21: Pooled prime editing maps functional human variants at scale

Herger M et al., Cell Genomics - Herger et al. present a pooled prime editing platform in haploid human cells that installs and assays thousands of short variants in their endogenous context. Using surrogate targets, co-selection and stringent pegRNA filtering, negative and positive selection screens identify loss-of-function variants in SMARCB1 and MLH1, including non-coding ClinVar variants that alter splicing. Key terms: prime editing, variant functional screening, SMARCB1, MLH1, HAP1 cells. Study Highlights:The authors develop a lentiviral pooled prime editing (PE) platform in HAP1 cells incorporating surrogate target (ST) sequences and co-selection to enrich edited cells. They optimize pegRNA scaffold design and show that ST editing rates are a useful proxy to filter active pegRNAs from inactive ones. Negative (essentiality) and positive (6-thioguanine) selection screens reveal novel loss-of-function variants in SMARCB1 and MLH1 across coding and non-coding regions. Stringent pegRNA activity filtering and orthogonal validation are necessary to reduce false positives and negatives. Conclusion:Pooled prime editing with surrogate targets and co-selection can scalably reveal functional effects of coding and non-coding human variants in their native genomic context, but accurate scoring depends on pegRNA activity and validation; improvements in pegRNA design and prime editors should expand genome-wide applicability. QC:This episode was checked against the original article PDF and publication metadata for the episode release published on 2025-05-16. QC Scope:- article metadata and core scientific claims from the narration- excludes analogies, intro/outro, and music QC Summary:- factual score: 10/10- metadata score: 10/10- supported core claims: 8- claims flagged for review: 0- metadata checks passed: 4- metadata issues found: 0 Metadata Audited:- article_doi- article_title- article_journal- license Factual Items Audited:- Pooled prime editing platform enables scalable installation of variants in haploid human cells (HAP1).- Surrogate targets (ST) and co-selection improve data quality by informing pegRNA activity and enriching edited cells.- Negative selection identifies LoF variants in SMARCB1; positive selection using 6TG identifies LoF variants in MLH1.- ST editing rates correlate with endogenous editing; ST editing is a proxy for ET editing.- Non-coding ClinVar variants in MLH1 can be functionally scored; 362 of 874 non-coding ClinVar variants were assigned function scores.- Pathogenic ClinVar variants in the MLH1 region show LoF scores; 54% called LoF; benign 2.4%. QC result: Pass.