic procedures and for the synthesis of new and effective vaccines and therapeutics.