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Scholarly Works (5 results)

  • Thesis
  • Peer Reviewed
UCLA Electronic Theses and Dissertations (2014)

Coenzyme Q (ubiquinone or Q) is an essential redox-active, polyisoprenylated benzoquinone lipid essential for electron and proton transport in the mitochondrial respiratory chain. Eleven genes products, Coq1-Coq9, Yah1 and Arh1, are required for Q biosynthesis in yeast Saccharomyces cerevisiae.

Chapter 2 details the investigation of the biological function of Coq8 and its human homolog ADCK3. Expression of ADCK3 harboring an amino-terminal yeast mitochondrial leader sequence successfully rescued growth of coq8 mutants on non-fermentable carbon source, partially restored Q biosynthesis and the phosphorylation states of Coq3, Coq5, and Coq7.

Chapter 3 investigates the use of over-expression of COQ8 as a tool to study Q biosynthesis pathway in S. cerevisiae. Over-expression of the Coq8 protein restores the steady state levels of the unstable Coq proteins. This stabilization results in the accumulation of several novel Q6 biosynthetic intermediates. Several of the new intermediates contain a C4-amine and provide information on the deamination reaction that takes place when para-aminobenzoci acid is used as a ring precursor in Q biosynthesis.

Chapter 4 studies the use of para-aminobenzoic acid, and resveratrol as alternative aromatic ring precursors in Q biosynthesis in E. coli, S. cerevisiae, mouse and human cells. In contrast to S. cerevisiae, neither E. coli nor mammalian cells could utilize pABA as ring precursors in Q biosynthesis. However, E. coli cells labeled with 13C6-pABA generated several novel N-containing early intermediates, suggesting UbiA, UbiD,X, and Ubil are capable of using pABA as substrates. E. coli, S. cerevisiae, human and mouse cells cultured in the presence of 13C6-resveratrol were able to synthesize 13C6-Q. Thus, future evaluation of the physiological and pharmacological responses to dietary polyphenols should consider their metabolism to Q.

    Cover page: Investigation of the roles of Coq8p, a putative kinase, and alternate ring precursors in coenzyme Q biosynthesis
    • Article
    • Peer Reviewed
    UCLA Previously Published Works (2014)
    Coenzyme Q biosynthesis in yeast requires a multi-subunit Coq polypeptide complex. Deletion of any one of the COQ genes leads to respiratory deficiency and decreased levels of the Coq4, Coq6, Coq7, and Coq9 polypeptides, suggesting that their association in a high molecular mass complex is required for stability. Over-expression of the putative Coq8 kinase in certain coq null mutants restores steady-state levels of the sensitive Coq polypeptides and promotes the synthesis of late-stage Q-intermediates. Here we show that over-expression of Coq8 in yeast coq null mutants profoundly affects the association of several of the Coq polypeptides in high molecular mass complexes, as assayed by separation of digitonin extracts of mitochondria by two-dimensional blue-native/SDS PAGE. The Coq4 polypeptide persists at high molecular mass with over-expression of Coq8 in coq3, coq5, coq6, coq7, coq9, and coq10 mutants, indicating that Coq4 is a central organizer of the Coq complex. Supplementation with exogenous Q6 increased the steady-state levels of Coq4, Coq7, and Coq9, and several other mitochondrial polypeptides in select coq null mutants, and also promoted the formation of late-stage Q-intermediates. Q supplementation may stabilize this complex by interacting with one or more of the Coq polypeptides. The stabilizing effects of exogenously added Q6 or over-expression of Coq8 depend on Coq1 and Coq2 production of a polyisoprenyl intermediate. Based on the observed interdependence of the Coq polypeptides, the effect of exogenous Q6, and the requirement for an endogenously produced polyisoprenyl intermediate, we propose a new model for the Q-biosynthetic complex, termed the CoQ-synthome.
      Cover page: Coenzyme Q supplementation or over-expression of the yeast Coq8 putative kinase stabilizes multi-subunit Coq polypeptide complexes in yeast coq null mutants
      • Article
      • Peer Reviewed
      UCLA Previously Published Works (2015)
      Coenzyme Q (Q or ubiquinone) is a redox-active polyisoprenylated benzoquinone lipid essential for electron and proton transport in the mitochondrial respiratory chain. The aromatic ring 4-hydroxybenzoic acid (4HB) is commonly depicted as the sole aromatic ring precursor in Q biosynthesis despite the recent finding that para-aminobenzoic acid (pABA) also serves as a ring precursor in Saccharomyces cerevisiae Q biosynthesis. In this study, we employed aromatic (13)C6-ring-labeled compounds including (13)C6-4HB, (13)C6-pABA, (13)C6-resveratrol, and (13)C6-coumarate to investigate the role of these small molecules as aromatic ring precursors in Q biosynthesis in Escherichia coli, S. cerevisiae, and human and mouse cells. In contrast to S. cerevisiae, neither E. coli nor the mammalian cells tested were able to form (13)C6-Q when cultured in the presence of (13)C6-pABA. However, E. coli cells treated with (13)C6-pABA generated (13)C6-ring-labeled forms of 3-octaprenyl-4-aminobenzoic acid, 2-octaprenyl-aniline, and 3-octaprenyl-2-aminophenol, suggesting UbiA, UbiD, UbiX, and UbiI are capable of using pABA or pABA-derived intermediates as substrates. E. coli, S. cerevisiae, and human and mouse cells cultured in the presence of (13)C6-resveratrol or (13)C6-coumarate were able to synthesize (13)C6-Q. Future evaluation of the physiological and pharmacological responses to dietary polyphenols should consider their metabolism to Q.
        Cover page: Resveratrol and para-coumarate serve as ring precursors for coenzyme Q biosynthesis[S]
        • Article
        • Peer Reviewed
        UCLA Previously Published Works (2011)
        Therapy of mitochondrial respiratory chain diseases is complicated by limited understanding of cellular mechanisms that cause the widely variable clinical findings. Here, we show that focal segmental glomerulopathy-like kidney disease in Pdss2 mutant animals with primary coenzyme Q (CoQ) deficiency is significantly ameliorated by oral treatment with probucol (1% w/w). Preventative effects in missense mutant mice are similar whether fed probucol from weaning or for 3 weeks prior to typical nephritis onset. Furthermore, treating symptomatic animals for 2 weeks with probucol significantly reduces albuminuria. Probucol has a more pronounced health benefit than high-dose CoQ(10) supplementation and uniquely restores CoQ(9) content in mutant kidney. Probucol substantially mitigates transcriptional alterations across many intermediary metabolic domains, including peroxisome proliferator-activated receptor (PPAR) pathway signaling. Probucol's beneficial effects on the renal and metabolic manifestations of Pdss2 disease occur despite modest induction of oxidant stress and appear independent of its hypolipidemic effects. Rather, decreased CoQ(9) content and altered PPAR pathway signaling appear, respectively, to orchestrate the glomerular and global metabolic consequences of primary CoQ deficiency, which are both preventable and treatable with oral probucol therapy.
          Cover page: Probucol ameliorates renal and metabolic sequelae of primary CoQ deficiency in Pdss2 mutant mice
          • Article
          • Peer Reviewed
          UCLA Previously Published Works (2013)
          Identification of single-gene causes of steroid-resistant nephrotic syndrome (SRNS) has furthered the understanding of the pathogenesis of this disease. Here, using a combination of homozygosity mapping and whole human exome resequencing, we identified mutations in the aarF domain containing kinase 4 (ADCK4) gene in 15 individuals with SRNS from 8 unrelated families. ADCK4 was highly similar to ADCK3, which has been shown to participate in coenzyme Q10 (CoQ10) biosynthesis. Mutations in ADCK4 resulted in reduced CoQ10 levels and reduced mitochondrial respiratory enzyme activity in cells isolated from individuals with SRNS and transformed lymphoblasts. Knockdown of adck4 in zebrafish and Drosophila recapitulated nephrotic syndrome-associated phenotypes. Furthermore, ADCK4 was expressed in glomerular podocytes and partially localized to podocyte mitochondria and foot processes in rat kidneys and cultured human podocytes. In human podocytes, ADCK4 interacted with members of the CoQ10 biosynthesis pathway, including COQ6, which has been linked with SRNS and COQ7. Knockdown of ADCK4 in podocytes resulted in decreased migration, which was reversed by CoQ10 addition. Interestingly, a patient with SRNS with a homozygous ADCK4 frameshift mutation had partial remission following CoQ10 treatment. These data indicate that individuals with SRNS with mutations in ADCK4 or other genes that participate in CoQ10 biosynthesis may be treatable with CoQ10.
            Cover page: ADCK4 mutations promote steroid-resistant nephrotic syndrome through CoQ10 biosynthesis disruption
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