Biochemical and genetic characterization of dengue virus methyltransferase.
We report that dengue virus (DENV) methyltransferase sequentially methylates the guanine N-7 and ribose 2'-O positions of viral RNA cap (GpppA-->m(7)GpppA-->m(7)GpppAm). The order of two methylations is determined by the preference of 2'-O methylation for substrate m(7)GpppA-RNA to GpppA-RNA, and the 2'-O methylation is not absolutely dependent on the prior N-7 methylation. A mutation that completely abolished the 2'-O methylation attenuated DENV replication in cell culture, whereas another mutation that abolished both methylations was lethal for viral replication, suggesting that N-7 methylation is more important than 2'-O methylation in viral replication. The latter mutant with lethal replication could be rescued by trans complementation using a wild-type DENV replicon. Furthermore, we found that chimeric DENVs containing the West Nile virus methyltransferase, polymerase, or full-length NS5 were nonreplicative, but the replication defect could also be rescued through trans complementation using the wild-type DENV replicon.
Dong
Hongping
H
Novartis Institute for Tropical Diseases, Chromos, Singapore 138670, Singapore. hongping.dong@novartis.com
Chang
David C
DC
Xie
Xuping
X
Toh
Ying Xiu
YX
Chung
Ka Yan
KY
Zou
Gang
G
Lescar
Julien
J
Lim
Siew Pheng
SP
Shi
Pei-Yong
PY
D000818:Animals; D002460:Cell Line; D006224:Cricetinae; D003716:Dengue Virus / Q000201:enzymology* / Q000235:genetics / Q000502:physiology; D008745:Methylation; D008780:Methyltransferases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008958:Models, Molecular; D012315:RNA Caps / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012367:RNA, Viral / Q000235:genetics / Q000378:metabolism; D014162:Transfection; D014764:Viral Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014779:Virus Replication
D012315:RNA Caps; D012367:RNA, Viral; D014764:Viral Proteins; D008780:Methyltransferases