Abstract
In in vitro experiments, prenylcysteine lyase (Pcly) cleaves the thioether bond of prenylcysteines to yield free cysteine and the aldehyde of the isoprenoid lipid. However, the importance of this enzyme has not yet been fully defined at the biochemical or physiologic level. In this study, we show that Pcly is expressed at high levels in mouse liver, kidney, heart, and brain. To test whether Pcly deficiency would cause prenylcysteines to accumulate in tissues and result in pathologic consequences, we produced Pcly-deficient cell lines and Pcly-deficient mice (Pcly-/-). Pcly activity levels were markedly reduced in Pcly-/- cells and tissues. Pcly-/- fibroblasts were more sensitive than wild-type fibroblasts to growth inhibition when prenylcysteines were added to the cell culture medium. To determine if the reduced Pcly enzyme activity levels led to an accumulation of prenylcysteines within cells, mass spectrometry was used to measure farnesylcysteine and geranylgeranylcysteine levels in the tissues of Pcly-/- mice and wild-type controls. These studies revealed a striking accumulation of both farnesylcysteine and geranylgeranylcysteine in the brain and liver of Pcly-/- mice. This accumulation did not appear to be accompanied by significant pathologic consequences. Pcly-/- mice were healthy and fertile, and surveys of more than 30 tissues did not uncover any abnormalities. We conclude that prenylcysteine lyase does play a physiologic role in cleaving prenylcysteines in mammals, but the absence of this activity does not lead to major pathologic consequences.
Highlights
In in vitro experiments, prenylcysteine lyase (Pcly) cleaves the thioether bond of prenylcysteines to yield free cysteine and the aldehyde of the isoprenoid lipid
To inactivate Pcly, we adopted a strategy to eliminate the carboxyl-terminal portion of the protein
No Pcly transcripts were detectable in Pcly–/– tissues when the Northern blot was hybridized with exon 6 sequences
Summary
Prenylcysteine lyase; BAC, bacterial artificial chromosome; ES, embryonic stem; FCME, farnesylcysteine methyl ester; GGCME, geranylgeranylcysteine methyl ester; FC, farnesylcysteine; GC, geranylcysteine; GGC, geranylgeranylcysteine; PPT, palmitoyl protein thioesterase. Teines are substrates for that transporter [10]. If prenylcysteines were to accumulate, would they cause cell toxicity or tissue pathology? Located within lysosomes is a thioesterase, palmitoyl-protein thioesterase 1 (PPT1), that cleaves fatty acids from acylated cysteines within proteins. Mutations in PPT1 cause a lysosomal storage disease in humans, infantile neuronal ceroid lipofuscinosis (Batten disease) [11]. Tschantz et al [8, 9] have speculated that the absence of prenylcysteine lyase might cause a lysosomal storage disease.
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