RESUMO
Nucleotide changes in gene regulatory elements are important determinants of neuronal development and diseases. Using massively parallel reporter assays in primary human cells from mid-gestation cortex and cerebral organoids, we interrogated the cis-regulatory activity of 102,767 open chromatin regions, including thousands of sequences with cell type-specific accessibility and variants associated with brain gene regulation. In primary cells, we identified 46,802 active enhancer sequences and 164 variants that alter enhancer activity. Activity was comparable in organoids and primary cells, suggesting that organoids provide an adequate model for the developing cortex. Using deep learning we decoded the sequence basis and upstream regulators of enhancer activity. This work establishes a comprehensive catalog of functional gene regulatory elements and variants in human neuronal development.
Assuntos
Córtex Cerebral , Neurogênese , Organoides , Humanos , Córtex Cerebral/embriologia , Córtex Cerebral/metabolismo , Cromatina/metabolismo , Cromatina/genética , Aprendizado Profundo , Elementos Facilitadores Genéticos , Regulação da Expressão Gênica no Desenvolvimento , Neurogênese/genética , Neurônios/metabolismo , Organoides/metabolismo , Sequências Reguladoras de Ácido Nucleico , Regiões Promotoras Genéticas , Elementos Reguladores de TranscriçãoRESUMO
Regulation of gene expression through enhancers is one of the major processes shaping the structure and function of the human brain during development. High-throughput assays have predicted thousands of enhancers involved in neurodevelopment, and confirming their activity through orthogonal functional assays is crucial. Here, we utilized Massively Parallel Reporter Assays (MPRAs) in stem cells and forebrain organoids to evaluate the activity of ~ 7000 gene-linked enhancers previously identified in human fetal tissues and brain organoids. We used a Gaussian mixture model to evaluate the contribution of background noise in the measured activity signal to confirm the activity of ~ 35% of the tested enhancers, with most showing temporal-specific activity, suggesting their evolving role in neurodevelopment. The temporal specificity was further supported by the correlation of activity with gene expression. Our findings provide a valuable gene regulatory resource to the scientific community.