Picornavirus Modification of a Host mRNA Decay Protein
Janet M. Rozovics,a,bAmanda J. Chase,a,bAndrea L. Cathcart,a,bWayne Chou,b,cPaul D. Gershon,b,cSaiprasad Palusa,d
Jeffrey Wilusz,dand Bert L. Semlera,b
Department of Microbiology and Molecular Genetics, School of Medicine, University of California, Irvine, California, USAa; Center for Virus Research, University of California,
Irvine, California, USAb; Department of Molecular Biology and Biochemistry, University of California, Irvine, California, USAc; and Department of Microbiology, Immunology
and Pathology, Colorado State University, Fort Collins, Colorado, USAd
shown to cleave all four isoforms of recombinant AUF1 at a specific N-terminal site in vitro. Finally, endogenous AUF1 was
Received 10 October 2012 Accepted 12 October 2012 Published 6 November 2012
Citation Rozovics JM, et al. 2012. Picornavirus modification of a host mRNA decay protein. mBio 3(6):e00431-12. doi:10.1128/mBio.00431-12.
Editor Anne Moscona, Weill Medical College-Cornell
Copyright © 2012 Rozovics et al. This is an open-access article distributed under the terms of the Creative Commons Attribution-Noncommercial-Share Alike 3.0 Unported
License, which permits unrestricted noncommercial use, distribution, and reproduction in any medium, provided the original author and source are credited.
Address correspondence to Bert L. Semler, firstname.lastname@example.org.
in picornavirus translation and RNA replication (for a review, see
references 1 and 2). Among these, members of the heterogeneous
ribonucleoprotein (hnRNP) family of proteins, including
hnRNP K, have been reported to have essential roles in picorna-
virus replication (3–16).
In addition to usurping the functions of host proteins during
infection, picornaviruses also modify host proteins via viral enzy-
shuts off host cell cap-dependent translation, thereby eliminating
liovirus proteinase 3C/3CD cleavage of PCBP2, a cellular RNA
binding protein required for poliovirus translation and RNA rep-
lication, is somewhat unique since cleavage of PCBP2 does not
simply disrupt cellular processes but may influence template se-
lection for RNA replication versus viral translation (18).
characteristics typical of an hnRNP, including nucleocytoplasmic
iven the relative genetic simplicity of their positive-sense,
single-stranded genomic RNAs, picornaviruses have evolved
shuttling properties and multiple isoforms (for a review, see ref-
These isoforms result from alternative splicing of a single mRNA
(20–22). Of the four AUF1 isoforms, p45 is the largest and is the
canonical form to which the other isoforms are compared (for a
review, see references 19 and 20). All four isoforms have a
dimerization domain (encoded in exon 1), as well as two RNA
AUF1 binds specific proto-oncogene and cytokine mRNAs con-
taining AU-rich element (ARE) sequences, such as c-myc (23),
which are then targeted for decay; all four isoforms have been
reported to have different roles in this process (22, 24). AUF1 has
also been reported to have a role in telomere binding (25) and
transcriptional activation (26, 27).
Much like other hnRNPs, AUF1 has been reported to have a
role in the replication cycles of different viruses. It is required for
efficient hepatitis C virus translation (28) and has been shown to
regulate the C promoter in Epstein-Barr virus (27). For picorna-
viruses, there is one report demonstrating an increase in AUF1
levels in the cytoplasm of primary cultures of human airway epi-
thelial cells infected with human rhinovirus 16 (HRV16) (29).
Although direct evidence for a role for AUF1 in picornavirus in-
fections is currently lacking, this host protein does interact with
November/December 2012 Volume 3 Issue 6 e00431-12
several proteins that have been reported to function during repli-
cation of poliovirus and other picornaviruses. These include
poly(A) binding protein (PABP) (30), nucleolin (21), and
PCBP1/2 (31). In uninfected cells, these interactions are thought
to facilitate a role in regulation of mRNA decay (30), transcrip-
tional activation (21), and the ?-globin mRNA stability complex
the genome via interaction with PCBP (32, 33). Nucleolin inter-
and has been observed to relocalize to the cytoplasm during po-
significantly decreases virus production (34). PCBP1/2 proteins
interact with the 5= NCR of poliovirus and are required for viral
translation and RNA replication (3–11).
In this paper, we describe data that suggest that AUF1 is a
player in picornavirus infection. We report modification of this
observed that all four isoforms of AUF1
are cleaved during infection with poliovi-
rus, HRV14, or HRV16 in HeLa cells. Us-
ing in vitro proteinase assays with recom-
binant enzymes and substrates, we
present evidence identifying the primary
cleavage site in all four isoforms that is
cleaved by viral proteinase 3CD. We have
5= NCR of poliovirus or human rhinovi-
rus 16 genomes in vitro. In addition,
sites adjacent to (but distinct from) puta-
ing poliovirus infection. Taken together,
these data strongly support a role for
or human rhinovirus infection. Given
the roles that different classes of RNA
ferent picornaviruses, we initially exam-
ined possible alterations of proteins in-
volved in cellular mRNA decay during
picornavirus infection. Cytoplasmic S10
lysates were generated from poliovirus-
protein modifications. Although AUF1,
HuR, Xrn1, Dcp2, and nucleophosmin
were explored via Western blot analysis,
AUF1 was the only protein observed to
TABLE 1 Examples of cellular proteins cleaved during picornavirus infectionsb
Cellular protein(s)Picornavirus proteinase(s)a
Effect(s) of cleavage
eIF4GI, eIF4GII, eIF4A
TBP, CREB, TFIIIC, Oct-1
PV 2A, PV 3C, HRV14 3C, CVB 2A
PV 2A, CVB 2A, FMDV 3C, L
PV 3C, 3CD
PV 3C, 3CD
Disrupts interaction with eIF4G, inhibits host cell translation
Disrupts cap-binding complex, shuts off host cell translation
Possible template selection for viral translation versus RNA replication
Induces morphological changes in host cells
Inhibits host cell RNA synthesis
Redistributes La to cytoplasm to enhance translation of viral mRNA
Regulates template selection for viral translation versus RNA replication
Possible role in P body disruption and RNA stability
aPV, poliovirus; FMDV, foot-and-mouth disease virus; CVB, coxsackievirus B; HRV, human rhinovirus; TBP, TATA-binding protein; references for cleavage events include
references 18, 58, and 65 to 79.
bAdapted from reference 18.
FIG 1 AUF1 exon structure schematic and proposed 3CD cleavage site. The figure shows the exon
structure of the human AUF1 gene and the different protein isoforms (p37, p40, p42, and p45) gener-
ated by alternative splicing. The RNA recognition motifs (RRM1 and RRM2), the Q-rich domain, and
sequence in exon 1 starting with amino acid residue 29 and extending through residue 42. This region
contains a putative picornavirus 3CD cleavage recognition site with the P1 and P1= positions Q-G
the experimental results displayed in Fig. 3, the Q-G pair was mutagenized to I-D (highlighted in red).
This amino acid pair is predicted not to be cleaved by the poliovirus or human rhinovirus 3CD protei-
nase. Adapted with data from the work of Gratacos and Brewer (19).
Rozovics et al.
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Cleavage of AUF1 during Picornavirus Infection
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