Molecular cloning, nucleotide sequence, and abscisic acid induction of a suberization-associated highly anionic peroxidase

Abstract

A highly anionic peroxidase induced in suberizing cells was suggested to be the key enzyme involved in polymerization of phenolic monomers to generate the aromatic matrix of suberin. The enzyme encoded by a potato cDNA was found to be highly homologous to the anionic peroxidase induced in suberizing tomato fruit. A tomato genomic library was screened using the potato anionic peroxidase cDNA and one genomic clone was isolated that contained two tandemly oriented anionic peroxidase genes. These genes were sequenced and were 96% and 87% identical to the mRNA for potato anionic peroxidase. Both genes consist of three exons with the relative positions of their two introns being conserved between the two genes. Primer extension analysis showed that only one of the genes is expressed in the periderm of 3 day wound-healed tomato fruits. Southern blot analyses suggested that there are two copies each of the two highly homologous genes per haploid genome in both potato and tomato. Abscisic acid (ABA) induced the accumulation of the anionic peroxidase transcripts in potato and tomato callus tissues. Northern blots showed that peroxidase mRNA was detectable at 2 days and was maximal at 8 days after transfer of potato callus to solid agar media containing 10(-4) M ABA. The transcripts induced by ABA in both potato and tomato callus were identical in size to those induced in wound-healing potato tuber and tomato fruit. The anionic peroxidase peptide was detected in extracts of potato callus grown on the ABA-containing media by western blot analysis. The results support the suggestion that stimulation of suberization by ABA involves the induction of the highly anionic peroxidase.

Developmentally regulated expression of the wound‐ and pathogen‐responsive tomato anionic peroxidase in green fruits

“…RNAs were gel-blotted and hybridized with a (32P)DNA probe corresponding to exon-1 of tap-1 (Roberts and Kolattukudy, 1989) and specific hybridization bands were quantified using a phosphor-imaging system (Figure 2b). The constitutive expression of tap in green fruits undergoes an approximately twofold increase during the developmental progression from immature to mature green fruits.…”

“…It has previously been reported (McGlasson, 1978) that the pericarp of maturing green fruits contains high levels of auxin, an increasing concentration of ABA (a known inducer of tap; Roberts and Kolattukudy, 1989), a very low concentration of ethylene and low but variable levels of cytokinin. Studies were conducted in which 72 h detached green fruits were supplied through their pedicels with either water or a nutrient solution containing one of these various growth regulators.…”

“…RNA gel-blots were probed using a 490 bp (32P)DNA corresponding to exon-1 of tap-1; this DNA was obtained from pTAP4.5 via digestion with restriction endonucleases Dral and Nsil (Roberts and Kolattukudy, 1989). In some cases TAP-specific band intensities were quantified using an ImageQuantTM phosphor-imaging system (Molecular Dynamics, Sunnyvale, CA).…”

“…However, the expression of tap is induced to high levels in tissues responding to mechanical wounding, pathogen assault or treatment with preparations of ABA or fungal elicitors (Mohan and Kolattukudy, 1990;Robb et al, 1991;Roberts and Kolattukudy, 1989). Using near-isogenic resistant and susceptible tomato varieties, we have demonstrated previously that when petioles or cell cultures derived from the resistant plants were exposed to Verficillium albo-atrum conidia or cell-free elicitor preparations, tap gene expression was rapidly induced to high levels.…”

Choline oxidase, a catabolic enzyme in Arthrobacter pascens, facilitates adaptation to osmotic stress in Escherichia coli

“…Thus, the cox gene may be suitable for enhancing osmotolerance in biological systems of interest. Native or, if necessary, heterologous H202-scavenging systems may be used to detoxify the additional H202 resulting from choline oxidation, and genes encoding catalases and peroxidases have been isolated from a number of sources (13,38,41,50). The cloning and overexpression of the cox gene also have applications in clinical biochemistry, where choline oxidase is used for estimation of choline-containing phospholipids in serum (48) and amniotic fluid (10).…”