Genome-wide CRISPR Screening of Chondrocyte Maturation Newly Implicates Multiple Genes in Longitudinal Skeletal Growth and Height-GWAS Associated Loci [GPLC_CRISPR_seq]

The current study pairs human height GWAS data with CRISPR-based genome-wide knock out (KO) screens using an in vitro assay of chondrocyte proliferation and maturation to identify genes and gene pathways of biological relevance to human growth plate maturation. Our CRISPR screen detects relevant candidates for regulating human growth plate maturation, illustrating the value of functional studies in tissues of biological relevance as orthogonal data sets to refine likely causal genes from large-scale human genetic studies. GPLCs transduced with Cas9 and a genome-wide CRISPRko library were grown for 4 days or 15 days in culture, and FACS sorted for the cell surface marker of chondrocyte maturation, CD200. gDNA-based gene barcode counts from sequencing were used to perform differential analysis between KOs in the top 10% of CD200 expressing cells and the bottom 10% of CD200 expressing cells. A smaller secondary screening CRISPRko library was designed based on performance in the primary screen, and used for validation of top targets