Expression of L protein of Hantaan virus 84FLi strain and its application for recovery of minigenomes.
Hantaan virus (HTNV) is an Old World hantavirus associated with hemorrhagic fever with renal syndrome (HFRS). To visualize the localization of the L protein of HTNV strain 84FLi within cells, a fusion protein composed of enhanced green fluorescent protein and L protein, EGFP-L, was expressed in Vero cells. The 273 KDa expressed fusion protein of EGFP-L localized in the perinuclear region. We also described the development of a reverse genetics system for HTNV strain 84FLi. The RNA polymerase I (pol I)-mediated transcription system was used to generate artificial viral RNA genome segments (minigenomes), which contained the chloramphenicol acetyltransferase (CAT) reporter gene in antisense (virus RNA) or sense (virus-complementary RNA) orientation flanked by the noncoding regions of HTNV 84FLi L segment. CAT could be detected in cells after transfection, indicating the successful encapsidation, transcription and replication of the pol I-derived minigenomes. The passaged transfer of CAT demonstrates that recombinant virus containing packaged pol I-derived minigenomes has been produced. This system may be helpful in studying the gene function and pathogenesis of HTNV.
Zhang
Ye
Y
Center of Infectious Diseases, Tangdu Hospital, Fourth Military Medical University, Xi'an, P. R. China.
Li
Xin-Hong
XH
Jiang
Hong
H
Huang
Chang-Xing
CX
Wang
Ping-Zhong
PZ
Mou
Dan-Lei
DL
Sun
Li
L
Xu
Zhe
Z
Wei
Xin
X
Bai
Xue-Fan
XF
D000818:Animals; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics; D002522:Chlorocebus aethiops; D003593:Cytoplasm / Q000378:metabolism; D015870:Gene Expression; D017930:Genes, Reporter; D016679:Genome, Viral; D049452:Green Fluorescent Proteins / Q000032:analysis / Q000096:biosynthesis; D018063:Hantaan virus / Q000235:genetics / Q000502:physiology*; D018778:Hantavirus Infections / Q000821:virology*; D006801:Humans; D012318:RNA Polymerase I / Q000378:metabolism; D012367:RNA, Viral / Q000235:genetics; D012324:RNA-Dependent RNA Polymerase / Q000032:analysis* / Q000096:biosynthesis; D011993:Recombinant Fusion Proteins / Q000037:antagonists & inhibitors* / Q000096:biosynthesis; D048348:Reverse Transcription; D014709:Vero Cells / Q000378:metabolism; D014764:Viral Proteins / Q000032:analysis* / Q000096:biosynthesis
D012367:RNA, Viral; D011993:Recombinant Fusion Proteins; D014764:Viral Proteins; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D015500:Chloramphenicol O-Acetyltransferase; C500570:L protein, hantavirus; D012324:RNA-Dependent RNA Polymerase; D012318:RNA Polymerase I