A Novel Cysteine Protease from Phytolacca americana Cleaves Pokeweed Antiviral Protein Generating Bioactive Fragments
Abstract
1. Introduction
2. Results
2.1. Identification of PAP Interactors
2.2. Characterization of PaCP1 Enzyme Activity
2.3. Support for PAP-PaCP1 Interaction
2.4. Co-Localization of PAP and PaCP1
2.5. PaCP1-Mediated Cleavage of PAP
2.6. Analysis of PAP Cleavage Products
2.7. Activation of MAPK Signaling by the Cleaved Peptides
3. Discussion
4. Materials and Methods
4.1. Plant Materials and Growth Conditions
4.2. Total Protein Isolation
4.3. Immunoprecipitation Assay and Mass Spectrometry
4.4. Bioinformatic and Structural Analyses of PaCP1
4.5. Plasmid Construction
4.6. Isolation of His-Tagged PaCP1
4.7. In Vitro Enzyme Activity Assay
4.8. Yeast-Two Hybrid Assay
4.9. Microscopy
4.10. Apoplastic Fluid Extraction
4.11. In Vitro Cleavage Assays
4.12. Immunoblotting
4.13. Peptide Treatment
4.14. Isolation of Ribosomes and Depurination Assay
Supplementary Materials
Author Contributions
Funding
Data Availability Statement
Acknowledgments
Conflicts of Interest
Abbreviations
AF | Apoplastic Fluid |
AtXCP1 | Arabidopsis thaliana xylem cysteine protease 1 |
MAPK | Mitogen activated protein kinase |
PaCP1 | Phytolacca americana cysteine protease 1 |
PAP | Pokeweed antiviral protein |
PLCP | Papain-like cysteine protease |
RFU | Relative fluorescence units |
RIP | Ribosome inactivating protein |
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Audet, A.; Chivers, J.A.; Hudak, K.A. A Novel Cysteine Protease from Phytolacca americana Cleaves Pokeweed Antiviral Protein Generating Bioactive Fragments. Plants 2025, 14, 2441. https://doi.org/10.3390/plants14152441
Audet A, Chivers JA, Hudak KA. A Novel Cysteine Protease from Phytolacca americana Cleaves Pokeweed Antiviral Protein Generating Bioactive Fragments. Plants. 2025; 14(15):2441. https://doi.org/10.3390/plants14152441
Chicago/Turabian StyleAudet, Annabelle, Jennifer A. Chivers, and Katalin A. Hudak. 2025. "A Novel Cysteine Protease from Phytolacca americana Cleaves Pokeweed Antiviral Protein Generating Bioactive Fragments" Plants 14, no. 15: 2441. https://doi.org/10.3390/plants14152441
APA StyleAudet, A., Chivers, J. A., & Hudak, K. A. (2025). A Novel Cysteine Protease from Phytolacca americana Cleaves Pokeweed Antiviral Protein Generating Bioactive Fragments. Plants, 14(15), 2441. https://doi.org/10.3390/plants14152441