Phosphorylation in stomatal vacuole fusion and tar

Control of stomatal opening and closing is essential for the uptake of CO2 while mitigating water vapor loss in plants. The opening of stomata is regulated in part by homotypic vacuole fusion, which is mediated by conserved homotypic vacuole protein sorting (HOPS) and vacuolar SNARE (soluble N-ethylmaleimide sensitive factor attachment protein receptors) complexes. HOPS-specific subunits VACUOLE PROTEIN SORTING39 (VPS39) and VPS41 are required for homotypic plant vacuole fusion, and a computational model predicted that post-translational modifications of HOPS may be needed for plant stomatal vacuole fusion. We recently characterized a viable T-DNA insertion allele of VPS39 that demonstrated a critical role of VPS39 in stomatal vacuole fusion. We also detected changes in VPS39 phosphorylation in stomata that indicate that VPS39 function in stomata and embryonic development requires dynamic changes in phosphorylation. I will also describe our efforts to use targeted protein degradation to characterize the function of essential proteins such as VPS39. The recently developed E3 DART system for protein ubiquitination is based on the specific and well-characterized interaction between a non-plant E3 ligase and its target. This ligase-target interaction is modular, which makes it ideal for targeted degradation of chimeric protein fusions. A specific domain of the target, or “degron”, is sufficient to target a chimeric protein for degradation. I will describe potential applications of E3 DART in plant research and biotech.