PMID- 29778185
OWN - NLM
STAT- MEDLINE
DCOM- 20181120
LR  - 20200407
IS  - 1873-2542 (Electronic)
IS  - 0378-1135 (Print)
IS  - 0378-1135 (Linking)
VI  - 219
DP  - 2018 Jun
TI  - Serine 105 and 120 are important phosphorylation sites for porcine reproductive 
      and respiratory syndrome virus N protein function.
PG  - 128-135
LID - S0378-1135(18)30209-8 [pii]
LID - 10.1016/j.vetmic.2018.04.010 [doi]
AB  - The nucleocapsid (N) protein is the most abundant protein of porcine reproductive 
      and respiratory syndrome virus (PRRSV). It has been shown to be 
      multiphosphorylated. However, the phosphorylation sites are still unknown. In 
      this study, we used liquid chromatography tandem mass spectrometry (LC-MS/MS) to 
      analyze the phosphorylation sites of N protein expressed in Sf9 cells. The 
      results showed that N protein contains two phosphorylation sites. Since N protein 
      can regulate IL-10, which may facilitate PRRSV replication, we constructed four 
      plasmids (pCA-XH-GD, pCA-A105, pCA-A120 and pCA-A105-120) and transfected them 
      into Pig alveolar macrophages (PAMs,3D4/2). The qPCR results showed that 
      mutations at residues 105 and 120 were associated with down-regulation of the 
      IL-10 mRNA level, potentially decreasing the viral growth ability. Then, we 
      mutated the phosphorylation sites (S105A and S120A) and rescued three mutated 
      viruses, namely, A105, A120 and A105-120. Compared with wild-type virus titers, 
      the titers of the mutated viruses at 48 hpi were significantly decreased. The 
      Nsp(non-structural protein) 9 qPCR results were consistent with the multistep 
      growth kinetics results. The infected PAMs (primary PAMs) results were similar 
      with Marc-145.The findings indicated that the mutations impaired the viral 
      replication ability. The confocal microscopy results suggested that mutations to 
      residues 105 and 120 did not affect N protein distribution. Whether the mutations 
      affected other functions of N protein and what the underlying mechanisms are need 
      further investigation. In conclusion, our results show that residues 105 and 120 
      are phosphorylation sites and are important for N protein function and for viral 
      replication ability.
CI  - Copyright (c) 2018 Elsevier B.V. All rights reserved.
FAU - Chen, Yao
AU  - Chen Y
AD  - Key Laboratory of Zoonosis Prevention and Control of Guangdong Province, College 
      of Veterinary Medicine, South China Agricultural University, Guangzhou, China; 
      College of Veterinary and National Engineering Research Center for Breeding Swine 
      Industry, South China Agricultural University, Guangzhou, China; National and 
      regional Joint Engineering Laboratory for Medicament of Zoonosis Prevention and 
      Control, China.
FAU - Xing, Xiulin
AU  - Xing X
AD  - Key Laboratory of Zoonosis Prevention and Control of Guangdong Province, College 
      of Veterinary Medicine, South China Agricultural University, Guangzhou, China; 
      College of Veterinary and National Engineering Research Center for Breeding Swine 
      Industry, South China Agricultural University, Guangzhou, China; National and 
      regional Joint Engineering Laboratory for Medicament of Zoonosis Prevention and 
      Control, China.
FAU - Li, Qi
AU  - Li Q
AD  - Key Laboratory of Zoonosis Prevention and Control of Guangdong Province, College 
      of Veterinary Medicine, South China Agricultural University, Guangzhou, China; 
      College of Veterinary and National Engineering Research Center for Breeding Swine 
      Industry, South China Agricultural University, Guangzhou, China; National and 
      regional Joint Engineering Laboratory for Medicament of Zoonosis Prevention and 
      Control, China.
FAU - Feng, Songlin
AU  - Feng S
AD  - Key Laboratory of Zoonosis Prevention and Control of Guangdong Province, College 
      of Veterinary Medicine, South China Agricultural University, Guangzhou, China; 
      College of Veterinary and National Engineering Research Center for Breeding Swine 
      Industry, South China Agricultural University, Guangzhou, China; National and 
      regional Joint Engineering Laboratory for Medicament of Zoonosis Prevention and 
      Control, China.
FAU - Han, Xiaoliang
AU  - Han X
AD  - Key Laboratory of Zoonosis Prevention and Control of Guangdong Province, College 
      of Veterinary Medicine, South China Agricultural University, Guangzhou, China; 
      College of Veterinary and National Engineering Research Center for Breeding Swine 
      Industry, South China Agricultural University, Guangzhou, China; National and 
      regional Joint Engineering Laboratory for Medicament of Zoonosis Prevention and 
      Control, China.
FAU - He, Shuyi
AU  - He S
AD  - Key Laboratory of Zoonosis Prevention and Control of Guangdong Province, College 
      of Veterinary Medicine, South China Agricultural University, Guangzhou, China; 
      College of Veterinary and National Engineering Research Center for Breeding Swine 
      Industry, South China Agricultural University, Guangzhou, China; National and 
      regional Joint Engineering Laboratory for Medicament of Zoonosis Prevention and 
      Control, China.
FAU - Zhang, Guihong
AU  - Zhang G
AD  - Key Laboratory of Zoonosis Prevention and Control of Guangdong Province, College 
      of Veterinary Medicine, South China Agricultural University, Guangzhou, China; 
      College of Veterinary and National Engineering Research Center for Breeding Swine 
      Industry, South China Agricultural University, Guangzhou, China; National and 
      regional Joint Engineering Laboratory for Medicament of Zoonosis Prevention and 
      Control, China. Electronic address: guihongzh@scau.edu.cn.
LA  - eng
PT  - Journal Article
DEP - 20180407
PL  - Netherlands
TA  - Vet Microbiol
JT  - Veterinary microbiology
JID - 7705469
RN  - 0 (Nucleocapsid Proteins)
RN  - 130068-27-8 (Interleukin-10)
RN  - 452VLY9402 (Serine)
SB  - IM
MH  - Animals
MH  - Chromatography, Liquid
MH  - Down-Regulation
MH  - Interleukin-10/genetics
MH  - Kinetics
MH  - Macrophages, Alveolar/virology
MH  - Mutation
MH  - Nucleocapsid Proteins/chemistry/*genetics/isolation & purification/*metabolism
MH  - Phosphorylation
MH  - Porcine Reproductive and Respiratory Syndrome/virology
MH  - Porcine respiratory and reproductive syndrome virus/*genetics/metabolism
MH  - Serine/genetics/*metabolism
MH  - Sf9 Cells
MH  - Swine
MH  - Tandem Mass Spectrometry
MH  - Virus Replication
PMC - PMC7117435
OTO - NOTNLM
OT  - N protein
OT  - PRRSV
OT  - Phosphorylation
EDAT- 2018/05/21 06:00
MHDA- 2018/11/21 06:00
PMCR- 2018/04/07
CRDT- 2018/05/21 06:00
PHST- 2018/02/17 00:00 [received]
PHST- 2018/04/04 00:00 [revised]
PHST- 2018/04/05 00:00 [accepted]
PHST- 2018/05/21 06:00 [entrez]
PHST- 2018/05/21 06:00 [pubmed]
PHST- 2018/11/21 06:00 [medline]
PHST- 2018/04/07 00:00 [pmc-release]
AID - S0378-1135(18)30209-8 [pii]
AID - 10.1016/j.vetmic.2018.04.010 [doi]
PST - ppublish
SO  - Vet Microbiol. 2018 Jun;219:128-135. doi: 10.1016/j.vetmic.2018.04.010. Epub 2018 
      Apr 7.