PROJECT SUMMARY Peroxisome Biogenesis, Dynamics, and Degradation Peroxisomes are eukaryotic organelles that are essential for life in plants and metazoans. Peroxisomes sequester various oxidative reactions, thereby improving metabolic efficiency while protecting cytosolic constituents from oxidative damage. Although our knowledge about peroxisome function and dysfunction is increasing, mechanistic understanding of how these critical organelles are formed, maintained, and turned over remains incomplete. The proposed studies aim to fill these gaps by answering the following questions: How do pre-peroxisomes originate? How do peroxisomal membrane complexities develop? How can chemical probes and genetic suppressors modulate peroxisome function? How are obsolete or damaged peroxisomes targeted for turnover? Does the ubiquitination machinery on the peroxisomal membrane moonlight in tasks beyond receptor recycling? These questions will be addressed using Arabidopsis thaliana; the peroxisomal functions, small size, and facile genetics of this model plant allow straightforward impairment and enhancement of peroxisomal processes in an intact multicellular organism. Moreover, the relatively large size of plant peroxisomes (compared to yeast and mammalian peroxisomes) offers unique opportunities to decipher peroxisome biogenesis and membrane intricacies using live-cell imaging. The proposed studies also will train the next generation of scientists in state-of-the-art genetic, biochemical, and cell biological approaches. Peroxisomal defects underlie the peroxisome biogenesis disorders, a group of inherited recessive syndromes that are generally fatal in infancy or childhood and are characterized by diverse symptoms including poor growth, multi-organ dysfunctions, hearing and vision loss, and psychomotor retardation. Peroxisome dysfunction also contributes to common age-related diseases (e.g., neurodegeneration, type 2 diabetes) that are exacerbated by oxidative stress. The proposed studies will exploit unique aspects of plant peroxisomes while taking advantage of knowledge from fungal and mammalian systems to provide insights that are likely to apply throughout eukaryotes. Continued development of evolutionarily distinct systems with unique advantages for elucidating peroxisome biology will advance hypotheses and mechanistic models to expand and refine our understanding of this essential organelle.