The effects in vivo of mutationally modified uroporphyrinogen decarboxylase in different hem12 mutants of baker's yeast (Saccharomyces cerevisiae)

Abstract

Nine new hem12 haploid mutants of baker's yeast (Saccharomyces cerevisiae), totally or partially deficient in uroporphyrinogen decarboxylase activity, were subjected to both genetic and biochemical analysis. The mutations sites studied are situated far apart within the HEM12 gene located on chromosome IV. Uroporphyrinogen decarboxylase activity in the cell-free extracts of the mutants was decreased by 50-100%. This correlated well with the decrease of haem formation and the increased accumulation and excretion of porphyrins observed in vivo. The pattern of porphyrins (uroporphyrin and its decarboxylation products) accumulated in the cells of mutants partially deficient in uroporphyrinogen decarboxylase activity did not differ significantly, although differences in vitro were found in the relative activity of the mutant enzyme at the four decarboxylation steps. The excreted porphyrins comprised mainly dehydroisocoproporphyrin or pentacarboxyporphyrin. In heterozygous hem12-1/HEM12 diploid cells, a 50% decrease in decarboxylase activity led to an increased accumulation of porphyrins as compared with the wild-type HEM12/HEM12 diploid, which points to the semi-dominant character of the hem12-1 mutation. The biochemical phenotypes of both the haploid and the heterozygous diploid resembles closely the situation encountered in porphyria cutanea tarda, the most common human form of porphyria.

Defense Responses to Tetrapyrrole-Induced Oxidative Stress in Transgenic Plants with Reduced Uroporphyrinogen Decarboxylase or Coproporphyrinogen Oxidase Activity1

“…Deficiency mutations of UROD have been described in humans and are similar to our antisense plants (Elder and Roberts, 1995, and refs. therein), Escherichia coli (Sasarman et al, 1975), and yeast (Kurlandzka et al, 1988). Patients with porphyria disease may suffer from severe light sensitivity as a consequence of accumulated tetrapyrroles.…”

Uroporphyrinogen decarboxylase in Saccharomyces cerevisiae

“…The mutant enzyme from G229. when assayed in cell-free extract, was totally unable to decarboxylate uroporphyrinogen and heptacarboxyporphyrinogen I or 111, while the enzyme from G230 could catalyze very small residual decarboxylation of these substrates into pentacarboxyporphyrinogen [17]; both were inactive with pentacarboxyporphyrinogen I as substrate. The hem12-8 and hem12-9 alleles present in these two mutants were cloned after in vitro PCR amplification of a 1.27-kb DNA fragment encompassing the entire HEM12 coding region plus 107 and 13 nucleotides of the 5'-and 3'-flanking regions, respectively.…”

Analysis of the role of theAspergillus nigeraminolevulinic acid synthase (hemA) gene illustrates the difference between regulation of yeast and fungal haem- and sirohaem-dependent pathways

“…Haem is an essential molecule for almost every organism owing to its requirement as a cofactor of proteins involved in many primary functions like cellular differentiation and gene regulation (Ferreira et al, 1993;Elrod et al, 1997;Panek & O'Brian, 2002;Hamza, 2006). Its biosynthesis in fungi has been extensively studied in Saccharomyces cerevisiae with mutants available for every step within the pathway (Gollub et al, 1977;Urban-Grimal & Labbe-Bois, 1981;Myers et al, 1987;Kurlandzka et al, 1988;Zagorec et al, 1988;Labbe-Bois, 1990;Keng et al, 1992;Amillet & Labbe-Bois, 1995;Camadro & Labbe, 1996;Hoffman et al, 2003). These mutants can be sustained by supplementing hemin to their growth media or by ergosterol or Tween80 addition to supply for essential unsaturated fatty acids (Gollub et al, 1977).…”