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bioRxiv · 10.1101/2025.06.11.659102

Processing and release of the maize phytocytokine Zip1

Koenig, M.·Sorger, Z.·Mantz, M.·Perrar, A.·Huesgen, P. F.·Stael, S.·Villamil, J. C. M.·Doehlemann, G.
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Abstract

Phytocytokines are endogenous peptides that modulate plant immunity outcomes, yet how their maturation and spatial deployment are controlled remains unclear. Here we show that the maize phytocytokine precursor PROZIP1 is controlled by a spatially separated, two-stage proteolytic pathway that mechanistically uncouples signal activation from extracellular attenuation. PROZIP1 associates with the endoplasmic reticulum and undergoes intracellular, arginine-dependent processing by type II metacaspases, generating a C-terminal PROZIP1 fragment (Ct-PROZIP1). This processing licenses PROZIP1 for export to the apoplast via an ER-Golgi-independent route. Proteomic mapping and mutational analyses identify arginine residues flanking the Zip1 peptide as critical for efficient processing and secretion. The calcium-dependent metacaspase ZmMC9 specifically processes PROZIP1, thereby efficiently generating the bioactive Ct-PROZIP1 fragment. In the apoplast, Ct-PROZIP1 is further processed by papain-like cysteine proteases and additional extracellular proteases, contributing to Zip1 turnover and signal clearance. While the free Zip1 peptide is detected at later stages, Ct-PROZIP appears to be the primary signaling entity in modulating pathogen-induced immune responses. Together, these findings demonstrate a previously unknown complexity in peptide signaling, suggesting a multilayered control of phytocytokine activity that provides spatial and temporal precision to disease modulation in maize.

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Koenig, M., Sorger, Z., Mantz, M., Perrar, A., Huesgen, P. F., Stael, S., Villamil, J. C. M., Doehlemann, G.. 2025-06-17. Processing and release of the maize phytocytokine Zip1. https://doi.org/10.1101/2025.06.11.659102

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