Functional analysis of apple stem pitting virus coat protein variants
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RESEARCH
Open Access
Functional analysis of apple stem pitting virus coat protein variants Xiaofang Ma1,2,3,4, Ni Hong1,2, Peter Moffett3, Yijun Zhou4 and Guoping Wang1,2*
Abstract Background: Although the canonical function of viral coat protein (CP) is to encapsidate the viral genome, they have come to be recognized as multifunctional proteins, involved in almost every stage of the viral infection cycle. However, CP functions of Apple stem pitting virus (ASPV) has not been comprehensively documented. This study aimed to characterize the functions of ASPV CP and any functional diversification caused by sequence diversity of six ASPV CP variants and studied their biological, serological, pathogenic and viral suppressor of RNA silencing (VSR) functions. Methods: Six ASPV CP variants that have previously been shown to belong to different subgroups were selected here to study their diversity functions. Agrobacterium mediated infiltration (Agroinfiltration) was used to express YFP-ASPV-CPs in Nicotiana. benthamiana and infect Nicotiana. occidental with PVX-ASPV-CPs in. Confocal microscopy was used to detect YFP-ASPV-CPs florescence. CPs expressed in Escherichia coli BL21 (DE3) were induced by IPTG. Results: In this study, we showed that recombinant CPs expressed in Escherichia coli BL21 (DE3) had different levels of serological reactivity to three anti-ASPV antibodies used to detect ASPV. Furthermore, fusion CPs with YFP (YFP-CPs) expressed in N. benthamiana cells differed in their ability to form aggregates. We also showed that ASPV isolates that harbour these CPs induced different biological symptoms on its herbaceous host N. occidentalis. At the same time, we found that all six CPs when expressed in PVX vector showed similar VSR activity and produced similar symptoms in N. occidentalis, despite their differences in amino acids. Conclusions: Different ASPV isolates induced different symptoms in N. occidentalis, however, ASPV CP variants expressed in PVX vector showed the same symptoms in N. occidentalis plants. Also, we showed that ASPV CP variants has the same level of VSR activity, but they have different abilities to aggregate in N. benthamiana. Keywords: Apple stem pitting virus, Coat protein, CP variants, Aggregate, RNA silencing suppressor
Background Apple stem pitting virus (ASPV) is the type species of the Foveavirus genus in the Betaflexiviridae family [1]. It possesses a single stranded positive RNA (+ssRNA) genome comprising of approximately 9300 nucleotides (nts), which encodes five open reading frames (ORFs, ORF1-ORF5) as well as the 5′ untranslated region (UTR) and 3’ UTR. ORF1 encodes the viral RNAdependent RNA polymerase (RdRP), ORF2-ORF4 * Correspondence: [email protected] 1 State Key Laboratory of Agricultural Microbiology, Huazhong Agricultural University, Wuhan, Hubei 430070, People’s Republic of China 2 Key Laboratory of Plant Pathology of Hubei Province, College of Plant Science and Technology, Huazhong Agricultural University, Wuhan, Hubei 430070, People’s Republic of China Full list
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