TELLchloroplasts - Trafficking of news on Extracellular Latest Likely danger to chloroplasts

01/03/2025 to 28/02/2027

Plants have evolved a sophisticated system to perceive and transmit external stimuli to the cell interior and inform growth anddefense. The host lab discovered a new pathway linking the PM to chloroplasts and regulating defence, based on inducible proteinre-localization, co-opted by pathogens during evolution. Both the plant protein CALCIUM DEPENDENT PROTEIN KINASE 16 (CPK16) and the viral protein C4 from tomato yellow leaf curl virus re-localize from the PM to chloroplasts upon activation of pathogenassociated molecular pattern (PAMP)-triggered immunity (PTI); from the organelle, CPK16 acts as a positive regulator of downstreamdefence responses, while C4 acts as a suppressor. However, the molecular and cellular events enabling and regulating proteintrafficking from PM to chloroplasts remain elusive. Preliminary data in the host lab found the colocalization of C4 with SYP61- andAra7-positive compartments upon flg22 treatment; among them, C4 colocalizes with Ara7 at the periphery of chloroplasts. Uponflg22 treatment, CPK16 was also able to form mobile dots in the cytoplasm. These results suggest that endocytosis is likely to beinvolved in C4/CPK16 trafficking from the PM to chloroplasts. In addition, phosphorylation regulation and N-terminal cleavage couldalso regulate the PM-to-chloroplast protein trafficking as evidenced by mass spectrometry analyses. In this project, I will utilize bothCPK16 and C4 to investigate the molecular determinants underlying the protein trafficking from the cell surface to chloroplasts, including how proteins are transported from the PM to chloroplasts (Objective 1) and how N-myristoylated C4 and CPK16 arereleased from membrane compartments prior to chloroplast import (Objectives 2). By using cell biology, protein biochemistry,molecular biology, and plant genetic methods, this project will expand our understanding of the PM-to-chloroplast protein traffickingand serve as stepping stone for the engineering of immune responses.