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Journal of Biochemistry 2006 140(1):121-133; doi:10.1093/jb/mvj136
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© 2006 The Japanese Biochemical Society.

Regular Paper

Mutational and Functional Analysis of the Cryptic N-Terminal Targeting Signal for Both Mitochondria and Peroxisomes in Yeast Peroxisomal Citrate Synthase Cit2p

Jeong Goo Lee1,*, Yong Joo Lee1, Cheong Ho Lee2 and Pil Jae Maeng1,{dagger}

1 Department of Microbiology, School of Biological Science & Biotechnology, Chungnam National University, Daejeon 305-764; and 2 KT&G Central Research Institute, Daejeon 305-805, Korea

{dagger} To whom correspondence should be addressed. Tel: +82-42-821-6415, Fax: +82-42-821-6415, E-mail: pjmaeng{at}cnu.ac.kr

We previously found that the peroxisomal citrate synthase of Saccharomyces cerevisiae, Cit2p, contains a cryptic targeting signal for both peroxisomes (PTS) and mitochondria (MTS) within its 20–amino acid N-terminal segment [Lee et al. (2000) J. Biochem. 128, 1059–1072]. In the present study, the fine structure of the cryptic signal was scrutinized using green fluorescent protein fusions led by variants of the N-terminal segment. The minimum ranges of the cryptic signals for mitochondrial and peroxisomal targeting were shown to consist of the first 15– and 10–amino acid N-terminal segments, respectively. Substitution of the 3rd Val, 6th Leu, 7th Asn, or 8th Ser with Ala abolished the cryptic MTS function, however, no single substitution causing an obvious defect in PTS function was found. Neither the 15–amino acid N-terminal segment nor the C-terminal SKL sequence (PTS1) was necessary for Cit2p to restore the glutamate auxotrophy caused by the double {Delta}cit1 {Delta}cit2 mutation. The Cit2p variant lacking PTS1 [Cit2({Delta}SKL)p] partially restored the growth of both the {Delta}cit1 {Delta}cit2 and {Delta}cit1 mutants on acetate, while that carrying intact PTS1 or lacking the N-terminal segment [Cit2p, Cit2({Delta}N{Delta}SKL)p, and Cit2({Delta}N)p] did not. It is thus suggested that the potential of the N-terminal segment as an ambidextrous targeting signal can be unmasked by deletion of PTS1.

* Present address: Doheny Eye Institute, Univ. Southern California, 1450 San Pablo St. DVRC 203, Los Angeles, CA 90033, USA.


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 Mol. Biol. CellHome page
Y. J. Lee, K. L. Hoe, and P. J. Maeng
Yeast Cells Lacking the CIT1-encoded Mitochondrial Citrate Synthase Are Hypersusceptible to Heat- or Aging-induced Apoptosis
Mol. Biol. Cell, September 1, 2007; 18(9): 3556 - 3567.
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