1
Scientific RepoRts | (2018) 8:4526 | DOI:10.1038/s41598-018-22778-9
www.nature.com/scientificreports
Hsc70-2 is required for Beet black
scorch virus infection through
interaction with replication and
capsid proteins
Xiaoling Wang
1
, Xiuling Cao
1
, Min Liu
1
, Ruiqi Zhang
1
, Xin Zhang
1
, Zongyu Gao
1
, Xiaofei Zhao
1
,
Kai Xu
2
, Dawei Li
1
& Yongliang Zhang
1
Dissecting the complex molecular interplay between the host plant and invading virus improves our
understanding of the mechanisms underlying viral pathogenesis. In this study, immunoprecipitation
together with the mass spectrometry analysis revealed that the heat shock protein 70 (Hsp70) family
homolog, Hsc70-2, was co-puried with beet black scorch virus (BBSV) replication protein p23 and
coat protein (CP), respectively. Further experiments demonstrated that Hsc70-2 interacts directly with
both p23 and CP, whereas there is no interaction between p23 and CP. Hsc70-2 expression is induced
slightly during BBSV infection of Nicotiana benthamiana, and overexpression of Hsc70-2 promotes
BBSV accumulation, while knockdown of Hsc70-2 in N. benthamiana leads to drastic reduction of
BBSV accumulation. Infection experiments revealed that CP negatively regulates BBSV replication,
which can be mitigated by overexpression of Hsc70-2. Further experiments indicate that CP impairs
the interaction between Hsc70-2 and p23 in a dose-dependent manner. Altogether, we provide
evidence that besides specic functions of Hsp70 family proteins in certain aspects of viral infection,
they can serve as a mediator for the orchestration of virus infection by interacting with dierent viral
components. Our results provide new insight into the role of Hsp70 family proteins in virus infection.
Beet black scorch virus (BBSV) is a positive-strand RNA virus belonging to the genus Betanecrovirus in the family
Tombusviridae
1
. BBSV genome consists of a 3644 nucleotides single-stranded RNA without a 5′ cap structure
nor 3′ polyA tail. e BBSV genome contains six open reading frames (ORFs). e 5′ proximal ORF encodes the
auxiliary replication protein p23, which induces the rearrangement of the ER to form virus replication complexes
(VRCs)
2
. e stop codon of p23 ORF is read-through to generate the RNA dependent RNA polymerase (RdRp)
protein, p82. p7a, p7b, and p5′ are translated from sub-genomic RNA1 (sgRNA1) and are responsible for viral
cell-to-cell movement. e CP ORF at the 3′ proximal region of the genome, is translated from sgRNA2. CP
expressed from sgRNA2 is not essential for cell-to-cell movement in Chenopodium amaranticolor, but is indispen-
sable for systemic infection in Nicotiana benthamiana
1,3,4
. In addition, BBSV virion assembly and virus systemic
movement require specic CP N-terminal basic amino acid and phosphorylation of the CP
5,6
.
Replication of positive-stranded RNA virus is a complicated but well-organized process, which involve the
coordinate functions of viral proteins and diverse host factors
7,8
. Because host factors involved in viral replica-
tion represent potential targets for virus control, identication and functional characterization of their roles is
one of the hot topics in virus research. Both auxiliary replicase protein and RNA-dependent RNA polymerase
(RdRp) are the key components of VRC and have potential to be baits for identifying host interactors. However,
molecular weight of auxiliary replicase protein is smaller than that of RdRp, which makes it relatively easy to be
manipulated for the subsequent biochemical analysis. More importantly, in contrast to RdRp with conned func-
tion in viral RNA synthesis, auxiliary replicase proteins, such as tomato bushy stunt virus (TBSV) p33
9
, cucumber
1
State Key Laboratory of Agro-Biotechnology and Ministry of Agriculture Key Laboratory of Soil Microbiology,
College of Biological Sciences, China Agricultural University, Beijing, 100193, P. R. China.
2
Jiangsu Key Laboratory for
Microbes and Functional Genomics, Jiangsu Engineering and Technology Research Center for Microbiology, College
of Life Sciences, Nanjing Normal University, Nanjing, 210046, P. R. China. Xiaoling Wang and Xiuling Cao contributed
equally to this work. Correspondence and requests for materials should be addressed to Y.Z. (email: cauzhangyl@
cau.edu.cn)
Received: 29 December 2017
Accepted: 27 February 2018
Published: xx xx xxxx
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