Giardia intestinalis parasites contain mitosomes, one of the simplest mitochondrion-related organelles. Strategies to identify the functions of mitosomes have been limited mainly to homology detection, which is not suitable for identifying species-specific proteins and their functions. An in vivo enzymatic tagging technique based on the Escherichia coli biotin ligase (BirA) has been introduced to G. intestinalis; this method allows for the compartment-specific biotinylation of a protein of interest. Known proteins involved in the mitosomal protein import were in vivo tagged, cross-linked, and used to copurify complexes from the outer and inner mitosomal membranes in a single step. New proteins were then identified by mass spectrometry. This approach enabled the identification of highly diverged mitosomal Tim44 (GiTim44), the first known component of the mitosomal inner membrane translocase (TIM). In addition, our subsequent bioinformatics searches returned novel diverged Tim44 paralogs, which mediate the translation and mitosomal insertion of mitochondrially encoded proteins in other eukaryotes. However, most of the identified proteins are specific to G. intestinalis and even absent from the related diplomonad parasite Spironucleus salmonicida, thus reflecting the unique character of the mitosomal metabolism. The in vivo enzymatic tagging also showed that proteins enter the mitosome posttranslationally in an unfolded state and without vesicular transport.

Centre for Microbial Diversity and Evolution Department of Botany University of British Columbia Vancouver BC Canada

Department of Parasitology Faculty of Science Charles University Prague Prague Czech Republic

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Mlf mediates proteotoxic response via formation of cellular foci for protein folding and degradation in Giardia

Installation of LYRM proteins in early eukaryotes to regulate the metabolic capacity of the emerging mitochondrion

Adaptation of the late ISC pathway in the anaerobic mitochondrial organelles of Giardia intestinalis

Structurally derived universal mechanism for the catalytic cycle of the tail-anchored targeting factor Get3

Inheritance of the reduced mitochondria of Giardia intestinalis is coupled to the flagellar maturation cycle

Retortamonads from vertebrate hosts share features of anaerobic metabolism and pre-adaptations to parasitism with diplomonads

The evolution of the Puf superfamily of proteins across the tree of eukaryotes

A Single Tim Translocase in the Mitosomes of Giardia intestinalis Illustrates Convergence of Protein Import Machines in Anaerobic Eukaryotes

Dynamic secretome of Trichomonas vaginalis: Case study of β-amylases

Giardia intestinalis mitosomes undergo synchronized fission but not fusion and are constitutively associated with the endoplasmic reticulum

Organelles that illuminate the origins of Trichomonas hydrogenosomes and Giardia mitosomes

Evolution of the Tim17 protein family