Reed College BA Biology, 1973
University of Washington Ph.D. Plant Biology, 1980
Organelle biogenesis in eucaryotes depends on the import of nuclear-encoded proteins from the cytosol. The chloroplast, the photosynthetic organelle in plants, imports at least 90% of its proteins, which are typically synthesized as precursors with an N-terminal extension, a transit peptide, that mediates multiple steps as proteins are targeted to the correct organelle, cross a defining membrane, and undergo proteolytically processing to reach their mature, functional form. We have identified a general stromal processing peptidase, SPP, responsible for transit peptide removal, a critical step in the import pathway. SPP is a member of a metallopeptidase family that depends on a zinc-binding motif for its activity. Our studies have explored the specificity of the protein-protein interactions that occur as SPP recognizes and cleaves the transit peptide. Downregulation of SPP in transgenic Arabidopsis yields many seedling lethals, demonstrating that SPP is encoded by an essential gene. Low levels of SPP severely affect other steps in the import pathway, as demonstrated by following the fate of green fluorescent protein (GFP) fused to a transit peptide in the same transgenic plants. Based on the GFP results, we have developed a genetic screen to identify chloroplast import mutants that depends both on the correct import of GFP and a chloroplast enzyme that confers herbicide resistance. We have identified novel mutants that should provide important insights into how proteins are selectively targeted to chloroplasts and the components that regulate this complex pathway during organelle biogenesis.