Determinants for stop‐transfer and post‐import pathways for protein targeting to the chloroplast inner envelope membrane

Abstract

Chloroplast biogenesis relies on the proper transport of thousands of nuclear encoded proteins into this organelle. The majority of nucleus‐encoded chloroplast proteins are imported via the TOC‐TIC translocons in the chloroplast envelope. Several of these proteins are further targeted to suborganellar compartments by additional targeting systems downstream of the import apparatus. The thylakoid targeting systems were presumably conserved from the original endosymbiont. In contrast, our knowledge of the mechanisms of protein targeting and integration at the inner envelope membrane (IM) is very limited. Previous reports have shown that IM proteins are transferred to the inner membrane via a stop‐transfer mechanism. Recent studies have demonstrated that one component of the Tic machinery is imported into the stroma and re‐inserted into the IM in a post‐import mechanism. This led us to investigate these pathways in more detail. Our analyses confirm the existence of both stop‐transfer and post‐import mechanisms of protein targeting to the IM. Furthermore, our results point to the conclusion that the transmembrane helices are not interchangeable and not sufficient for the choice of the insertion pathway. Further deletion and substitution analyses are being performed to better understand the features that determine protein insertion and topology into the chloroplast IM. Supported by NIH, CAPES/Fulbright.