02403nas a2200397   4500000000100000000000100001008004100002260001500043653003100058653002400089653002600113653001500139653002800154100001600182700002200198700001900220700001800239700001800257700002400275700002000299700001900319700002100338700001900359700002600378700001500404700003400419700002100453700002100474700001700495245010700512856005500619300000900674490000700683520130100690022001401991       2021                            d  c2021-09-1710aCRISPR-Cas9 genome editing10aGenetic engineering10aintestinal stem cells10aSARS-CoV-210aStem-cell biotechnology1 aJoep Beumer1 aMaarten H. Geurts1 aMart M. Lamers1 aJens Puschhof1 aJingshu Zhang1 aJelte van der Vaart1 aAnna Z. Mykytyn1 aTim I. Breugem1 aSamra Riesebosch1 aDebby Schipper1 aPetra B. van den Doel1 aWim de Lau1 aCayetano Pleguezuelos-Manzano1 aGeorg Busslinger1 aBart L. Haagmans1 aHans Clevers00aA CRISPR/Cas9 genetically engineered organoid biobank reveals essential host factors for coronaviruses  uhttps://www.nature.com/articles/s41467-021-25729-7  a54980 v123 aRapid identification of host genes essential for virus replication may expedite the generation of therapeutic interventions. Genetic screens are often performed in transformed cell lines that poorly represent viral target cells in vivo, leading to discoveries that may not be translated to the clinic. Intestinal organoids are increasingly used to model human disease and are amenable to genetic engineering. To discern which host factors are reliable anti-coronavirus therapeutic targets, we generate mutant clonal IOs for 19 host genes previously implicated in coronavirus biology. We verify ACE2 and DPP4 as entry receptors for SARS-CoV/SARS-CoV-2 and MERS-CoV respectively. SARS-CoV-2 replication in IOs does not require the endosomal Cathepsin B/L proteases, but specifically depends on the cell surface protease TMPRSS2. Other TMPRSS family members were not essential. The newly emerging coronavirus variant B.1.1.7, as well as SARS-CoV and MERS-CoV similarly depended on TMPRSS2. These findings underscore the relevance of non-transformed human models for coronavirus research, identify TMPRSS2 as an attractive pan-coronavirus therapeutic target, and demonstrate that an organoid knockout biobank is a valuable tool to investigate the biology of current and future emerging coronaviruses.  a2041-1723