Our studies have also uncovered novel surrogate function of an epigenetic effector molecule in the host defense against pathogenic bacteria. A homolog of the Drosophila protein, Suppressor of Variegation 3(9), SUV39H1, is an evolutionary conserved protein. In all the previous studies that have examined suppressor of variegation 3(9) or its homologs, this protein has been shown to function by modulating chromatin organization, through its role as a histone H3K9 methyltransferase in the nucleus. We show that during infection, SUV39H1 is relocalised to the cell surface where it interacts with the mycobacterial bacilli and methylates the mycobacterial cell surface protein HupB, a histone like protein from mycobacteria. SUV39H1 mediated HupB methylation inhibits the adhesion capabilities of the mycobacterial bacilli and interfere with their ability to interact with the host cell. This surrogate role of SUV39H1 as a methyltransferase of the mycobacterial protein HupB, adds a novel epigenetic paradigm to the duel between the host cell and the infecting pathogen.

These studies opened new avenues to better understand host–pathogen interactions and to develop novel, more effective antimicrobial strategies.have

Chromatin organisation and genomic imprinting

The fusion of two haploid cells (a sperm and an egg) to produce the diploid zygote heralds the development of a new organism. According to Mendel's laws of inheritance the genetic information contributed by the sperm (from the father) and the egg (from the mother) is equal. This would in effect mean that given the right conditions, fusion of two sperm nuclei or two egg nuclei should lead to normal development of an organism. However, this was found not to be the case in mammals, some insects and plants. Mouse embryos with two paternal copies (androgenetic) or two maternal copies (parthenogenetic) of the genome die very early during embryonic development (Barton et al 1984 Role of paternal and maternal genomes in mouse development Nature 311(5984):374-6; Surani et al 1984 Development of reconstituted mouse eggs suggests imprinting of the genome during gametogenesis. Nature 308(5959):548-50; McGrath and Solter 1984 Completion of mouse embryogenesis requires both the maternal and paternal genomes. Cell 37(1):179-83). Therefore, in mammals, maternal and paternal contribution to the progeny is not equal and for normal development contribution from both parents is required. This non-equivalence of the parental genomes is epigenetic in nature and forms the basis of the phenomenon known as genomic imprinting.

For imprinted genes one copy of a DNA molecule is transcribed and the other copy is silent. What distinguishes two molecules of the same DNA sequence within the same nucleus? The answer lies not in the genetic information encoded by the DNA sequence but in its epigenetic modifications. These modifications include differential modification of the DNA itself and differential organisation of DNA within the chromatin scaffold. One important modification of DNA known in mammals is methylation of cytosine residues in the CpG dinucleotides. However, methylation does not answer all mechanistic questions related to imprinted genes. If one allele of a gene becomes methylated what prevents methylation of the other allele? Does the unmethylated allele carry another epigenetic imprint? To investigate the mechanisms of genomic imprinting, it would therefore be essential to identify the alternative imprint on the unmethylated allele.

It has been found that within the differentially methylated regions of some imprinted genes there are sub-regions that also show differential organisation of chromatin. Interestingly, the regions which show differential chromatin organisation have also been genetically defined as Imprinting Control Regions (ICRs) and shown to control the imprinting status of genes within that locus. In all the imprinted loci analysed, specialised chromatin structures were found exclusively on the unmethylated allele. This suggests that for these regions, methylation and specialised chromatin structures are mutually exclusive and reflects alternate epigenetic states. How is this mutual exclusiveness between methylation and specialised chromatin structures maintained? Do the chromatin modifications on unmethylated allele prevent it from getting methylated? More importantly, during development when are these chromatin structures established? Can such chromatin conformations behave as imprints marking the parental alleles? The aim of the ongoing project is to answer these questions and identify the role of chromatin organisation in mechanisms underlying genomic imprinting in particular and as a epigenetic regulator of gene expression in general (Feil and Khosla 1999, John et al 2001, Sowpati et al 2008, Thamban et al 2019a).

Epigenetic Inheritance

The inheritance of epigenetic modifications across multiple generations results from events during germ cell development and fertilization that allow epigenetic marks acquired during the life time of an organism to be passed on to the progeny. Epigenetic inheritance could be intergenerational as has been documented for genomic imprinting or transgenerational (see figure on transgenerational epigenetic inheritance) where epigenetic modification acquired in one generation is inherited unchanged across several generations. Work from our laboratory has shown the role of DNMT3L in epigenetic inheritance.

Figure Legend: Mechanisms of Transgenerational Inheritance. A hypothetical model describing the possible mechanisms by which epimutations acquired by the somatic cells are transmitted to the germ cells and inherited to the next generation (ref: Thamban et al 2019b)

DNMT3L is an epigenetic modifier that interacts with de novo methyltransferases DNMT3A/3B and histone H3. Work from our laboratory has indicated a role of DNMT3L in nuclear reprogramming. Nuclear reprogramming observed in both DNMT3L expressing Drosophila and mammalian cells overexpressing DNMT3L was gradual and progressive. In Drosophila, ectopic DNMT3L expression caused progressive misregulation of genes and by 5th generation a very large number of genes were aberrantly expressed. Some of the transgenic flies expressing DNMT3L developed melanotic tumors and did not survive. The progressive misregulation of genes across several generations in transgenic Drosophila involved germline passage through meiotic divisions. Most epigenetic marks are normally erased and reset in the germline. Therefore, any epimutation acquired during one generation would be lost in the germ cells and hence not inherited. Even for imprinted genes, the epigenetic imprints inherited from the previous generation are erased before new epigenetic modifications determined by the sex of the individual are initiated. However, we observed a progressive decrease in the genome-wide levels of H3K4me3 and H3K36me3 across more than one generation. This accumulated decrease in these modifications (or accumulation of epimutations) suggested that in the presence of DNMT3L new epigenetic modifications were being added over and above of what was inherited from the previous generation. It is possible that epimutation accumulation could be incapacitation of protein(s) responsible for erasure or resetting of epigenetic modification, as a secondary event in Drosophila expressing DNMT3L. Hence, in each generation DNMT3L would keep adding new epimutations to the ones inherited from the previous generation. On the other hand, it is possible that the presence of DNMT3L prevented erasure of epimutations inherited from the previous generation. In this scenario, epigenetic modifiers responsible for setting up of new epigenetic marks in the germ cells would add on to the epigenetic modifications that were not erased due to the presence of DNMT3L. Further work is being carried out to examine these hypotheses (Basu et al Scientific Reports 2016, Thamban et al 2019b)