The cellular RNA helicase DDX1 interacts with coronavirus nonstructural protein 14 and enhances viral replication.
The involvement of host proteins in the replication and transcription of viral RNA is a poorly understood area for many RNA viruses. For coronaviruses, it was long speculated that replication of the giant RNA genome and transcription of multiple subgenomic mRNA species by a unique discontinuous transcription mechanism may require host cofactors. To search for such cellular proteins, yeast two-hybrid screening was carried out by using the nonstructural protein 14 (nsp14) from the coronavirus infectious bronchitis virus (IBV) as a bait protein, leading to the identification of DDX1, a cellular RNA helicase in the DExD/H helicase family, as a potential interacting partner. This interaction was subsequently confirmed by coimmunoprecipitation assays with cells coexpressing the two proteins and with IBV-infected cells. Furthermore, the endogenous DDX1 protein was found to be relocated from the nucleus to the cytoplasm in IBV-infected cells. In addition to its interaction with IBV nsp14, DDX1 could also interact with the nsp14 protein from severe acute respiratory syndrome coronavirus (SARS-CoV), suggesting that interaction with DDX1 may be a general feature of coronavirus nsp14. The interacting domains were mapped to the C-terminal region of DDX1 containing motifs V and VI and to the N-terminal portion of nsp14. Manipulation of DDX1 expression, either by small interfering RNA-induced knockdown or by overexpression of a mutant DDX1 protein, confirmed that this interaction may enhance IBV replication. This study reveals that DDX1 contributes to efficient coronavirus replication in cell culture.
Xu
Linghui
L
Institute of Molecular and Cell Biology, 61 Biopolis Drive, Proteos, Singapore.
Khadijah
Siti
S
Fang
Shouguo
S
Wang
Li
L
Tay
Felicia P L
FP
Liu
Ding Xiang
DX
D000818:Animals; D002467:Cell Nucleus / Q000378:metabolism; D002522:Chlorocebus aethiops; D018352:Coronavirus Infections / Q000235:genetics / Q000378:metabolism* / Q000821:virology; D003593:Cytoplasm / Q000378:metabolism; D053487:DEAD-box RNA Helicases / Q000235:genetics / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D001351:Infectious bronchitis virus / Q000235:genetics / Q000502:physiology*; D011485:Protein Binding; D021381:Protein Transport; D020798:Two-Hybrid System Techniques; D014709:Vero Cells; D017361:Viral Nonstructural Proteins / Q000235:genetics / Q000378:metabolism*; D014779:Virus Replication
D017361:Viral Nonstructural Proteins; C087633:nonstructural protein, coronavirus; C086252:DDX1 protein, human; D053487:DEAD-box RNA Helicases