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1. Genetic determinants of global developmental delay and intellectual disability in Ukrainian children

   https://doi.org/10.1186/s11689-024-09528-x  

   Shchubelka, Khrystyna; Turova, Liudmyla; Wolfsberger, Walter; Kalanquin, Kelly; Williston, Krista; Kurutsa, Oleksii; Makovetska, Anastasiia; Hasynets, Yaroslava; Mirutenko, Violeta; Vakerych, Mykhailo; et al (December 2024, Journal of Neurodevelopmental Disorders)

   Abstract BackgroundGlobal developmental delay or intellectual disability usually accompanies various genetic disorders as a part of the syndrome, which may include seizures, autism spectrum disorder and multiple congenital abnormalities. Next-generation sequencing (NGS) techniques have improved the identification of pathogenic variants and genes related to developmental delay. This study aimed to evaluate the yield of whole exome sequencing (WES) and neurodevelopmental disorder gene panel sequencing in a pediatric cohort from Ukraine. Additionally, the study computationally predicted the effect of variants of uncertain significance (VUS) based on recently published genetic data from the country’s healthy population. MethodsThe study retrospectively analyzed WES or gene panel sequencing findings of 417 children with global developmental delay, intellectual disability, and/or other symptoms. Variants of uncertain significance were annotated using CADD-Phred and SIFT prediction scores, and their frequency in the healthy population of Ukraine was estimated. ResultsA definitive molecular diagnosis was established in 66 (15.8%) of the individuals. WES diagnosed 22 out of 37 cases (59.4%), while the neurodevelopmental gene panel identified 44 definitive diagnoses among the 380 tested patients (12.1%). Non-diagnostic findings (VUS and carrier) were reported in 350 (83.2%) individuals. The most frequently diagnosed conditions were developmental and epileptic encephalopathies associated with severe epilepsy and GDD/ID (associated genesARX, CDKL5, STXBP1, KCNQ2, SCN2A, KCNT1, KCNA2). Additionally, we annotated 221 VUS classified as potentially damaging, AD or X-linked, potentially increasing the diagnostic yield by 30%, but 18 of these variants were present in the healthy population of Ukraine. ConclusionsThis is the first comprehensive study on genetic causes of GDD/ID conducted in Ukraine. This study provides the first comprehensive investigation of the genetic causes of GDD/ID in Ukraine. It presents a substantial dataset of diagnosed genetic conditions associated with GDD/ID. The results support the utilization of NGS gene panels and WES as first-line diagnostic tools for GDD/ID cases, particularly in resource-limited settings. A comprehensive approach to resolving VUS, including computational effect prediction, population frequency analysis, and phenotype assessment, can aid in further reclassification of deleterious VUS and guide further testing in families. 

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2. Structure and Sequence of the Sex Determining Locus in Two Wild Populations of Nile Tilapia

   https://doi.org/10.3390/genes11091017  

   Triay, Cécile; Conte, Matthew A.; Baroiller, Jean-François; Bezault, Etienne; Clark, Frances E.; Penman, David J.; Kocher, Thomas D.; D’Cotta, Helena (September 2020, Genes)

   null (Ed.)

   In domesticated strains of the Nile tilapia, phenotypic sex has been linked to genetic variants on linkage groups 1, 20 and 23. This diversity of sex-loci might reflect a naturally polymorphic sex determination system in Nile tilapia, or it might be an artefact arising from the process of domestication. Here, we searched for sex-determiners in wild populations from Kpandu, Lake Volta (Ghana-West Africa), and from Lake Koka (Ethiopia-East Africa) that have not been subjected to any genetic manipulation. We analysed lab-reared families using double-digest Restriction Associated DNA sequencing (ddRAD) and analysed wild-caught males and females with pooled whole-genome sequencing (WGS). Strong sex-linked signals were found on LG23 in both populations, and sex-linked signals with LG3 were observed in Kpandu samples. WGS uncovered blocks of high sequence coverage, suggesting the presence of B chromosomes. We confirmed the existence of a tandem amh duplication in LG23 in both populations and determined its breakpoints between the oaz1 and dot1l genes. We found two common deletions of ~5 kb in males and confirmed the presence of both amhY and amh∆Y genes. Males from Lake Koka lack both the previously reported 234 bp deletion and the 5 bp frameshift-insertion that creates a premature stop codon in amh∆Y. 

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3. Natural genetic variation in the pheromone production of C. elegans

   https://doi.org/10.1073/pnas.2221150120  

   Lee, Daehan; Fox, Bennett W.; Palomino, Diana Fajardo; Panda, Oishika; Tenjo, Francisco J.; Koury, Emily J.; Evans, Kathryn S.; Stevens, Lewis; Rodrigues, Pedro R.; Kolodziej, Aiden R.; et al (June 2023, Proceedings of the National Academy of Sciences)

   From bacterial quorum sensing to human language, communication is essential for social interactions. Nematodes produce and sense pheromones to communicate among individuals and respond to environmental changes. These signals are encoded by different types and mixtures of ascarosides, whose modular structures further enhance the diversity of this nematode pheromone language. Interspecific and intraspecific differences in this ascaroside pheromone language have been described previously, but the genetic basis and molecular mechanisms underlying the variation remain largely unknown. Here, we analyzed natural variation in the production of 44 ascarosides across 95 wild Caenorhabditis elegans strains using high-performance liquid chromatography coupled to high-resolution mass spectrometry. We discovered wild strains defective in the production of specific subsets of ascarosides ( e.g. , the aggregation pheromone icas#9) or short- and medium-chain ascarosides, as well as inversely correlated patterns between the production of two major classes of ascarosides. We investigated genetic variants that are significantly associated with the natural differences in the composition of the pheromone bouquet, including rare genetic variants in key enzymes participating in ascaroside biosynthesis, such as the peroxisomal 3-ketoacyl-CoA thiolase, daf-22 , and the carboxylesterase cest-3 . Genome-wide association mappings revealed genomic loci harboring common variants that affect ascaroside profiles. Our study yields a valuable dataset for investigating the genetic mechanisms underlying the evolution of chemical communication. 

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4. Multiple Loci Control Eyespot Number Variation on the Hindwings of Bicyclus anynana Butterflies

   https://doi.org/10.1534/genetics.120.303059  

   Rivera-Colón, Angel G.; Westerman, Erica L.; Van Belleghem, Steven M.; Monteiro, Antónia; Papa, Riccardo (April 2020, Genetics)

   The underlying genetic changes that regulate the appearance and disappearance of repeated traits, or serial homologs, remain poorly understood. One hypothesis is that variation in genomic regions flanking master regulatory genes, also known as input–output genes, controls variation in trait number, making the locus of evolution almost predictable. Another hypothesis implicates genetic variation in up- or downstream loci of master control genes. Here, we use the butterfly Bicyclus anynana , a species that exhibits natural variation in eyespot number on the dorsal hindwing, to test these two hypotheses. We first estimated the heritability of dorsal hindwing eyespot number by breeding multiple butterfly families differing in eyespot number and regressing eyespot numbers of offspring on midparent values. We then estimated the number and identity of independent genetic loci contributing to eyespot number variation by performing a genome-wide association study with restriction site-associated DNA sequencing from multiple individuals varying in number of eyespots sampled across a freely breeding laboratory population. We found that dorsal hindwing eyespot number has a moderately high heritability of ∼0.50 and is characterized by a polygenic architecture. Previously identified genomic regions involved in eyespot development, and novel ones, display high association with dorsal hindwing eyespot number, suggesting that homolog number variation is likely determined by regulatory changes at multiple loci that build the trait, and not by variation at single master regulators or input–output genes. 

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5. Discovery of target genes and pathways at GWAS loci by pooled single-cell CRISPR screens

   https://doi.org/10.1126/science.adh7699  

   Morris, John A.; Caragine, Christina; Daniloski, Zharko; Domingo, Júlia; Barry, Timothy; Lu, Lu; Davis, Kyrie; Ziosi, Marcello; Glinos, Dafni A.; Hao, Stephanie; et al (May 2023, Science)

   INTRODUCTION Genome-wide association studies (GWASs) have identified thousands of human genetic variants associated with diverse diseases and traits, and most of these variants map to noncoding loci with unknown target genes and function. Current approaches to understand which GWAS loci harbor causal variants and to map these noncoding regulators to target genes suffer from low throughput. With newer multiancestry GWASs from individuals of diverse ancestries, there is a pressing and growing need to scale experimental assays to connect GWAS variants with molecular mechanisms. Here, we combined biobank-scale GWASs, massively parallel CRISPR screens, and single-cell sequencing to discover target genes of noncoding variants for blood trait loci with systematic targeting and inhibition of noncoding GWAS loci with single-cell sequencing (STING-seq). RATIONALE Blood traits are highly polygenic, and GWASs have identified thousands of noncoding loci that map to candidate cis -regulatory elements (CREs). By combining CRE-silencing CRISPR perturbations and single-cell readouts, we targeted hundreds of GWAS loci in a single assay, revealing target genes in cis and in trans . For select CREs that regulate target genes, we performed direct variant insertion. Although silencing the CRE can identify the target gene, direct variant insertion can identify magnitude and direction of effect on gene expression for the GWAS variant. In select cases in which the target gene was a transcription factor or microRNA, we also investigated the gene-regulatory networks altered upon CRE perturbation and how these networks differ across blood cell types. RESULTS We inhibited candidate CREs from fine-mapped blood trait GWAS variants (from ~750,000 individual of diverse ancestries) in human erythroid progenitors. In total, we targeted 543 variants (254 loci) mapping to candidate CREs, generating multimodal single-cell data including transcriptome, direct CRISPR gRNA capture, and cell surface proteins. We identified target genes in cis (within 500 kb) for 134 CREs. In most cases, we found that the target gene was the closest gene and that specific enhancer-associated biochemical hallmarks (H3K27ac and accessible chromatin) are essential for CRE function. Using multiple perturbations at the same locus, we were able to distinguished between causal variants from noncausal variants in linkage disequilibrium. For a subset of validated CREs, we also inserted specific GWAS variants using base-editing STING-seq (beeSTING-seq) and quantified the effect size and direction of GWAS variants on gene expression. Given our transcriptome-wide data, we examined dosage effects in cis and trans in cases in which the cis target is a transcription factor or microRNA. We found that trans target genes are also enriched for GWAS loci, and identified gene clusters within trans gene networks with distinct biological functions and expression patterns in primary human blood cells. CONCLUSION In this work, we investigated noncoding GWAS variants at scale, identifying target genes in single cells. These methods can help to address the variant-to-function challenges that are a barrier for translation of GWAS findings (e.g., drug targets for diseases with a genetic basis) and greatly expand our ability to understand mechanisms underlying GWAS loci. Identifying causal variants and their target genes with STING-seq. Uncovering causal variants and their target genes or function are a major challenge for GWASs. STING-seq combines perturbation of noncoding loci with multimodal single-cell sequencing to profile hundreds of GWAS loci in parallel. This approach can identify target genes in cis and trans , measure dosage effects, and decipher gene-regulatory networks. 

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