The DNA Helicase, Hmi1p, Is Transported into Mitochondria by a C-terminal Cleavable Targeting Signal

Abstract

We have identified a novel mitochondrial targeting signal in the precursor of the DNA helicase Hmi1p of Saccharomyces cerevisiae that is located at the C terminus of the protein. Similar to classical N-terminal presequences, this C-terminal targeting signal consists of a stretch of positively charged amino acids that has the potential to form an amphipathic ␣-helix. Deletion of the C-terminal 36 amino acids of helicase resulted in loss of import into mitochondria, while deletion of the N-terminal 40 amino acids had no effect. When C-terminal regions of the helicase were placed at the C terminus of a passenger protein, dihydrofolate reductase, the resulting fusion proteins were directed into the mitochondrial matrix, and the C-terminal region of helicase became proteolytically processed. Import of helicase occurs in a C-to N-terminal direction; it requires a membrane potential and the TIM17-23 translocase together with mitochondrial Hsp70. Helicase is the only mitochondrial matrix protein identified thus far with a cleavable targeting signal at its C terminus.

14-3-3 protein is a regulator of the mitochondrial and chloroplast ATP synthase

“…Recently, it has been shown that 14-3-3 forms part of a guidance complex targeting nuclear-encoded proteins to the chloroplast (15). It is known that many 14-3-3 isoforms possess N-and C-terminal amphipathic ␣-helices (42,43), but these ␣-helices carry a negative charge, in contrast to the mitochondrial targeting presequences that form positively charged amphipathic ␣-helices (44). The elucidation of the exact mechanism of 14-3-3 import into organelles is a challenge for the future.…”

Dual localization of human DNA topoisomerase IIIα to mitochondria and nucleus

“…Examination of the amino acid sequences of known mammalian RecQ helicases revealed no apparent mitochondria-targeting signal peptides (results not shown). Although several mitochondrial helicases have been identified (41,59,60), the plausible association of hTop3␣ with any one of these, or a presently unknown mitochondrial DNA helicase, is yet to be studied.…”

Plant organellar protein targeting: a traffic plan still under construction

“…The protein is targeted by a 115 amino acid peptide at the N-terminus of the protein that lacks the amphipathic α-helical structure characteristic of targeting presequences. Moreover, a yeast mitochondrial helicase and a dual nuclear-mitochondrial Caenorhabditis elegans host cell factor contain mitochondrial targeting information at the carboxy end of the proteins [47,48]. The computer-based algorithms for protein targeting prediction currently available are simply not adequate to address this degree of variation in protein targeting determinants, and new algorithms are going to be needed to assess the extent to which these variations occur.…”