TY - JOUR
T1 - CRISPR/Cas9 screens identify key host factors that enhance rotavirus reverse genetics efficacy and vaccine production
AU - Zhu, Yinxing
AU - Sullender, Meagan E.
AU - Campbell, Danielle E.
AU - Wang, Leran
AU - Lee, Sanghyun
AU - Kawagishi, Takahiro
AU - Hou, Gaopeng
AU - Dizdarevic, Alen
AU - Jais, Philippe H.
AU - Baldridge, Megan T.
AU - Ding, Siyuan
N1 - Publisher Copyright:
© The Author(s) 2024.
PY - 2024/12
Y1 - 2024/12
N2 - Rotaviruses pose a significant threat to young children. To identify novel pro- and anti-rotavirus host factors, we performed genome-wide CRISPR/Cas9 screens using rhesus rotavirus and African green monkey cells. Genetic deletion of either SERPINB1 or TMEM236, the top two antiviral factors, in MA104 cells increased virus titers in a rotavirus strain independent manner. Using this information, we optimized the existing rotavirus reverse genetics systems by combining SERPINB1 knockout MA104 cells with a C3P3-G3 helper plasmid. We improved the recovery efficiency and rescued several low-titer rotavirus reporter and mutant strains that prove difficult to rescue otherwise. Furthermore, we demonstrate that TMEM236 knockout in Vero cells supported higher yields of two live-attenuated rotavirus vaccine strains than the parental cell line and represents a more robust vaccine-producing cell substrate. Collectively, we developed a third-generation optimized rotavirus reverse genetics system and generated gene-edited Vero cells as a new substrate for improving rotavirus vaccine production.
AB - Rotaviruses pose a significant threat to young children. To identify novel pro- and anti-rotavirus host factors, we performed genome-wide CRISPR/Cas9 screens using rhesus rotavirus and African green monkey cells. Genetic deletion of either SERPINB1 or TMEM236, the top two antiviral factors, in MA104 cells increased virus titers in a rotavirus strain independent manner. Using this information, we optimized the existing rotavirus reverse genetics systems by combining SERPINB1 knockout MA104 cells with a C3P3-G3 helper plasmid. We improved the recovery efficiency and rescued several low-titer rotavirus reporter and mutant strains that prove difficult to rescue otherwise. Furthermore, we demonstrate that TMEM236 knockout in Vero cells supported higher yields of two live-attenuated rotavirus vaccine strains than the parental cell line and represents a more robust vaccine-producing cell substrate. Collectively, we developed a third-generation optimized rotavirus reverse genetics system and generated gene-edited Vero cells as a new substrate for improving rotavirus vaccine production.
UR - http://www.scopus.com/inward/record.url?scp=85208746401&partnerID=8YFLogxK
U2 - 10.1038/s41541-024-01007-7
DO - 10.1038/s41541-024-01007-7
M3 - Article
C2 - 39505878
AN - SCOPUS:85208746401
SN - 2059-0105
VL - 9
JO - npj Vaccines
JF - npj Vaccines
IS - 1
M1 - 211
ER -