A Systematic Screen to Discover and Analyze Apicoplast Proteins Identifies a Conserved and Essential Protein Import Factor

PLoS Pathogens

Boris Striepen¹  Jessica L. Demerly¹  Lilach Sheiner¹  Michael W. White²  Olivier Lucas²  Michael S. Behnke³  Wandy L. Beatty³  Nicole Poulsen⁴ 

[1] Center for Tropical and Emerging Global Diseases, University of Georgia, Athens, Georgia, United States of America;Department of Molecular Medicine, University of South Florida, Tampa, Florida, United States of America;Department of Molecular Microbiology, Washington University School of Medicine, St. Louis, Missouri, United States of America;School of Chemistry and Biochemistry, Georgia Institute of Technology, Atlanta, Georgia, United States of America

关键词: Plastids;    Cellular structures;    organelles;    Algae;    Apicomplexa;    Sequence motif analysis;    Toxoplasma gondii;    Multiple alignment calculation;    Fluorescence microscopy;   

DOI  :  10.1371/journal.ppat.1002392

学科分类：生物科学（综合）

来源: Public Library of Science

【 摘 要 】

Parasites of the phylum Apicomplexa cause diseases that impact global health and economy. These unicellular eukaryotes possess a relict plastid, the apicoplast, which is an essential organelle and a validated drug target. However, much of its biology remains poorly understood, in particular its elaborate compartmentalization: four membranes defining four different spaces. Only a small number of organellar proteins have been identified in particular few proteins are known for non-luminal apicoplast compartments. We hypothesized that enlarging the catalogue of apicoplast proteins will contribute toward identifying new organellar functions and expand the realm of targets beyond a limited set of characterized pathways. We developed a bioinformatic screen based on mRNA abundance over the cell cycle and on phyletic distribution. We experimentally assessed 57 genes, and of 30 successful epitope tagged candidates eleven novel apicoplast proteins were identified. Of those, seven appear to target to the lumen of the organelle, and four localize to peripheral compartments. To address their function we then developed a robust system for the construction of conditional mutants via a promoter replacement strategy. We confirm the feasibility of this system by establishing conditional mutants for two selected genes – a luminal and a peripheral apicoplast protein. The latter is particularly intriguing as it encodes a hypothetical protein that is conserved in and unique to Apicomplexan parasites and other related organisms that maintain a red algal endosymbiont. Our studies suggest that this peripheral plastid protein, PPP1, is likely localized to the periplastid compartment. Conditional disruption of PPP1 demonstrated that it is essential for parasite survival. Phenotypic analysis of this mutant is consistent with a role of the PPP1 protein in apicoplast biogenesis, specifically in import of nuclear-encoded proteins into the organelle.

【 授权许可】

CC BY