National Eye Institute (NEI) researchers have mapped the organization of human retinal cell chromatin, the fibers that package three billion nucleotide-long DNA molecules into compact structures that fit into chromosomes within each cell’s nucleus. The resulting comprehensive gene regulatory network offers insights into the regulation of gene expression in general, and in retinal function, in both rare and common eye diseases.
“This is the first detailed integration of retinal regulatory genome topology with genetic variants associated with age-related macular degeneration (AMD) and glaucoma, two leading causes of vision loss and blindness,” said the study’s lead investigator, Anand Swaroop, PhD, senior investigator and chief of the neurobiology neurodegeneration and repair laboratory at the NEI, part of the National Institutes of Health.
Swaroop and colleagues describe their findings in Nature Communications, in a paper titled, “High-resolution genome topology of human retina uncovers super enhancer-promoter interactions at tissue-specific and multifactorial disease loci,” in which they concluded: “Our studies thus provide a framework for connecting regulatory variants with retinal disease phenotypes and may assist in design of targeted translational paradigms by modulating genomic regulation.”
Adult human retinal cells are highly specialized sensory neurons that do not divide, and can be used to explore how the chromatin’s three-dimensional structure contributes to the expression of genetic information. “The adult retina is comprised of nondividing and highly-specialized sensory neurons and thus offers a relatively stable environment for investigating the role of 3D genome in controlling genetic information,” the team noted.
Chromatin fibers package long strands of DNA, which are spooled around histone proteins and then repeatedly looped to form highly compact structures. All those loops create multiple contact points where genetic sequences that code for proteins interact with gene regulatory sequences, such as super enhancers, promoters, and transcription factors.
Such noncoding sequences were long considered “junk DNA.” But more advanced studies demonstrate ways these sequences control which genes get transcribed and when, shedding light on the specific mechanisms by which noncoding regulatory elements exert control even when their location on a DNA strand is remote from the genes they regulate.
As the authors explained, “Across the 3D hierarchy, genome topology undergoes dynamic and contextual physical alterations in distinct tissues and cell types. These adaptations correlate with activation of specific cis-regulatory elements (CREs) that contribute to the establishment of unique gene expression patterns.”
Using deep Hi-C sequencing, a tool used for studying 3D genome organization, the researchers created a high-resolution map that included 704 million contact points within retinal cell chromatin. Maps were constructed using post-mortem retinal samples from four human donors. The researchers then integrated that chromatin topology map with datasets on retinal genes and regulatory elements. “To elucidate genomic regulation in human retina, we mapped chromatin contacts at high resolution and integrated with super-enhancers (SEs), histone marks, binding of CTCF, and select transcription factors,” they commented.
What emerged was a dynamic picture of interactions within chromatin over time, including gene activity hot spots and areas with varying degrees of insulation from other regions of DNA. “Further integration of chromatin contacts with histone marks, chromatin accessibility, selected TF [transcription factor] binding, and gene expression datasets reveals targets of CREs and uncovers properties of 3D chromatin organization of SEs in human retina.”
They found distinct patterns of interaction at retinal genes suggesting how chromatin’s 3D organization plays an important role in tissue-specific gene regulation. “Having such a high-resolution picture of genomic architecture will continue to provide insights into the genetic control of tissue-specific functions,” Swaroop said. Furthermore, similarities between mice and human chromatin organization suggest conservation across species, underscoring the relevance of chromatin organizational patterns for retinal gene regulation. More than a third (35.7%) of gene pairs interacting through a chromatin loop in mice also did so in human retina.
The researchers integrated the chromatin topology map with data on genetic variants identified from genome-wide association studies for their involvement in AMD and glaucoma, two leading causes of vision loss and blindness. The findings point to specific candidate causal genes involved in those diseases. “Merging genome-wide expression quantitative trait loci (eQTLs) with topology map reveals physical links between 100 eQTLs and corresponding eGenes associated with retinal neurodegeneration … by combining our findings with AMD and glaucoma GWAS, we have uncovered candidate causal genes contributing to these complex traits,” they wrote.
The integrated genome regulatory map will also assist in evaluating genes associated with other common retina-associated diseases such as diabetic retinopathy, determining missing heritability and understanding genotype-phenotype correlations in inherited retinal and macular diseases. The authors concluded, “In summary, we have generated a significant resource for the human retina, by integrating high-resolution Hi-C data with epigenetic profiles and CREs, thereby facilitating investigations of genomic regulation, identification of missing heritability in retinopathies, and candidate causal genes and variants for common blinding diseases including AMD and glaucoma.”