Evolutionary Biology has always fascinated us with fancy but effective theories on how ancient and extinct organisms might have looked like, along with their habitats, habits, and ways of life. A major limiting factor in these studies is the precise mimicry of the environment persisting in those times. We know that the environment plays a major role in shaping up the lives of all the organisms, which leads them to adopt various adaptation in order to cope up with the changing environment.Thus if we do not know the environment that caused a particular adaptation which, let’s say, was naturally selected and persisted to this day, we get to see half the picture and our understanding has little value.
Our traditional methods of studying past environments is completely based on archaeology and paleontology. These methods, however, are not competent enough to give us the true and complete picture. This is because both rely on preserved artifacts, which is biased towards the objects which are sturdier and less prone to degradation. Also, the interpretation is usually subjective and open-ended, and our modern eyes cannot accurately tell the usage of an object, without knowing the pre-existing environmental conditions. For example, the Great Bath structure found in the ruins of Indus Valley Civilisation might have been a storage tank of water for an entire city; or it could have been just a swimming pool. Thus we see that archaeology and paleontology cannot accurately depict the prevailing environmental conditions of the past and thus cannot aid in the study of evolution in the truest sense.
Progress in extracting and sequencing DNA from fossils has carved out a novel means to reconstruct the past. The study of the changes in the genetic makeup can provide us with the long-term changes in a population but cannot point out the environmental conditions persisting in those times. In contrast, epigenetic changes – defined as somatically heritable chemical changes of the DNA which do not alter its sequence, is quite responsive to environmental changes. There are a host of epigenetic ways of control which affect gene expression like methylation, histone modification, chromatin remodeling and nucleosome positioning. Of these, methylation is the most studied way of control, and we will focus on the same throughout this article. Methylation patterns are chiefly governed by two factors. Firstly, intrinsic genetic makeup, where the sequence of the DNA itself regulates the site of methylation, and secondly the changes in the environment. The first regulation is not very useful for assisting in the task at hand, but the second is. The environment here encompasses both external like temperature, humidity etc as well as internal like quantity of food intake, ingestion or inhalation of toxins etc. We can very easily see that the internal factors directly or indirectly depend on the external factors and are regulated by it.
Epigenetics and Ancient DNA:
Usually, epigenetics studies help us to decipher the changes in the epigenome upon environmental changes. With ancient DNA in hand, we are going to work backward. So a standard methylation map of the entire methylome is essential for us. The knowledge that methylations are usually done on the cytosine residue of a CpG island and the deamination of such methylated cytosine yields thymine, which is not removed by the DNA repair mechanism helps in the reconstruction of methylation maps. From the methylation map, genetically driven methylations are segregated from the methylations due to environmental changes, the latter was also known as Environmentally Responsive Loci (ERL). However, it must be noted that DNA can only be recovered from the persisting body parts of the organism, like the bones, teeth, hair of humans, and those may not necessarily respond to all environmental changes. For instance, in case of prolonged hunger and malnutrition, the DNA of adipose tissues might undergo methylations, but DNA from the bones are very unlikely to do so. Studies have shown that methylations and formation of ERLs also occur during the process of embryogenesis. Since embryonic cells divide and eventually form every other cell in the body, the epigenetic markers are also carried on and are found in every cell of the body in this case. A kind of such methylations in embryonic cells is known as Metastable Epialleles. These alleles are different in genetically similar people but are more or less similar in all the tissues of the body. Therefore, Metastable Epialleles are important in the study of past environments.
Analysis of Epigenetic Studies:
Effect of Nutrition:
Taking the nutritional aspect in focus, which is the widely studied factor, a study of the Dutch hunger winter of 1944-1945 yielded that 5’- UTR region of IGF2 gene was hypomethylated. Later it was found that several other genes like INSIGF, IL10, ABCA1, GNASAS, MEG3 and LEP genes have a change in their methylation levels following exposure to famine conditions. In contrast, studies have shown that a high-fat diet in mice corresponded with hypermethylation in about 30 CPGs in the intron 2 of igf2r. Also the availability of methyl donors has a limiting role in the methylation of gene sequences. When the diet of pregnant female mice was supplemented with excess folic acid, vitamin B12, choline and betaine – all of which are potent methyl donors, it resulted in hypermethylation of several loci in the offspring. Results such as these are used as standards in the nutritional assay of past humans, by comparing the methylome obtained from the DNA of archaic human fossils with the standard methylome.As an elaboration of how information such as above is used to reconstruct the past environments, we look into a study done on the rural Gambian population, where the amount of food intake is less in the rainy season, owing to scarcity of resources, which is termed as the ‘hunger season’; on the other hand the quantity of food intake increases in the dry season as harvesting is done and far greater resources are available. It was seen that babies conceived during the ‘hunger season’ had methylations in about 6 metastable epialleles. When compared to the Neanderthals and Denisovans, 3 of the above metastable epialleles feature in the DMRs, namely EXD3, RBM46, ZNF678. So we can fairly speculate that these archaic humans must have faced a similar ‘hunger season’ where there was an acute shortage of resources, which might have been periodical like the Gambians, or for short or a long span in their lives.
Effect of Pollutants and Toxins:
Pollutants and toxins also affect the methylome. Smoking, consumption of ethanol, exposure to arsenic, cadmium etc alters the methylome and leaves behind epigenetic markers. By assessing these alterations, we can decipher the trends of use of these drugs. The alterations due to arsenic, cadmium, if found in the methylomes of archaic humans, can reflect on the pollution levels of the soil, water etc persisting in those times.
Effect of Diseases:
Another factor which changes the methylome is diseases. It has been shown that diseases cause changes in the methylome and vice versa. Offsprings of mothers who have type 2 diabetes mellitus or gestational diabetes mellitus undergo hypomethylation in lipoprotein lipase gene(LPL), hypermethylation in the promoters of leptin gene and many other genes. These alterations result in decreased levels of leptin, which causes a person to eat more food, which is also accompanied by increased production of lipoprotein lipase, both of which ultimately make the offspring obese and likely to develop diabetes in the future.
This field of study, formally known as environmental paleoepigenetics, is at its infancy. So naturally, it has a lot of ground to cover. Here I list a few of the limitations:
- Firstly, DNA is a molecule which degrades over time in nature and the degradation is accelerated by heat, water, and sunlight. It has been found out that DNA has a specific life span, beyond which, no DNA can survive, even when kept in the ideal conditions possible.
- Mutations are incorporated in the DNA with time which changes the sequence of the DNA, along with the epigenome.The presence of the various mutagenic substance in the vicinity accelerates the process. Thus the epigenome we see may not be necessarily the exact premortem epigenome.
- The most popular method currently used to prepare methylation maps is Reduced Representation Bisulfite Sequencing (RRBS) which further degrades the DNA in the process of acquiring the methylation map and thus gives us information of about 10% of the Cp
- All the standard methylome of the modern man in different environmental conditions created to serve as a standard methylation map are created by using adult tissues other than the hard and usually persisting ones like teeth, bones, and hair which are usually found as fossils. So the relevance of this standard map in the study of recreating the past environments is still to be determined.
- A single ERL may respond to a number of environmental conditions as is seen with the genes NR3C1, SLC6A4 etc. Thus a complete knowledge of the full complement of ERLs that respond to each environmental factor is necessary for us to deduce the past environments.
Environmental paleoepigenetics is a multidisciplinary field of study which is primarily aimed at aiding the medical sector in the welfare of humans. Although this field is at its infancy and has quite a few limitations, some of which are listed above, it still has the potential of opening our eyes to the day to day environmental changes around archaic humans, which is considered impossible to this extent by the other methods present now. As the knowledge of how environment influences diseases are becoming more and more clear and complex than it was earlier thought of, knowing the effect of environment on the epigenome and ultimately on the human is crucial. Recent studies have shown that the chance a drug is potent, or ineffective, or toxic on a person depends on the person’s microbiome, which is different for each person, and it changes according to the lifestyle of the person, which is an amalgamation of the person’s internal and external environment. It is likely that the epigenome dictates the growth and the diversity of the microbiome in the human body. Thus it is very important for us to completely understand the interactions between the environment and the epigenome so that we can incorporate the knowledge and continue to make new age drugs and therapies to combat deadly diseases that haunt the human society. In this aspect, environmental paleoepigenetics does promise to be an exciting and an open field for scientists to unveil its riches and enrich the lives of the entire humanity.
By Suharto Banerjee
St. Xavier’s College
Dept. of Biotechnology
References: David Gokhman, Anat Malul, Liran Carmel; Inferring Past Environments from Ancient Epigenomes, Molecular Biology and Evolution, Volume 34, Issue 10, 1 October 2017, Pages 2429–2438, https://doi.org/10.1093/molbev/msx211
 Justeson JS. 1973. Limitations of archaeological inference: an information-theoretic approach with applications in methodology. Am Antiq. 38(02): 131
 Feil R, Fraga MF. 2012. Epigenetics and the environment: emerging patterns and implications. Nat Rev Genet. 13(2): 97–109
 Lienert F, Wirbelauer C, Som I, Dean A, Mohn F, Schu¨beler D. 2011. Identification of genetic elements that autonomously determine DNA methylation states. Nat Genet. 43(11): 1091–1097
 Briggs AW, Stenzel U, Meyer M, Krause J, Kircher M, P € a€ abo S. 2010. Removal of deaminated cytosines and detection of in vivo methylation in ancient DNA.Nucleic Acids Res. 6: e87
 Rakyan VK, Blewitt ME, Druker R, Preis JI, Whitelaw E. 2002. Metastable epialleles in mammals. Trends Genet. 18(7): 348–351
 Heijmans BT, Tobi EW, Stein AD, Putter H, Blauw GJ, Susser ES, Slagboom PE, Lumey LH. 2008. Persistent epigenetic differences associated with prenatal exposure to famine in humans. Proc Natl Acad Sci U S A. 105(44): 17046–17049
 TobiEW,LumeyLH,TalensRP,KremerD,PutterH,SteinAD,Slagboom PE, Heijmans BT. 2009. DNA methylation differences after exposure to prenatal famine are common and timing- and sex-specific. Hum Mol Genet. 18(21): 4046–4053
 Gallou-KabaniC,GaboryA,TostJ,KarimiM,MayeurS,LesageJ,Boudadi E, Gross MS, Taurelle J, Vige´ A, et al. 2010. Sex- and diet-specific changes of imprinted gene expression and dna methylation in mouse placenta under a high-fat diet.PLoS One 5(12): e14398
 Waterland RA, Dolinoy DC, Lin JR, Smith CA, Shi X, Tahiliani KG. 2006. Maternal methyl supplements increase offspring DNA methylation at Axin fused. Genesis 44(9): 401–406
 Waterland RA, Kellermayer R, Laritsky E, Rayco-Solon P, Harris RA, Travisano M, Zhang W, Torskaya MS, Zhang J, Shen L, et al. 2010. Season of conception in rural gambia affects DNA methylation at putative human metastable epialleles. PLoS Genet. 6(12): 1–10
 Gokhman D, Lavi E, Pru¨fer K, Fraga MF, Riancho JA, Kelso J, P€ a€ abo S, Meshorer E, Carmel L. 2014. Reconstructing the DNA methylation maps of the Neandertal and the Denisovan. Science 344(6183): 523–527
 Del Rosario MC, OssowskiV, KnowlerWC, Bogardus C, Baier LJ,Hanson RL.2014.Potentialepigeneticdysregulationofgenesassociatedwith MODY and type 2 diabetes in humans exposed to a diabetic intrauterineenvironment:ananalysisofgenome-wideDNAmethylation. Metab Clin Exp. 63: 654–660
 Houde AA, St-Pierre J, Hivert MF, Baillargeon JP, Perron P, Gaudet D, BrissonD,BouchardL.2014.PlacentallipoproteinlipaseDNAmethylation levels are associated with gestational diabetes mellitus and maternal and cord blood lipid profiles. J Dev Orig Health Dis. 5(2): 132–141
 LesseurC,ArmstrongDA,PaquetteAG,LiZ,PadburyJF,MarsitCJ.2014. Maternal obesity and gestational diabetes are associated with placental leptin DNA methylation. Am J Obstet Gynecol. 211(6): 654.e1–654.e9.
 Alexander Meissner, Andreas Gnirke, George W. Bell, Bernard Ramsahoye, Eric S. Lander, Rudolf Jaenisch; Reduced representation bisulfite sequencing for comparative high-resolution DNA methylation analysis, Nucleic Acids Research, Volume 33, Issue 18, 1 January 2005, Pages 5868–5877, https://doi.org/10.1093/nar/gki901