Integrating genetic regulation and single-cell expression with GWAS prioritizes causal genes and cell types for glaucoma.

Primary open-angle glaucoma (POAG), characterized by retinal ganglion cell death, is a leading cause of irreversible blindness worldwide. However, its molecular and cellular causes are not well understood. Elevated intraocular pressure (IOP) is a major risk factor, but many patients have normal IOP. Colocalization and Mendelian randomization analysis of >240 POAG and IOP genome-wide association study (GWAS) loci and overlapping expression and splicing quantitative trait loci (e/sQTLs) in 49 GTEx tissues and retina prioritizes causal genes for 60% of loci. These genes are enriched in pathways implicated in extracellular matrix organization, cell adhesion, and vascular development. Analysis of single-nucleus RNA-seq of glaucoma-relevant eye tissues reveals that the POAG and IOP colocalizing genes and genome-wide associations are enriched in specific cell types in the aqueous outflow pathways, retina, optic nerve head, peripapillary sclera, and choroid. This study nominates IOP-dependent and independent regulatory mechanisms, genes, and cell types that may contribute to POAG pathogenesis.

Single-nucleus cross-tissue molecular reference maps toward understanding disease gene function.

Understanding gene function and regulation in homeostasis and disease requires knowledge of the cellular and tissue contexts in which genes are expressed. Here, we applied four single-nucleus RNA sequencing methods to eight diverse, archived, frozen tissue types from 16 donors and 25 samples, generating a cross-tissue atlas of 209,126 nuclei profiles, which we integrated across tissues, donors, and laboratory methods with a conditional variational autoencoder. Using the resulting cross-tissue atlas, we highlight shared and tissue-specific features of tissue-resident cell populations; identify cell types that might contribute to neuromuscular, metabolic, and immune components of monogenic diseases and the biological processes involved in their pathology; and determine cell types and gene modules that might underlie disease mechanisms for complex traits analyzed by genome-wide association studies.

Population structure of ocular Streptococcus pneumoniae is highly diverse and formed by lineages that escape current vaccines.

Streptococcus pneumoniae is a leading cause of ocular infections including serious and sight-threatening conditions. The use of pneumococcal conjugate vaccines (PCV) has substantially reduced the incidence of pneumonia and invasive pneumococcal diseases, but has had limited impact on ocular infections. Additionally, widespread vaccine use has resulted in ongoing selective pressure and serotype replacement in carriage and disease. To gain insight into the population structure of pneumococcal isolates causing ocular infections in a post-PCV-13 time period, we investigated the genomic epidemiology of ocular S. pneumoniae isolates (n=45) collected at Massachusetts Eye and Ear between 2014 and 2017. By performing a series of molecular typing methods from draft genomes, we found that the population structure of ocular S. pneumoniae is highly diverse with 27 sequence types (grouped into 18 clonal complexes) and 17 serotypes being identified. Distribution of these lineages diverged according to the site of isolation, with conjunctivitis being commonly caused by isolates grouped in the Epidemic Conjunctivitis Cluster-ECC (60 %), and ST448 (53.3 %) being most frequently identified. Conversely, S. pneumoniae keratitis cases were caused by a highly diverse population of isolates grouping within 15 different clonal complexes. Serotyping inference demonstrated that 95.5 % of the isolates were non-PCV-13 vaccine types. Most of the conjunctivitis isolates (80 %) were unencapsulated, with the remaining belonging to serotypes 15B, 3 and 23B. On the other hand, S. pneumoniae causing keratitis were predominantly encapsulated (95.2 %) with 13 different serotypes identified, mostly being non-vaccine types. Carriage of macrolide resistance genes was common in our ocular S. pneumoniae population (42.2 %), and usually associated with the mefA +msrD genotype (n=15). These genes were located in the Macrolide Efflux Genetic Assembly cassette and were associated with low-level in vitro resistance to 14- and 15-membered macrolides. Less frequently, macrolide-resistant isolates carried an ermB gene (n=4), which was co-located with the tetM gene in a Tn-916-like transposon. Our study demonstrates that the population structure of ocular S. pneumoniae is highly diverse, mainly composed by isolates that escape the PCV-13 vaccine, with patterns of tissue/niche segregation, adaptation and specialization. These findings suggest that the population structure of ocular pneumococcus may be shaped by multiple factors including PCV-13 selective pressure, microbial-related and niche-specific host-associated features.

Gene Set Enrichment Analsyes Identify Pathways Involved in Genetic Risk for Diabetic Retinopathy.

To identify functionally related genes associated with diabetic retinopathy (DR) risk using gene set enrichment analyses applied to genome-wide association study meta-analyses. METHODS: We analyzed DR GWAS meta-analyses performed on 3246 Europeans and 2611 African Americans with type 2 diabetes. Gene sets relevant to 5 key DR pathophysiology processes were investigated: tissue injury, vascular events, metabolic events and glial dysregulation, neuronal dysfunction, and inflammation. Keywords relevant to these processes were queried in 4 pathway and ontology databases. Two GSEA methods, Meta-Analysis Gene set Enrichment of variaNT Associations (MAGENTA) and Multi-marker Analysis of GenoMic Annotation (MAGMA), were used. Gene sets were defined to be enriched for gene associations with DR if the P value corrected for multiple testing (Pcorr) was <.05. RESULTS: Five gene sets were significantly enriched for numerous modest genetic associations with DR in one method (MAGENTA or MAGMA) and also at least nominally significant (uncorrected P < .05) in the other method. These pathways were regulation of the lipid catabolic process (2-fold enrichment, Pcorr = .014); nitric oxide biosynthesis (1.92-fold enrichment, Pcorr = .022); lipid digestion, mobilization, and transport (1.6-fold enrichment, P = .032); apoptosis (1.53-fold enrichment, P = .041); and retinal ganglion cell degeneration (2-fold enrichment, Pcorr = .049). The interferon gamma (IFNG) gene, previously implicated in DR by protein-protein interactions in our GWAS, was among the top ranked genes in the nitric oxide pathway (best variant P = .0001). CONCLUSIONS: These GSEA indicate that variants in genes involved in oxidative stress, lipid transport and catabolism, and cell degeneration are enriched for genes associated with DR risk. NOTE: Publication of this article is sponsored by the American Ophthalmological Society.

Genome-wide meta-analysis identifies 127 open-angle glaucoma loci with consistent effect across ancestries.

Primary open-angle glaucoma (POAG), is a heritable common cause of blindness world-wide. To identify risk loci, we conduct a large multi-ethnic meta-analysis of genome-wide association studies on a total of 34,179 cases and 349,321 controls, identifying 44 previously unreported risk loci and confirming 83 loci that were previously known. The majority of loci have broadly consistent effects across European, Asian and African ancestries. Cross-ancestry data improve fine-mapping of causal variants for several loci. Integration of multiple lines of genetic evidence support the functional relevance of the identified POAG risk loci and highlight potential contributions of several genes to POAG pathogenesis, including SVEP1, RERE, VCAM1, ZNF638, CLIC5, SLC2A12, YAP1, MXRA5, and SMAD6. Several drug compounds targeting POAG risk genes may be potential glaucoma therapeutic candidates.