Photosensitisation properties of mitochondrially localised green fluorescent protein

Biochemical and Biophysical Research Communications
C ZhangP Nagley

Abstract

The photosensitisation properties of a mitochondrially localised green fluorescent protein (GFP) variant were established in cultured monkey kidney cells. We first cloned into a mammalian expression vector the thermostable variant GFP5, fused at its N-terminus to the 16-amino acid mitochondrial targeting sequence of human 3-oxoacyl-CoA thiolase. The recombinant plasmid thus constructed, pCZ34, was transfected into COS7 cells, under conditions for transient expression. GFP5 was shown to have a mitochondrial localisation by confocal microscopy, confirmed by its almost identical pattern with that of MitoTracker Red which selectively stains mitochondria. After photoirradiation of such transfected COS7 cells with 390-570 nm light to excite the mitochondrially localised GFP5, significant cell killing was observed. DAPI-staining of the dead cells detached from the growth support revealed cells to have undergone apoptosis. Thus, GFP5 can be used in the development of an organelle-specific photosensitiser since it can be targeted to different subcellular locations by protein engineering of the signal sequences.

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Citations

Jun 15, 2004·Journal of Virology·Kenneth RajPeter Beard
Nov 27, 2007·Advanced Drug Delivery Reviews·Kentaro KogureHideyoshi Harashima
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Apoptosis

Apoptosis is a specific process that leads to programmed cell death through the activation of an evolutionary conserved intracellular pathway leading to pathognomic cellular changes distinct from cellular necrosis