Exploring Transit Peptides through Photosynthetic Restoration in Chlamydomonas

• Applicant: Oliver Caspari, Ph.D.
• Subject Area: Plant Biochemistry and Biophysics
• Term: since 2024
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 551520827

The vast majority of chloroplast proteins are encoded in the nucleus and imported into the organelle. This import relies on N-terminal transit peptides. Over 40 years of research into these transit peptides has produced a large body of work, and many sequence elements have been proposed to play a role in determining chloroplast import. Yet an overarching vision of how transit peptides specify import has remained elusive. There is an urgent need to extract the fundamental rules that govern cTP efficacy and specificity in order to generate custom cTPs that reliably target heterologous proteins. The project proposed in these pages is designed to systematically test the confusing array of targeting determinants described in the literature, and then unify that knowledge in a generative model enabling reliable cTP design. To this end, a screen is proposed, whereby photosynthetic mutants of the green alga Chlamydomonas reinhardtii are complemented with constructs testing a large number of transit peptide variants. Since complementation can only occur if a given peptide correctly specifies chloroplast import, the screen provides a simple tool to dissect the molecular determinants of chloroplast targeting.

• DFG Programme: Research Grants