Coenzyme Q (CoQ) is a component of the electron transport chain that participates in aerobic cellular respiration to produce ATP. In addition, CoQ acts as an electron acceptor in several enzymatic ...reactions involving oxidation-reduction. Biosynthesis of CoQ has been investigated mainly in Escherichia coli and Saccharomyces cerevisiae, and the findings have been extended to various higher organisms, including plants and humans. Analyses in yeast have contributed greatly to current understanding of human diseases related to CoQ biosynthesis. To date, human genetic disorders related to mutations in eight COQ biosynthetic genes have been reported. In addition, the crystal structures of a number of proteins involved in CoQ synthesis have been solved, including those of IspB, UbiA, UbiD, UbiX, UbiI, Alr8543 (Coq4 homolog), Coq5, ADCK3, and COQ9. Over the last decade, knowledge of CoQ biosynthesis has accumulated, and striking advances in related human genetic disorders and the crystal structure of proteins required for CoQ synthesis have been made. This review focuses on the biosynthesis of CoQ in eukaryotes, with some comparisons to the process in prokaryotes.
CoQ (coenzyme Q), an isoprenylated benzoquinone, is a well‐known component of the electron‐transfer system in eukaryotes. The main role of CoQ is to transfer electrons from NADH dehydrogenase and ...succinate dehydrogenase to CoQ:cytochrome c reductase in the respiratory chain. However, recent evidence indicates that an involvement in respiration is not the only role of CoQ. The second apparent role of CoQ is its anti‐oxidation property, and other novel roles for CoQ, such as in disulfide‐bond formation, sulfide oxidation and pyrimidine metabolism, have been reported. CoQ10, having ten isoprene units in the isoprenoid side chain, has been used as a medicine and is now commercially popular as a food supplement. Two yeast species, namely the budding yeast Saccharomyces cerevisiae, which produces CoQ6, and the fission yeast Schizosaccharomyces pombe, which produces CoQ10, are the main subjects of the present minireview because they have greatly contributed to our basic knowledge of CoQ biosynthesis among eukaryotes. The biosynthetic pathway that converts p‐hydroxybenzoate into CoQ consists of eight steps in yeasts. The five enzymes involved in the biosynthetic pathway have been identified in both yeasts, yet the functions of three proteins were still not known. Analyses of the biosynthetic pathway in yeasts also contribute to the understanding of human genetic diseases related to CoQ deficiency. In the present minireview I focus on the biochemical and commercial aspects of CoQ in yeasts and in other organisms for comparison.
The phospholipase B homolog Plb1 and the cAMP-dependent protein kinase (PKA) pathway are required by fission yeast, also known as to
Schizosaccharomyces pombe
, to grow under KCl-stress conditions. ...Here, we report the relative contributions of Plb1 and the cAMP/PKA pathway during the hypertonic stress response. We show that the
plb1∆
,
cyr1∆
, and
pka1∆
single mutants are sensitive to high concentrations of KCl but insensitive to sorbitol-induced osmotic stress. In contrast, the
plb1∆ cyr1∆
and
plb1∆ pka1∆
double mutants are hypersensitive to KCl and sorbitol. The
cyr1∆ pka1∆
double mutants showed the same phenotype of each single mutant. Growth inhibition due to hypertonic stress in the
plb1∆
,
plb1∆ cyr1∆
, and
plb1∆ pka1∆
strains was partially rescued by
cgs1
deletion—
cgs1∆
has constitutively active Pka1—or by the deletion of transcription factor Rst2, which is negatively regulated by Pka1. Pka1-GFP localized in the nucleus and cytoplasm in
plb1∆
, whereas it is localized only in the cytoplasm in
cyr1∆
, indicating that Plb1 does not regulate Pka1 localization. Glucose limitation downregulates the PKA pathway, and it was accordingly observed that glucose limitation in
plb1∆
further increased the strain’s sensitivity to KCl. Growth inhibition by KCl in
plb1∆
under glucose-limited conditions was significantly rescued by
cgs1∆
and slightly rescued by
rst2∆
. These findings indicate that, in fission yeast, Plb1 and the glucose-sensing cAMP/PKA pathway play a synergistic role in responding to hypertonic stress.
Some marine fishes are algae-feeding, and the microorganisms in their digestive tracts produce carbohydrate hydrolyzing enzymes such as agarose and fucosidase, which are potentially interesting ...resource for new functional enzymes. The purpose of this study was to establish a method for identifying and utilizing characteristic bacteria from the intestines of two algae-eating fish species: Andamia tetradactylus, which exclusively eats algae on the rock surface, and stellar rockskipper Entomacrodus stellifer, which feeds on both algae and invertebrates. We tested the species composition of the intestinal bacterial flora and found that Proteobacteria were commonly found both in species as in the common gut communities of marine fish, whereas Spirochaetes and Tenericutes occupied the flora of A. tetradactylus. We then performed anaerobic and aerobic cultures and isolated 34 and 44 strains including 48 strains belonged to Vibrio species from A. tetradactylus and E. stellifer. We observed that some Vibrio strains formed a clear boundary to avoid contacting other strains of bacteria. Whole-genome sequencing of such two Vibrio alginolyticus strains revealed two cyclic chromosomes commonly found in the genome of Vibrio species, and some unique genes encoding alginate lyase, chitinases, and type I-F CRISPR-associated endoribonuclease for the first time in Vibrio alginolyticus.
The homothallic fission yeast Schizosaccharomyces pombe undergoes sexual differentiation when starved, but sam (skips the requirement of starvation for mating) mutants such as those carrying ...mutations in adenylate cyclase (cyr1) or protein kinase A (pka1) mate without starvation. Here, we identified sam3, a dominant negative allele of rad24, encoding one of two 14-3-3 proteins. Genetic mapping and whole-genome sequencing showed that the sam3 mutation comprises a change in nucleotide at position 959 from guanine to adenine, which switches the amino acid at position 185 from glutamic acid to lysine (E185K). We generated the rad24-E185K integrated mutant and its phenotype was similar to that of the sam3 mutant, including calcium sensitivity and UV non-sensitivity, but the phenotype is different from that of the DELTArad24 strain. While the UV-sensitive phenotype was observed in the DELTArad24 mutant, it was not observed in the sam3 and rad24-E185K mutants. The expression of the rad24-E185K gene in wild type cells induced spore formation in the nutrient rich medium, confirming rad24-E185K is dominant. This dominant effect of rad24-E185K was also observed in DELTAras1 and DELTAbyr2 diploid mutants, indicating that rad24-E185K generate stronger phenotype than rad24 null mutants. Ste11, the key transcription factor for sexual differentiation was expressed in sam3 mutants without starvation and it predominantly localized to the nucleus. The Rad24-E185K mutant protein retained its interaction with Check point kinase1 (Chk1), whereas it reduced interaction with Ste11, an RNA binding protein Mei2, and a MAPKKK Byr2, freeing these proteins from negative regulation by Rad24, that account for the sam phenotype and UV non-sensitive phenotype. Glucose depletion in rad24-E185K or DELTApka1 DELTArad24 double mutation induced haploid meiosis, leading to the formation of spores in haploid. The position of glutamic acid 185 is conserved in all major 14-3-3s; hence, our finding of a dominant negative allele of 14-3-3 is useful for understanding 14-3-3s in other organisms.
Coenzyme Q (CoQ) is an essential factor for aerobic growth and oxidative phosphorylation in the electron transport system. The biosynthetic pathway for CoQ has been proposed mainly from biochemical ...and genetic analyses of Escherichia coli and Saccharomyces cerevisiae; however, the biosynthetic pathway in higher eukaryotes has been explored in only a limited number of studies. We previously reported the roles of several genes involved in CoQ synthesis in the fission yeast Schizosaccharomyces pombe. Here, we expand these findings by identifying ten genes (dps1, dlp1, ppt1, and coq3-9) that are required for CoQ synthesis. CoQ10-deficient S. pombe coq deletion strains were generated and characterized. All mutant fission yeast strains were sensitive to oxidative stress, produced a large amount of sulfide, required an antioxidant to grow on minimal medium, and did not survive at the stationary phase. To compare the biosynthetic pathway of CoQ in fission yeast with that in higher eukaryotes, the ability of CoQ biosynthetic genes from humans and plants (Arabidopsis thaliana) to functionally complement the S. pombe coq deletion strains was determined. With the exception of COQ9, expression of all other human and plant COQ genes recovered CoQ10 production by the fission yeast coq deletion strains, although the addition of a mitochondrial targeting sequence was required for human COQ3 and COQ7, as well as A. thaliana COQ6. In summary, this study describes the functional conservation of CoQ biosynthetic genes between yeasts, humans, and plants.
The cAMP-dependent protein kinase (PKA) pathway in
Schizosaccharomyces pombe
plays an important role in microtubule organization and chromosome segregation. Typically, loss of functional Pka1 induces ...sensitivity to the microtubule-destabilizing drug thiabendazole (TBZ) and chromosome mis-segregation. To determine the mechanism via which Pka1 is involved in these events, we explored the relevance of transcription factors by creating a double-deletion strain of
pka1
and 102 individual genes encoding transcription factors. We found that
rst2∆
,
tfs1∆
,
mca1∆
, and
moc3∆
suppressed the TBZ-sensitive phenotype of the
pka1∆
strain, among which
tfs1∆
was the strongest suppressor. All single mutants (
rst2∆
,
tfs1∆
,
mca1∆
, and
moc3∆
) showed a TBZ-tolerant phenotype. Tfs1 has two transcriptional domains (TFIIS and Zn finger domains), both of which contributed to the suppression of the
pka1∆
-induced TBZ-sensitive phenotype.
pka1∆
-induced chromosome mis-segregation was rescued by
tfs1∆
in the presence of TBZ.
tfs1
overexpression induced the TBZ-sensitive phenotype and a high frequency of chromosome mis-segregation, suggesting that the amount of Tfs1 must be strictly controlled. However, Tfs1-expression levels did not differ between the wild-type and
pka1∆
strains, and the Tfs1-GFP protein was localized to the nucleus and cytoplasm in both strains, which excludes the direct regulation of expression and localization of Tfs1 by Pka1. Growth inhibition by TBZ in
pka1∆
strains was notably rescued by double deletion of
rst2
and
tfs1
rather than single deletion of
rst2
or
tfs1
, indicating that Rst2 and Tfs1 contribute independently to counteract TBZ toxicity. Our findings highlight Tfs1 as a key transcription factor for proper chromosome segregation.
The cAMP-dependent protein kinase Pka1 is known as a regulator of glycogenesis, transition into meiosis, chronological aging, and stress responses in the fission yeast, Schizosaccharomyces pombe. We ...demonstrated here that Pka1 is responsible for normal growth in the presence of the microtubule-destabilization drug TBZ and proper chromosome segregation. The deletion of the pka1 gene resulted in the TBZ-sensitive phenotype and chromosome mis-segregation. We isolated the mal3 gene as a multi-copy suppressor of the TBZ-sensitive phenotype in the pka1Δ strains. Overexpression of the CH domain (1-143) or the high-affinity microtubule binding mutant (1-143 Q89R) of Mal3 rescued the TBZ-sensitive phenotype in the pka1Δ and mal3Δ strains, while the EB1 domain (135-308) and the mutants defective in microtubule binding (1-143 Q89E) failed to do so in the same strains. Chromosome mis-segregation caused by TBZ in the pka1Δ or mal3Δ strains was suppressed by the overexpression of the Mal3 CH domain (1-143), Mal3 CH domain with the coiled-coil domain (1-197), or full-length Mal3. Overexpression of EB1 orthologs from Saccharomyces cerevisiae, Arabidopsis thaliana, Mus musculus, or Homo sapiens suppressed the TBZ-sensitive phenotype in the pka1Δ strains, indicating their conserved functions. These findings suggest that Pka1 and the microtubule binding of the Mal3 CH domain play a role in the maintenance of proper chromosome segregation.
The cAMP-dependent protein kinase (Pka1) regulates many cellular events, including sexual development and glycogenesis, and response to the limitation of glucose, in
Schizosaccharomyces pombe
. ...Despite its importance in many cellular events, the targets of the cAMP/PKA pathway have not been fully investigated. Here, we demonstrate that the expression of
mug14
is induced by downregulation of the cAMP/PKA pathway and limitation of glucose. This regulation is dependent on the function of Rst2, a transcription factor that regulates transition from mitosis to meiosis. The loss of the C2H2-type zinc finger domain in Rst2, termed Rst2 (C2H2∆), abolished the induction of Mug14 expression. Upon deletion of the stress starvation response element of the
S. pombe
(STREP: CCCCTC) sequence, which is a potential binding site of Rst2 on
mug14
, in the
pka1∆
strain, its induction was abolished. The expression of Mug14 was significantly reduced and delayed by the limitation of glucose and also by nitrogen starvation in the
rst2∆
strain. Mug14 is known to share a common function with Mde1 and Mta3 in the methionine salvage pathway, but the expression of
mde1
and
mta3
mRNAs was not enhanced by
pka1
deletion and limitation of glucose. We conclude that the expression of Mug14 is upregulated by Rst2 under the control of the cAMP/PKA signaling pathway, which senses the limitation of glucose.
Coenzyme Q (CoQ, ubiquinone) is an essential component of the electron transport system in aerobic organisms. Human type CoQ10, which has 10 units of isoprene in its quinone structure, is especially ...valuable as a food supplement. Therefore, studying the biosynthesis of CoQ10 is important not only for increasing metabolic knowledge, but also for improving biotechnological production. Herein, we show that Schizosaccharomyces pombe utilizes p-aminobenzoate (PABA) in addition to p-hydroxybenzoate (PHB) as a precursor for CoQ10 synthesis. We explored compounds that affect the synthesis of CoQ10 and found benzoic acid (Bz) at >5 μg/mL inhibited CoQ biosynthesis without accumulation of apparent CoQ intermediates. This inhibition was counteracted by incubation with a 10-fold lower amount of PABA or PHB. Overexpression of PHB-polyprenyl transferase encoded by ppt1 (coq2) also overcame the inhibition of CoQ biosynthesis by Bz. Inhibition by Bz was efficient in S. pombe and Schizosaccharomyces japonicus, but less so in Saccharomyces cerevisiae, Aureobasidium pullulans, and Escherichia coli. Bz also inhibited a S. pombe ppt1 (coq2) deletion strain expressing human COQ2, and this strain also utilized PABA as a precursor of CoQ10. Thus, Bz is likely to inhibit prenylation reactions involving PHB or PABA catalyzed by Coq2.
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