Virus Research 138 (2008) 119–129
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Virus Research
journal homepage: www.elsevier.com/locate/virusres
Cauliflower mosaic virus gene VI product N-terminus contains regions
involved in resistance-breakage, self-association and interactions with
movement protein
Michael Hapiak
a
, Yongzhong Li
1
, Keli Agama
2
, Shaddy Swade
a
, Genevieve Okenka
a
, Jessica Falk
a
,
Sushant Khandekar
a
, Gaurav Raikhy
a
, Alisha Anderson
a
, Justin Pollock
a
, Wendy Zellner
a
,
James Schoelz
b
, Scott M. Leisner
a,∗
a
Department of Biological Sciences, The University of Toledo, Toledo, OH 43606, United States
b
Plant Science Unit, College of Agriculture, Food, and Natural Resources, University of Missouri-Columbia, Columbia, MO 65211, United States
article info
Article history:
Received 17 June 2008
Received in revised form
15 September 2008
Accepted 15 September 2008
Available online 28 October 2008
Keywords:
CaMV
P6
TAV
P1
Arabidopsis
Inclusion body protein
abstract
Cauliflower mosaic virus (CaMV) gene VI encodes a multifunctional protein (P6) involved in the translation
of viral RNA, the formation of inclusion bodies, and the determination of host range. Arabidopsis thaliana
ecotype Tsu-0 prevents the systemic spread of most CaMV isolates, including CM1841. However, CaMV
isolate W260 overcomes this resistance. In this paper, the N-terminal 110 amino acids of P6 (termed D1)
were identified as the resistance-breaking region. D1 also bound full-length P6. Furthermore, binding
of W260 D1 to P6 induced higher -galactosidase activity and better leucine-independent growth in
the yeast two-hybrid system than its CM1841 counterpart. Thus, W260 may evade Tsu-0 resistance by
mediating P6 self-association in a manner different from that of CM1841. Because Tsu-0 resistance pre-
vents virus movement, interaction of P6 with P1 (CaMV movement protein) was investigated. Both yeast
two-hybrid analyses and maltose-binding protein pull-down experiments show that P6 interacts with P1.
Although neither half of P1 interacts with P6, the N-terminus of P6 binds P1. Interestingly, D1 by itself
does not interact with P1, indicating that different portions of the P6 N-terminus are involved in different
activities. The P1–P6 interactions suggest a role for P6 in virus transport, possibly by regulating P1 tubule
formation or the assembly of movement complexes.
© 2008 Elsevier B.V. All rights reserved.
1. Introduction
Plant virus resistance is mediated at multiple levels includ-
ing: elicitation of active defense responses involving programmed
cell death (a hypersensitive response), elicitation of active defense
responses without necrosis, and the degradation of viral RNAs by
post-transcriptional gene silencing (Goodman and Novacky, 1994;
Goodrick et al., 1991; Hull, 2002; Waterhouse et al., 1999). Although
plant resistance can be effective in limiting viruses to the inoc-
ulated leaves or even individual cells, certain strains are able to
infect even these protected hosts (Meshi et al., 1988, 1989; Padgett
and Beachy, 1993). Plant viruses may overcome resistance by a
∗
Corresponding author.
E-mail address: sleisne@uoft02.utoledo.edu (S.M. Leisner).
1
Now at: Department of Medicine, University of Illinois at Chicago, Chicago, IL
60612, United States.
2
Now at: Laboratory of Molecular Pharmacology, Center for Cancer Research,
National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, United
States.
passive means, i.e., they are not recognized by the host. For exam-
ple, passive resistance-breakage was observed in the relationship
between specific Cauliflower mosaic virus (CaMV) isolates and Ara-
bidopsis thaliana ecotypes (Agama et al., 2002). While certain A.
thaliana ecotypes, such as Col-0, are susceptible to many isolates
of CaMV others, such as Tsu-0, are resistant to many viral iso-
lates, such as CM1841 (Balazs and Lebeurier, 1981; Leisner and
Howell, 1992; Melcher, 1989). Tsu-0 resistance prevents systemic
spread of CaMV, whereas isolate W260 evades detection by the Tsu-
0 resistance machinery, which allows this CaMV strain to invade
the host systemically (Agama et al., 2002). Analysis of chimeric
DNA genomes generated from W260 and CM1841 localized the
resistance-breaking determinant to the region of gene VI encod-
ing the N-terminal 184 amino acid residues (termed RBR-1), of the
protein product (P6).
In addition to its role as a host range determinant, P6 is involved
in many activities including regulating translation of viral pro-
teins and forming the characteristic amorphous inclusion bodies
observed in the cytoplasm of infected cells (Bonneville et al., 1989;
Covey and Hull, 1981; De Tapia et al., 1993; Schoelz et al., 1986;
0168-1702/$ – see front matter © 2008 Elsevier B.V. All rights reserved.
doi:10.1016/j.virusres.2008.09.002