Heterogeneous distribution of pyruvate dehydrogenase in the matrix of mitochondria

A fusion protein between GFP and the Elalpha subunit of the pyruvate dehydrogenase (PDH) complex was created and shown to assemble into functional PDH complexes using immunoprecipitation and activity assays. The expression of this GFP-Elalpha chimera is specific to mitochondria and results in two different fluorescence patterns. These patterns have been distinguished by immunolabeling experiments using monoclonal antibodies against PDH subunits and GFP. The bright, localized fluorescent spots represent the assembled form of the GFP-Elalpha in PDH complexes. The uniform, dim fluorescence is given by the unassembled chimera free to diffuse throughout the mitochondrial reticulum. This study reveals a discrete, heterogeneous distribution of PDH complexes in the matrix of mitochondria, both in cells with normal and reduced levels of PDH. The uneven arrangement of PDH complexes is maintained over time and most likely reflects the structural and metabolic compartmentalization of mitochondria. (C) 2002 Elsevier Science B.V. and Mitochondria Research Society. All rights reserved.