Systematic functional interrogation of human pseudogenes

Human genome encodes >14,000 pseudogenes that are evolutionary relics and have long been considered as nonfunctional genomic elements. Emerging evidence suggests that pseudogene can exert important regulatory function. However, function of most pseudogenes remains unknown. To fill this gap, we developed an integrated computational pipeline and performed to date the first set of pseudogene-focused CRISPRi screens in human cells. Our screens identified >100 pseudogenes that are important for cell fitness, with a more cell-type specific function compared to parent genes. In addition, we discovered a cancer-testis unitary pseudogene MGAT4EP that interacts with FOXA1, a key regulator in luminal A breast cancer. Overall design: 5703 sgRNAs target pseudogenes and 3727 sgRNAs target their parent genes were included in the library. The CRISPRi screen was conducted in MCF7 cell line. MGAT4EP expression was down-regulated by sgRNA in dcas9 expressed MCF7 cells, and then next generation sequencing was performed to identify MGAT4EP regulated genes.