Helper component-proteinase (HC-Pro) protein of Papaya ringspot virus interacts with papaya calreticulin.
Potyviral helper component-proteinase (HC-Pro) is a multifunctional protein involved in plant-virus interactions. In this study, we constructed a Carica papaya L. plant cDNA library to investigate the host factors interacting with Papaya ringspot virus (PRSV) HC-Pro using a Sos recruitment two-hybrid system (SRS). We confirmed that the full-length papaya calreticulin, designated PaCRT (GenBank accession no. FJ913889), interacts specifically with PRSV HC-Pro in yeast, in vitro and in plant cells using SRS, in vitro protein-binding assay and bimolecular fluorescent complementation assay, respectively. SRS analysis of the interaction between three PaCRT deletion mutants and PRSV HC-Pro demonstrated that the C-domain (residues 307-422), with a high Ca(2+)-binding capacity, was responsible for binding to PRSV HC-Pro. In addition, quantitative real-time reverse transcriptase-polymerase chain reaction assay showed that the expression of PaCRT mRNA was significantly upregulated in the primary stage of PRSV infection, and decreased to near-basal expression levels in noninoculated (healthy) papaya plants with virus accumulation inside host cells. PaCRT is a new calcium-binding protein that interacts with potyviral HC-Pro. It is proposed that the upregulated expression of PaCRT mRNA may be an early defence-related response to PRSV infection in the host plant, and that interaction between PRSV HC-Pro and PaCRT may be involved in plant calcium signalling pathways which could interfere with virus infection or host defence.
Shen
Wentao
W
Key Biotechnology Laboratory for Tropical Crops, Ministry of Agriculture, Institute of Tropical Bioscience and Biotechnology (ITBB), Chinese Academy of Tropical Agriculture Sciences (CATAS), 4 Xueyuan Road, Haikou, 571101, China.
Yan
Pu
P
Gao
Le
L
Pan
Xueying
X
Wu
Jinyan
J
Zhou
Peng
P
D000595:Amino Acid Sequence; D037282:Calreticulin / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D029441:Carica / Q000235:genetics / Q000378:metabolism / Q000821:virology*; D003546:Cysteine Endopeptidases / Q000378:metabolism*; D018506:Gene Expression Regulation, Plant; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017800:Potyvirus / Q000378:metabolism*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000378:metabolism; D016415:Sequence Alignment; D020798:Two-Hybrid System Techniques; D015854:Up-Regulation / Q000235:genetics; D014764:Viral Proteins / Q000378:metabolism*
D037282:Calreticulin; D050505:Mutant Proteins; D012333:RNA, Messenger; D014764:Viral Proteins; D003546:Cysteine Endopeptidases; C072775:HC-Pro protein, potyvirus
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