미토콘드리아 내부 단백질들의 이동 신호 분석과 단백질 이동 기작 연구

Abstract

The majority of mitochondrial proteins are encoded in the nuclear genome and imported into mitochondria postranslationally from the cytosol. An N-terminal presequence functions as the signal for the import of mitochondrial proteins. However, the sequence information in the presequence remains elusive, and the targeting mechanism how specific protein targeting to mitochondria without mis-targeting to other organelles occurs is still unclear. This study reports the identification of critical sequence motifs from the presequence of Arabidopsis F1-ATPase γ subunit (pFAγ). pFAγ was divided into six 10- amino acid (aa) segments, designated P1 to P6 from the N- to the C-terminus, each of which was further divided into two 5-aa subdivisions. These P segments and their subdivisions were substituted with alanines and fused to GFP. Protoplast targeting experiments using these GFP constructs revealed that pFAγ contains several functional sequence motifs that are dispersed throughout the presequence. The sequence motifs DQEEG (P4a) and VVRNR (P5b) were involved in translocation across the mitochondrial membranes. The sequence motifs IAARP (P2b) and IAAIR (P3a) participated in binding to mitochondria by Tom20- dependent manner, and the sequence motifs RLLPS (P2a) and SISTQ (P5a) assisted in pulling proteins into the matrix. In addition, there are several pairs of sequence motifs that exhibit compensatorily or synergistic functions. The residues in P1 segment is involved in determining the targeting specificity of pFAγ to mitochondria. Especially, the arginine residues in P1 segment play a critical role to evade the mis-targeting of mitochondrial protein to chloroplasts. This feature is also conserved in other mitochondrial presequences, and these targeting sequences without the N-terminus or arginine residues have the ability to be imported exclusively into chloroplasts. Thus, I propose that the mitochondrial presequence is divided to two functionally different parts: the N-terminus determining the organelle specificity for efficient protein targeting into mitochondria, and the C-terminus containing multiple sequence motifs to function cooperatively during protein translocation into endosymbiotic organelles.