PMID- 33127640
OWN - NLM
STAT- MEDLINE
DCOM- 20210325
LR  - 20240407
IS  - 1083-351X (Electronic)
IS  - 0021-9258 (Print)
IS  - 0021-9258 (Linking)
VI  - 295
IP  - 52
DP  - 2020 Dec 25
TI  - Molecular characterization of the RNA-protein complex directing -2/-1 programmed 
      ribosomal frameshifting during arterivirus replicase expression.
PG  - 17904-17921
LID - S0021-9258(17)50671-7 [pii]
LID - 10.1074/jbc.RA120.016105 [doi]
AB  - Programmed ribosomal frameshifting (PRF) is a mechanism used by arteriviruses 
      like porcine reproductive and respiratory syndrome virus (PRRSV) to generate 
      multiple proteins from overlapping reading frames within its RNA genome. PRRSV 
      employs -1 PRF directed by RNA secondary and tertiary structures within its viral 
      genome (canonical PRF), as well as a noncanonical -1 and -2 PRF that are 
      stimulated by the interactions of PRRSV nonstructural protein 1beta (nsp1beta) and host 
      protein poly(C)-binding protein (PCBP) 1 or 2 with the viral genome. Together, 
      nsp1beta and one of the PCBPs act as transactivators that bind a C-rich motif near 
      the shift site to stimulate -1 and -2 PRF, thereby enabling the ribosome to 
      generate two frameshift products that are implicated in viral immune evasion. How 
      nsp1beta and PCBP associate with the viral RNA genome remains unclear. Here, we 
      describe the purification of the nsp1beta:PCBP2:viral RNA complex on a scale 
      sufficient for structural analysis using small-angle X-ray scattering and 
      stochiometric analysis by analytical ultracentrifugation. The proteins associate 
      with the RNA C-rich motif as a 1:1:1 complex. The monomeric form of nsp1beta within 
      the complex differs from previously reported homodimer identified by X-ray 
      crystallography. Functional analysis of the complex via mutational analysis 
      combined with RNA-binding assays and cell-based frameshifting reporter assays 
      reveal a number of key residues within nsp1beta and PCBP2 that are involved in 
      complex formation and function. Our results suggest that nsp1beta and PCBP2 both 
      interact directly with viral RNA during formation of the complex to coordinate 
      this unusual PRF mechanism.
CI  - (c) 2020 Patel et al.
FAU - Patel, Ankoor
AU  - Patel A
AD  - Department of Microbiology, University of Manitoba, Winnipeg, Manitoba, Canada.
FAU - Treffers, Emmely E
AU  - Treffers EE
AD  - Molecular Virology Laboratory, Department of Medical Microbiology, Leiden 
      University Medical Center, Leiden, The Netherlands.
FAU - Meier, Markus
AU  - Meier M
AD  - Department of Chemistry, University of Manitoba, Winnipeg, Manitoba, Canada.
FAU - Patel, Trushar R
AU  - Patel TR
AD  - Alberta RNA Research and Training Institute, Department of Chemistry and 
      Biochemistry, University of Lethbridge, Lethbridge, Alberta, Canada.
FAU - Stetefeld, Jorg
AU  - Stetefeld J
AD  - Department of Chemistry, University of Manitoba, Winnipeg, Manitoba, Canada.
FAU - Snijder, Eric J
AU  - Snijder EJ
AD  - Molecular Virology Laboratory, Department of Medical Microbiology, Leiden 
      University Medical Center, Leiden, The Netherlands.
FAU - Mark, Brian L
AU  - Mark BL
AD  - Department of Microbiology, University of Manitoba, Winnipeg, Manitoba, Canada. 
      Electronic address: brian.mark@umanitoba.ca.
LA  - eng
SI  - PDB/2P2R
SI  - PDB/2JZX
SI  - PDB/2PQU
SI  - PDB/3MTV
SI  - PDB/2PY9
GR  - 201610PJT-152935/CIHR/Canada
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
DEP - 20201030
PL  - United States
TA  - J Biol Chem
JT  - The Journal of biological chemistry
JID - 2985121R
RN  - 0 (DNA-Binding Proteins)
RN  - 0 (PCBP1 protein, human)
RN  - 0 (RNA, Viral)
RN  - 0 (RNA-Binding Proteins)
RN  - 0 (Viral Nonstructural Proteins)
SB  - IM
MH  - Animals
MH  - DNA-Binding Proteins/genetics/*metabolism
MH  - Frameshifting, Ribosomal/*physiology
MH  - Host-Pathogen Interactions/*immunology
MH  - Humans
MH  - Immune Evasion
MH  - Porcine Reproductive and Respiratory Syndrome/immunology/*virology
MH  - Porcine respiratory and reproductive syndrome virus/*physiology
MH  - RNA, Viral
MH  - RNA-Binding Proteins/genetics/*metabolism
MH  - Swine
MH  - Viral Nonstructural Proteins/genetics/*metabolism
MH  - *Virus Replication
PMC - PMC7939443
OTO - NOTNLM
OT  - PCBP
OT  - PCPB2
OT  - PRRSV
OT  - RNA
OT  - RNA virus
OT  - nidovirus
OT  - nonstructural protein 1
OT  - nsp1
OT  - nsp1beta
OT  - poly(C)-binding protein
OT  - ribosome
OT  - ribosome function
OT  - sedimentation velocity
OT  - small-angle X-ray scattering
OT  - structural biology
OT  - translation
OT  - viral replication
OT  - virology
OT  - beta
COIS- Conflict of interest-E. J. S. and E. E. T. have a patent that relates to aspects 
      of this work (patent number: US9623103).
EDAT- 2020/11/01 06:00
MHDA- 2021/03/26 06:00
PMCR- 2021/01/13
CRDT- 2020/10/31 05:29
PHST- 2020/09/18 00:00 [received]
PHST- 2020/10/22 00:00 [revised]
PHST- 2020/11/01 06:00 [pubmed]
PHST- 2021/03/26 06:00 [medline]
PHST- 2020/10/31 05:29 [entrez]
PHST- 2021/01/13 00:00 [pmc-release]
AID - S0021-9258(17)50671-7 [pii]
AID - 10.1074/jbc.RA120.016105 [doi]
PST - ppublish
SO  - J Biol Chem. 2020 Dec 25;295(52):17904-17921. doi: 10.1074/jbc.RA120.016105. Epub 
      2020 Oct 30.