Pseudomonas reinekei MT1 has previously been reported to degrade 4-and 5-chlorosalicylate by a pathway with 4-chlorocatechol, 3-chloromuconate, 4-chloromuconolactone, and maleylacetate as intermediates, and a gene cluster channeling various salicylates into an intradiol cleavage route has been reported. We now report that during growth on 5-chlorosalicylate, besides a novel (chloro)catechol 1,2-dioxygenase, C12O ccaA , a novel (chloro)muconate cycloisomerase, MCI ccaB , which showed features not yet reported, was induced. This cycloisomerase, which was practically inactive with muconate, evolved for the turnover of 3-substituted muconates and transforms 3-chloromuconate into equal amounts of cis-dienelactone and protoanemonin, suggesting that it is a functional intermediate between chloromuconate cycloisomerases and muconate cycloisomerases. The corresponding genes, ccaA (C12O ccaA ) and ccaB (MCI ccaB ), were located in a 5.1-kb genomic region clustered with genes encoding trans-dienelactone hydrolase (ccaC) and maleylacetate reductase (ccaD) and a putative regulatory gene, ccaR, homologous to regulators of the IclR-type family. Thus, this region includes genes sufficient to enable MT1 to transform 4-chlorocatechol to 3-oxoadipate. Phylogenetic analysis showed that C12O ccaA and MCI ccaB are only distantly related to previously described catechol 1,2-dioxygenases and muconate cycloisomerases. Kinetic analysis indicated that MCI ccaB and the previously identified C12O salD , rather than C12O ccaA , are crucial for 5-chlorosalicylate degradation. Thus, MT1 uses enzymes encoded by a completely novel gene cluster for degradation of chlorosalicylates, which, together with a gene cluster encoding enzymes for channeling salicylates into the ortho-cleavage pathway, form an effective pathway for 4-and 5-chlorosalicylate mineralization.
Plants are sessile organisms and, as such, their survival relies on their ability to respond quickly all along their life cycle to any kind of environmental stimuli, including abiotic and biotic stresses. In this respect, plants have developed efficient mechanisms of protection and/or adaptation to minimize deleterious effects of stress on their growth and development. In a stress type-dependent manner, external signals are firstly sensed. This step is then followed by the activation of particular signalling pathways, resulting ultimately in the rapid and specific modulation of the plant transcriptome. Currently, transcriptional regulation is considered as a central process in the build-up of plant responses to both abiotic and biotic stresses. Among mechanisms involved in transcriptional regulation, the combined effect of different histone tail post-translational modifications PTMs e.g. acetylation and methylation through the activity of particular histone-modifying enzymes can lead to changes in the local chromatin structure environment and hence the underlying DNA accessibility.By focusing on histone lysine methylation, in this chapter we highlight our current understanding of the transcriptional roles played by chromatin-remodelling mechanisms in regulating plant response/adaptation to different biotic and abiotic stresses.Based on recent advances, we further discuss the stability and transmission of such methylation marks to subsequent generations, with the underlying idea of an epigenetically based transcriptional memory of stresses in plants.
Chloroaromatic pollutants from bleached Kraft pulp mill effluents (BKME) are difficult to degrade, because bacterial strains present in BKME aerobic treatments, only partially degrade these compounds, accumulating the corresponding chlorocatechol intermediates. To improve the catabolic performance of chlorocatecholaccumulating strains, we introduced, by chromosomal insertion, the tfd I CDEF gene cluster from Ralstonia eutropha JMP134 (pJP4). This gene cluster allows dechlorination and channelling of chlorocatechols into the intermediate metabolism. Two bacterial strains, R. eutropha JMP222 and Pseudomonas putida KT2442, able to produce chlorocatechols from 3-chlorobenzoate (3-CB) were used. Acinetobacter lwoffii RB2 isolated from BKME by its ability to grow on guaiacol as sole carbon source and shown to be able to produce the corresponding chlorocatechols from the BKME pollutants 4-, and 5-chloroguaiacol, was also used. The Bobadilla, R. et al. 163tfd I CDEF gene cluster was inserted in the chromosome of these strains using miniTn5-derived vectors that allow expression of the Tfd enzymes driven by the lacI q /P trc or tfdR/P tfd-I regulatory systems, and therefore, responding to the inducers isopropyl-ß-Dthiogalactopyranoside (IPTG) or 3-CB, respectively. Crude extracts of cells from strains JMP222, KT2442 or RB2 engineered with the tfd genes, grown on benzoate and induced with IPTG or 3-CB showed Tfd specific activities of about 15% -80% of that of the strain JMP134. Dechlorination rates for 3-CB or chloroguaiacols correlated with levels of Tfd enzymes. However, none of the strains containing the chromosomal copy of the tfd I CDEF cluster grew on monochloroaromatics as sole carbon source. Experiments with BKME aerobic treatment microcosms showed that the catabolic performance of the engineered bacteria was also lower than the wildtype R. eutropha strain JMP134.
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