Requirement for HC-Pro processing during genome amplification of tobacco etch potyvirus

Kristin D. Kasschau, James C. Carrington

Research output: Contribution to journalArticlepeer-review

11 Scopus citations


The helper component-proteinase (HC-Pro) of tobacco etch potyvirus (TEV) is a multifunctional protein with several known activities. The N-terminal region is required for aphid transmission and efficient genome amplification, the central region is required for long-distance movement in plants, and the C-terminal domain is a cysteine-type proteinase that autocatalytically cleaves between itself and the P3 protein. To investigate the requirement for HC-Pro-mediated proteolysis during viral replication, a variety of mutations resulting in amino acid substitutions and insertions in the proteolytic domain and cleavage site motif were introduced into the vital genome. Mutations affecting the active site residues, His722 and Cys649, or the cleavage site P1' residue, Gly764, inhibited proteolytic activity of HC-Pro. Mutant genomes containing these modifications were amplification-defective in protoplasts and plants. Mutants with substitutions affecting several conserved, but non-active site, residues (Ser610, Cys694, and Asp715) within the proteinase domain encoded active HC-Pro proteinases and were similar to parental virus in protoplasts and plants. To determine if the replication defect of the proteinase-debilitated mutants was due to inactivation of HC- Pro proteolytic activity or simply to the inability of HC-Pro and P3 protein to separate, a sequence coding for a heterologous cleavage site recognized by the TEV Nla proteinase was inserted between the HC-Pro and P3 coding regions of an active site mutant. This cleavage site was functional in vitro using purified Nla proteinase. However, this modification was insufficient to restore amplification activity to the mutant. In addition, the active site mutant was not complemented by wild-type HC-Pro supplied in trans by transgenic plants. These results suggest that an active HC-Pro proteinase is required in cis for TEV genome amplification.

Original languageEnglish
Article number71254
Pages (from-to)268-273
Number of pages6
Issue number1
StatePublished - May 10 1995


Dive into the research topics of 'Requirement for HC-Pro processing during genome amplification of tobacco etch potyvirus'. Together they form a unique fingerprint.

Cite this