Histone H3 lysine 27 acetylation profile undergoes two global shifts in undernourished children and suggests altered one-carbon metabolism

Abstract

Background: Stunting is a condition in which a child does not reach their full growth potential due to chronic undernutrition. It arises during the first 2 years of a child's life and is associated with developmental deficiencies and life-long health problems. Current interventions provide some benefit, but new approaches to prevention and treatment grounded in a molecular understanding of stunting are needed. Epigenetic analyses are critical as they can provide insight into how signals from a poor environment lead to changes in cell function.

Results: Here we profiled histone H3 acetylation on lysine 27 (H3K27ac) in peripheral blood mononuclear cells (PBMCs) of 18-week-old (n = 14) and 1-year-old children (n = 22) living in an urban slum in Dhaka, Bangladesh. We show that 18-week-old children destined to become stunted have elevated levels of H3K27ac overall, functional analysis of which indicates activation of the immune system and stress response pathways as a primary response to a poor environment with high pathogen load. Conversely, overt stunting at 1-year-of age is associated with globally reduced H3K27ac that is indicative of metabolic rewiring and downregulation of the immune system and DNA repair pathways that are likely secondary responses to chronic exposure to a poor environment with limited nutrients. Among processes altered in 1-year-old children, we identified one-carbon metabolism, the significance of which is supported by integrative analysis with results from histone H3 trimethylation on lysine 4 (H3K4me3). Together, these results suggest altered one-carbon metabolism in this population of stunted children.

Conclusions: The epigenomes of stunted children undergo two global changes in H3K27ac within their first year of life, which are associated with probable initial hyperactive immune responses followed by reduced metabolic capacity. Limitation of one-carbon metabolites may play a key role in the development of stunting. Trial registration ClinicalTrials.gov NCT01375647. Registered 17 June 2011, retrospectively registered, https://ichgcp.net/clinical-trials-registry/NCT01375647 .

Keywords: Epigenetics; Histone acetylation; One-carbon metabolism; Stunting; Undernutrition.

Conflict of interest statement

The authors declare no competing interests.

© 2021. The Author(s).

Figures

Fig. 1

Vicious cycle of chronic undernutrition. Multiple factors contribute to chronic undernutrition in children. Enteric infections (with or without diarrhea) are key determinants of a child’s nutritional status, as they often lead to gut dysfunction with accompanying health issues listed in the figure. An optimal time for intervention is from conception until 2 years of age, a crucial time in child development [2]. A stunted child faces a high risk of life-long health consequences, which may be passed on to the next generation through poorly understood mechanisms. Health problems associated with the introduction of high calorie diet are marked with an asterisk. T2D type 2 diabetes, CVD cardiovascular diseases

Fig. 2

The H3K27ac landscape undergoes global changes in stunted children within their first year of life. a Overview of the experimental design. Each line in the plot with HAZ scores represents the change of HAZ score from birth to 1 year of age for a child whose PBMC sample was used for the study. The left panel shows HAZ scores for 18-week-old children (n = 14), the right panel for 1-year-old children (n = 22). Red dots indicate HAZ <  − 2 at a given age. Lines are colored based on ΔHAZ between birth and 18 weeks or between birth and 1 year. Bottom panel illustrates three different scenarios for differential analysis in which blue regions are downregulated in stunting, grey regions unchanged or with no significant trend; and red regions are upregulated in stunted children. b MA-plot shows changes in H3K27ac regions in 18-week-old children as the ΔHAZ (18 wk) score increases. Each dot is an H3K27ac region, the x-axis represents the mean read coverage over the region, and the y-axis indicates the log2(fold-change) of read coverage per unit increase of ΔHAZ (18 wk) score. Colored dots indicate significantly affected regions with false discovery rate (FDR) corrected p value < 0.05. c Same as b for 1-year-old children. d Alluvial plot showing changes to log2(fold-change) of corresponding regions between 18 weeks and 1 year. “Up”/“down”—H3K27ac levels are increased/decreased, accordingly, with higher risk of stunting at a given age, “not-present”—region was not identified at the given age

Fig. 3

Functional annotation of genes associated with significantly upregulated H3K27ac regions in 18-week-old stunted children. a Genome browser snapshot of H3K27ac signal tracks showing a representative region in which the normalized coverage increased with decreasing ΔHAZ (18 wk) score, i.e. H3K27ac was higher in stunted children. The color of individual signal tracks corresponds to the ΔHAZ (18 wk) scores. Genes associated with the highlighted region are labeled. b Biological terms (databases: GO biological process, KEGG, Rectome, WikiPathways) that were significantly enriched for genes associated with differential H3K27ac regions in 18-week-old children. Each node is a biological term, the node size corresponds to the term size, the node color indicates the significance of enrichment (bright orange: low q-value, light orange: higher q-value, all q-values < 0.05), and the edges are based on the number of genes shared between terms. Groups of terms were manually organized in different colored clusters with the summary term for the cluster shown in the label with the same color. c Gene network constructed with STRING (see “Methods” section) using genes found in the highlighted biological terms in b. Red genes belong to the “viral processes” GO term

Fig. 4

Functional annotation of genes associated with significantly downregulated H3K27ac regions in 1-year-old stunted children. a Genome browser screenshot showing a representative region in 1-year-old children in which read coverage decreased with decreasing ΔHAZ (1 year) score, i.e. H3K27ac was lower in stunted children. Track colors correspond to the child’s ΔHAZ (1 year) score. Genes associated with the region are labeled. b Network of biological terms as in Fig. 3b applied on data from 1-year-old children. c Genes from highlighted biological terms in b are shown in a pathway diagram. PPP pentose phosphate pathway, SSP serine synthesis pathway

Fig. 5

Integrative analysis of H3K27ac and H3K4me3 in 1-year-old stunted children. a Overview of the “gene-centric” approach in which each differential H3K27ac and H3K4me3 region was assigned to its target gene(s). Only genes predicted to be regulated by both H3K27ac and H3K4me3 were functionally annotated. b, d Venn diagrams show overlap of genes regulated by H3K27ac regions that were downregulated in stunting (red circle), and H3K4me3 regions that were downregulated (blue circle) and upregulated (grey circle) in stunting respectively. c, e Networks of biological terms as in Fig. 3b generated for overlapping genes from b, and d respectively. The color code of network edges corresponds to the color code of H3K4me3 circles in the associated Venn diagrams. f Overview of the “region-centric” approach. Significantly affected H3K27ac regions that overlap significantly affected H3K4me3 regions were assigned to target genes, which were functionally annotated. g, h Venn diagrams showing intersections of significantly downregulated H3K27ac regions (red circle) in stunting with significantly downregulated (blue circle) or upregulated (grey circle) H3K4me3 regions in stunting. i Pathway enrichment of genes associated with overlapping regions from g, h. Bars are color-coded accordingly with H3K4me3 circles in Venn diagrams

Fig. 6

Working model depicting changes to the epigenetic landscape in stunting. The model highlights changes to H3K27ac and H3K4me3 landscapes in 18-week-old and 1-year-old children as they become more severely stunted, along with biological terms associated with these changes