Nuclear control of respiratory chain expression by nuclear respiratory factors and PGC-1-related coactivator

Abstract

Expression of the respiratory apparatus depends on both nuclear and mitochondrial genes. Although these genes are sequestered in distinct cellular organelles, their transcription relies on nucleus-encoded factors. Certain of these factors are directed to the mitochondria, where they sponsor the bi-directional transcription of mitochondrial DNA. Others act on nuclear genes that encode the majority of the respiratory subunits and many other gene products required for the assembly and function of the respiratory chain. The nuclear respiratory factors, NRF-1 and NRF-2, contribute to the expression of respiratory subunits and mitochondrial transcription factors and thus have been implicated in nucleo-mitochondrial interactions. In addition, coactivators of the PGC-1 family serve as mediators between the environment and the transcriptional machinery governing mitochondrial biogenesis. One family member, peroxisome proliferator-activated receptor gamma coactivator PGC-1-related coactivator (PRC), is an immediate early gene product that is rapidly induced by mitogenic signals in the absence of de novo protein synthesis. Like other PGC-1 family members, PRC binds NRF-1 and activates NRF-1 target genes. In addition, PRC complexes with NRF-2 and HCF-1 (host cell factor-1) in the activation of NRF-2-dependent promoters. HCF-1 functions in cell-cycle progression and has been identified as an NRF-2 coactivator. The association of these factors with PRC is suggestive of a role for the complex in cell growth. Finally, shRNA-mediated knock down of PRC expression results in a complex phenotype that includes the inhibition of respiratory growth on galactose and the loss of respiratory complexes. Thus, PRC may help integrate the expression of the respiratory apparatus with the cell proliferative program.

Figure 1

Nuclear respiratory factors (NRF-1 and NRF-2) in the expression of nuclear genes governing mitochondrial respiratory function. NRFs act on the majority of nuclear genes that specify subunits of the five respiratory complexes of the mitochondrial inner membrane. In addition, they act on many other genes whose products direct the expression and assembly of the respiratory apparatus. Promoters for most of the nuclear genes encoding mtDNA transcription and replication factors have functional recognition sites for NRF-1, NRF-2, or both. These factors are required for the expression of respiratory subunits from complexes I, III, IV, and V encoded by mtDNA. Similarly, genes for mitochondrial translational components—including ribosomal proteins and tRNA synthetases as well as heme biosynthetic enzymes localized to the mitochondrial matrix—are NRF-dependent. Increasing evidence also suggests that a number of genes specifying proteins of the mitochondrial protein import and assembly machinery are NRF targets, including subunits of the import receptor complexes and COX assembly factors. Thus, NRF-1 and NRF-2 are part of a unifying mechanism for the coordinate transcriptional control of respiratory chain expression. MRP = mitochondrial RNA processing; POL = polymerase; TOM = translocase of the outer mitochondrial membrane; cytc = cytochrome c.

Figure 2

Arrangement of conserved sequence motifs in PGC-1-related coactivator (PRC). PRC is the largest member of the PGC-1 coactivator family. Sequence similarities between PRC and the other PGC-1 family coactivators are confined to distinct sequence blocks that are spatially conserved among the three family members. These include a potent amino-terminal transcriptional activation domain (vertical hatched box), an expanded proline-rich region (filled box), an arginine/serine (R/S) rich domain (open box) and an RNA recognition motif (horizontal hatched box). In addition, PRC has the LXXLL coactivator signature sequence and the DHDY host cell factor 1 (HCF-1) binding site present in the other family members.

Figure 3

Summary of molecular interactions between PRC and its cognate transcription factors, NRF-1 and CREB. NRF-1 and CREB engage in specific in vitro and in vivo interactions with PRC, as evidenced by pull-down assays and co-immunoprecipitations (see text for references). Deletion fine mapping revealed that both transcription factors share distinct binding sites located between the activation domain and the proline-rich region and within the R/S domain. A fragment containing the upstream binding site inhibits cytochrome c promoter activity and respiratory growth on galactose when expressed in trans. As observed for the interaction between PPARγ and PGC-1α, the PRC-transcription factor interactions occur through the DNA-binding domains (cross hatched boxes) of both NRF-1 and CREB.

Figure 4

Summary of PRC-transcription factor complexes. The expression of PRC mRNA and protein is induced in quiescent fibroblasts by serum growth factors and downregulated upon exit from the cell cycle brought about either by serum withdrawal or contact inhibition. PRC engages in a direct interaction with NRF-1 and CREB in vitro and exists in a complex with each factor in cell extracts. PRC occupancy of the NRF-1- and CREB-dependent cytochrome c promoter also increases upon serum stimulation, along with transcription factor phosphorylation. By contrast, PRC does not bind any of the NRF-2 subunits in vitro. However, antibodies directed against PRC can immunoprecipitate both NRF-2β and HCF-1 from cell extracts. This, combined with the fact that HCF-1 engages in direct interactions with both PRC and NRF-2β suggests that all three proteins exist in a complex in vivo. This is supported by the observation that mutations in the DHDY HCF-1 binding site on PRC, or in key hydrophobic residues in the NRF-2β activation domain, that are required for interaction with HCF-1 inhibit PRC trans-activation through NRF-2.