A novel TAT-mitochondrial signal sequence fusion protein is processed, stays in mitochondria, and crosses the placenta

Victoria Del Gaizo, R. Payne

Research output: Contribution to journalArticle

91 Citations (Scopus)

Abstract

Mutations in nuclear and mitochondrial genomes can lead to defects in mitochondrial function. To date, repair of these defects with exogenous proteins or gene transfer has been difficult with either viral or nonviral vectors. We hypothesized that TAT fusion proteins would cross both mitochondrial membranes and that incorporation of a mitochondrial signal sequence into a TAT fusion protein would allow processing and localization of exogenous proteins in mitochondria. A TAT-mitochondrial malate dehydrogenase signal sequence (mMDH)-enhanced green fluorescent protein (eGFP) fusion protein was constructed. TAT-mMDH-eGFP allowed rapid transduction and localization of fusion protein into mitochondria of multiple cell types. In contrast, TAT-GFP, without a mitochondrial signal sequence, rapidly transduced into cells and mitochondria, displayed pseudo-first-order kinetics, but did not remain there. Mice injected 5 days prior with TAT-mMDH-eGFP had detectable eGFP activity in multiple tissue types. Western blotting of cytosolic and mitochondrial fractions isolated from their livers confirmed eGFP localization to mitochondria and that the mMDH transit peptide was recognized and processed. Furthermore, TAT-mMDH-eGFP fusion protein injected into pregnant mice crossed the placenta and was detectable in both the fetus and the newborn pups. TAT fusion proteins containing a mitochondrial signal sequence are a viable method to localize proteins to mitochondria.

Original languageEnglish (US)
Pages (from-to)720-730
Number of pages11
JournalMolecular Therapy
Volume7
Issue number6
DOIs
StatePublished - Jun 1 2003
Externally publishedYes

Fingerprint

Protein Sorting Signals
Placenta
Mitochondria
Malate Dehydrogenase
Proteins
Mitochondrial Genome
Mitochondrial Membranes
enhanced green fluorescent protein
Fetus
Western Blotting
Mutation
Liver

Keywords

  • eGFP
  • Mitochondria
  • Placenta
  • Protein sorting signals
  • Protein transduction domain
  • TAT

ASJC Scopus subject areas

  • Molecular Biology

Cite this

A novel TAT-mitochondrial signal sequence fusion protein is processed, stays in mitochondria, and crosses the placenta. / Del Gaizo, Victoria; Payne, R.

In: Molecular Therapy, Vol. 7, No. 6, 01.06.2003, p. 720-730.

Research output: Contribution to journalArticle

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