Twin-arginine translocation (Tat) pathways have been well-characterized in bacteria and chloroplasts. Genes encoding a TatC protein are found in almost all plant mitochondrial genomes but to date these have not been extensively investigated. For the first time it could be demonstrated that this mitochondrial-encoded TatC is a functional gene that is translated into a protein in the model plant Arabidopsis thaliana. A TatB-like subunit localized to the inner membrane was also identified that is nuclear-encoded and is essential for plant growth and development, indicating that plants potentially require a Tat pathway for mitochondrial biogenesis.