Role of Calcium-Dependent Kinases in Arabidopsis Root Hair Responses to Mechanical Forces

Plants increasingly struggle with mechanical stress in compacted soils, a growing problem amplified by climate change. Although we know root hairs are essential for absorbing water and nutrients, the exact molecular mechanisms enabling them to sense soil pressure have remained mostly unclear. In my master's research, I aimed to discover how specific calcium-dependent protein kinases (CPKs) namely CPK4 and CPK11 and the protein ANNEXIN1 influence the responses of Arabidopsis thaliana root hairs to mechanical stress. To accurately study these delicate cellular processes, I developed two highly specific root-hair isolation methods: Cryo-Mechanical Disruption and freeze–thaw lysis, that consistently achieved over 90% hair RNA purity. Using knockout mutants (cpk4, cpk11, annexin1) and agar gradients simulating soil compaction (0.5%, 0.8%, 1%, 1.25%, 1.5% Agar Concentration), I found distinct responses: cpk4 hairs showed early stress sensitivity but adapted at higher pressures; cpk11 initially outperformed wild-type but sharply declined under severe stress; ANNEXIN1 was critical, with knockout lines almost completely halting hair growth at high stress. qPCR analysis revealed compensatory gene expression between CPK4 and CPK11, and similar responses in rice highlighted evolutionary conservation. These findings enhance our understanding of root hair mechanosensory systems and provide valuable insights for developing stress-resilient crops.