Single cell RNA sequencing from zebrafish embryos

Through an in vivo genome-wide CRISPR/Cas9 transcriptional activation screen using human Ewing sarcoma (ES) cells as xenografts, we identified the human inaF homolog (INAFM2) as a strong driver of metastasis. Therefore, we named the vertebrate gene and its protein product ROME (Regulator of Metastasis). We showed that ROME negatively regulates the Ca2+ and canonical Wnt pathways and interacts directly with the CAV1, FLOT1, and vimentin proteins. Blocking ROME expression in zebrafish embryos resulted in severe developmental defects and early mortality. ROME overexpression increased the metastasis of ES xenografts in zebrafish and immunodeficient mice, whereas ROME knockout in ES xenografts reduced metastasis. RNA sequencing data from human tumors revealed a negative correlation between ROME expression and patient survival. In summary, we discovered a previously unstudied human plasma membrane glycoprotein that can regulate cell motility and invasion, is essential for normal development in vertebrates, and exacerbates metastasis-related phenotypes in human tumor cells.