TIM29 is required for enhanced stem cell activity during regeneration in the flatworm Macrostomum lignano

Stijn Mouton,Kirill Ustyantsev,Eugene Berezikov,Frank Beltman,Lisa Glazenburg

doi:10.1038/s41598-020-80682-7

Abstract

TIM29 is a mitochondrial inner membrane protein that interacts with the protein import complex TIM22. TIM29 was shown to stabilize the TIM22 complex but its biological function remains largely unknown. Until recently, it was classified as one of the Domain of Unknown Function (DUF) genes, with a conserved protein domain DUF2366 of unclear function. Since characterizing DUF genes can provide novel biological insight, we used previously established transcriptional profiles of the germline and stem cells of the flatworm Macrostomum lignano to probe conserved DUFs for their potential role in germline biology, stem cell function, regeneration, and development. Here, we demonstrate that DUF2366/TIM29 knockdown in M. lignano has very limited effect during the normal homeostatic condition but prevents worms from adapting to a highly proliferative state required for regeneration.

Highlights

TIM29 is a mitochondrial inner membrane protein that interacts with the protein import complex the Inner Membrane 22 (TIM22)
To facilitate the discovery of novel genes involved in stem cell function, germline biology, regeneration, and development, we identified all genes in the M. lignano genome-guided transcriptome assembly M lig_3_7_DV1_v312 encoding uncharacterized proteins
Based on the expression level of the Domain of Unknown Function (DUF) in different conditions and using previously established neoblast and germline transcriptional s ignatures[11], categories were provided to predict their functional role in stem cells, the germline, regeneration, and development (Table 1)

Summary

Introduction

TIM29 is a mitochondrial inner membrane protein that interacts with the protein import complex TIM22. Evolutionary conservation suggests that many of these DUFs are important, but studies indicated that they more likely represent biological functions specific to certain conditions, or certain groups of organisms, rather than being part of the core machinery common to all life[1,3]. In 2016, we established transcriptional signatures of proliferating somatic neoblasts and germline cells by performing RNAseq of FACS-isolated cells of worms in different conditions[11] This dataset represents a convenient resource to identify DUFs with functions related to in vivo stem cell and germline regulation, regeneration, and development. It was suggested that TIM29 contacts the Translocase of the Outer Membrane (TOM) complex, enabling transport of hydrophobic carrier substrates across the aqueous intermembrane space[13] Both papers studied the effect of TIM29 RNA interference (RNAi) on HEK cell proliferation, and reported contradicting results. We identify the DUFs conserved in M. lignano and demonstrate the crucial role of Mlig-DUF2366/ TIM29 for adapting to highly proliferative conditions during whole-body regeneration by means of RNA interference studies

Methods

Results

Conclusion