Effects of the nsP2-726 Pro mutation on infectivity and pathogenesis of Sindbis virus derived from a full-length infectious cDNA clone.
The point mutations at residue 726 Pro in the nonstructural gene 2 (nsP2-726P) could make Sindbis virus (SINV) replicons lacking the structural protein-coding region less cytopathic and capable of persisting in some vertebrate cell lines. However, the effects of nsP2-726P mutations on characteristics of SINV in the context of genomic-RNA are poorly understood. To investigate the effects of point mutations at nsP2-726P on the infectivity and the pathogenesis of SINV, based on the infectious clone (pBR-XJ160) of a Sindbis-like XJ-160 virus, we constructed mutants BR-726L, BR-726S, BR-726V and BR-726A containing point mutations Pro-to-Leu, Pro-to-Ser, Pro-to-Val and Pro-to-Ala. The BR-726V virus and BR-726A virus exhibited similar growth characteristics to the wild-type BR-XJ160 in cultured cells, including cytopathic effects (CPE), plaque morphology and growth kinetics. For the Leu substitution, no CPE or plaques were seen after six passages through BHK-21 cells, although expression of XJ-160 virus-specific protein was detectable by indirect immunofluorescence assay (IFA). The Ser substitutions gave an intermediate phenotype. The mutant viruses exhibited different levels of neurovirulence in 3-day-old suckling mice, which did not match their propagation in cultured cells or in the mouse brain. Compared with BR-XJ160, BR-726A with the Ala substitution showed highly increased neurovirulence, while BR-726V with the Val substitution exhibited an attenuated phenotype. In contrast, BR-726S, with reduced growth capacity in cultured cells and mouse brain, showed intermediate neurovirulence. BR-726L virus produced no lethality or morbidity in suckling mice. Thus, the nsP2-726 Pro residue regulates virus-host cell interactions directly and is also important in viral pathogenesis in suckling mice.
Zhu
Wu-yang
WY
State Key Laboratory for Infectious Disease Prevention and Control (SKLID), Institute for Viral Disease Control and Prevention, Chinese Center for Disease Control and Prevention (IVDC, China CDC), Xuan Wu District, Beijing 100052, China.
Fu
Shi-hong
SH
Wang
Jing-lin
JL
He
Ying
Y
Tang
Qing
Q
Liang
Guo-dong
GD
D018354:Alphavirus Infections / Q000821:virology*; D000818:Animals; D001921:Brain / Q000821:virology; D002460:Cell Line; D006224:Cricetinae; D003546:Cysteine Endopeptidases / Q000235:genetics* / Q000378:metabolism; D018076:DNA, Complementary / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008647:Mesocricetus; D051379:Mice; D017354:Point Mutation; D012845:Sindbis Virus / Q000235:genetics / Q000472:pathogenicity* / Q000502:physiology*; D014774:Virulence
D018076:DNA, Complementary; D003546:Cysteine Endopeptidases; C078306:nsP2 proteinase