UCP3 expressed in yeast is primarily localized in extramitochondrial particles

Previously it was concluded (1) that, differently from UCP1, on expression in Saccharomyces cerevisiae, UCP3, and UCP3 short (UCP3s) are in a deranged state, allowing for unregulated uncoupling. Here we show that the bulk of UCP3 and UCP3s is in extramitochondrial aggregates whether expressed with h igh or medium expression vectors. The evidence is based on the insolubility of most UCP3 and UCP3s in nonionic detergents such as Triton X100, in cont rast to UCP1. Using very high expression vector, macroscopic evidence for e xtramitochondrial UCP3 containing particles is a viscous white sediment sur rounding the mitochondrial fraction which contains UCP3 as inclusion body t ype aggregate. Together with the previous data it is concluded that uncoupl ing due to small amounts of incorporated, deranged, and nucleotide insensit ive UCP3 prevents incorporation of the bulk of UCP3 into mitochondria, This finding also provides a simple and stringent assay for the state of hetero logously expressed in mitochondrial membrane proteins. (C) 2001 Academic Pr ess.