Analysis of the maize cytokinin receptor Zea mays Histidine Kinase 1 function using Saccharomyces cerevisiae
Cytokinins (CK) regulate a diverse assortment of processes in plants, including cellular division, biosynthesis of chloroplasts, and differentiation within root and apical meristems. Response to CK is regulated through a two-component signal transduction system consisting of a receptor and a response regulator. Two-component signaling systems are highly conserved in bacteria, fungi and plants and allow organisms to sense and respond to external and internal stimuli. Our analysis of the semi-dominant, leaf patterning maize mutant Hairy Sheath Frayed1 (Hsf1) identified the maize CK receptor Zea mays Histidine Kinase1 (ZmHK1) as the underlying gene. The Hsf1 phenotype is marked by the outgrowth of proximal leaf tissue (sheath, auricle and ligule) in the distal leaf blade, reduced leaf size, and increased leaf pubescence. Missense mutations in the CK binding domain of ZmHK1 increase ligand binding affinity, resulting in CK hypersignaling and giving rise to altered leaf patterning in Hsf1. We are using a two-component signaling assay in Saccharomyces cerevisiae to understand the relationship between these amino acid changes and altered ZmHK1 activity. We have assayed the three independent Hsf1 alleles (Hsf1-1595, Hsf1-1603, and Hsf1-AEWL) using the yeast system and found some signal in the absence of added CK. We are making additional targeted amino acid changes near the CK binding domain in ZmHK1 to determine which residues are critical for ligand recognition, binding and signaling. Our current results will be presented.