Cyanide detoxification in an insect herbivore: Molecular identification of β-cyanoalanine synthases from Pieris rapae

Ohlen, Maike van; Herfurth, Anna-Maria; Kerbstadt, Henrike; Wittstock, Ute

doi:10.1016/j.ibmb.2015.12.004

Cited by 42 publications

(54 citation statements)

References 52 publications

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“…oleracea , cv. Rosella (Brassicaceae) [13]. Larvae of A. cardamines were collected in Braunschweig, Germany, and kept on Alliaria petiolata (Brassicaceae) until dissection.…”

Section: Methodsmentioning

confidence: 99%

“…Three gut-expressed β-substituted alanine synthases (BSAS) with β-cyanoalanine synthase activity, PrBSAS1-PrBSAS3, have been cloned and characterized from Pieris rapae [13]. In phylogenetic trees, they are embedded in bacterial sequences together with uncharacterized proteins from other lepidopterans and with β-cyanoalanine synthase from the two-spotted spider mite, Tetranychus urticae (Trombidiformes: Tetranychidae), Tu-CAS [14].…”

Section: Introductionmentioning

confidence: 99%

“…The mite enzyme has been recruited by horizontal gene transfer from bacterial endosymbionts [14]. As a likely scenario, the presence of BSAS in lepidopteran genomes is due to an independent gene transfer event from bacteria to ancient lepidoptera [13,14]. Thus, arthropod BSAS are among the few known examples of horizontal gene transfer providing herbivores with tools to overcome plant chemical defenses [15].…”

Section: Introductionmentioning

confidence: 99%

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β-Cyanoalanine Synthases and Their Possible Role in Pierid Host Plant Adaptation

Herfurth

Ohlen

Wittstock

2017

Insects

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Cyanide is generated in larvae of the glucosinolate-specialist Pieris rapae (Lepidoptera:Pieridae) upon ingestion of plant material containing phenylalanine-derived glucosinolates as chemical defenses. As these glucosinolates were widespread within ancient Brassicales, the ability to detoxify cyanide may therefore have been essential for the host plant shift of Pierid species from Fabales to Brassicales species giving rise to the Pierinae subfamily. Previous research identified β-cyanoalanine and thiocyanate as products of cyanide detoxification in P. rapae larvae as well as three cDNAs encoding the β-cyanoalanine synthases PrBSAS1-PrBSAS3. Here, we analyzed a total of eight species of four lepidopteran families to test if their cyanide detoxification capacity correlates with their feeding specialization. We detected β-cyanoalanine synthase activity in gut protein extracts of all six species tested, which included Pierid species with glucosinolate-containing host plants, Pierids with other hosts, and other Lepidoptera with varying food specialization. Rhodanese activity was only scarcely detectable with the highest levels appearing in the two glucosinolate-feeding Pierids. We then amplified by polymerase chain reaction (PCR) 14 cDNAs encoding β-cyanoalanine synthases from seven species. Enzyme characterization and phylogenetic analysis indicated that lepidopterans are generally equipped with one PrBSAS2 homolog with high affinity for cyanide. A second β-cyanoalanine synthase which grouped with PrBSAS3 was restricted to Pierid species, while a third variant (i.e., homologs of PrBSAS1), was only present in members of the Pierinae subfamily. These results are in agreement with the hypothesis that the host shift to Brassicales was associated with the requirement for a specialized cyanide detoxification machinery.

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“…oleracea , cv. Rosella (Brassicaceae) [13]. Larvae of A. cardamines were collected in Braunschweig, Germany, and kept on Alliaria petiolata (Brassicaceae) until dissection.…”

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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β-Cyanoalanine Synthases and Their Possible Role in Pierid Host Plant Adaptation

Herfurth

Ohlen

Wittstock

2017

Insects

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“…Cyanide is cytotoxic and kills the cell by inhibiting the mitochondrial electron transport chain enzyme, cytochrome oxidase; it is, thus, an inhibitor of cellular respiration (Ohlen et al, 2016). Through cytochrome oxidase inhibition, cyanide adversely affects higher plants.…”

Section: Author(s) Agree That This Article Remain Permanently Open Acmentioning

confidence: 99%

Comparative studies on the partial purification and characterization of rhodanese from seed and mesocarp of snake tomatoes (Trichosanthes cucumerina Linn.)

Okonji¹,

Fagbohunka²,

Ehigie³

et al. 2017

J. Agric. Biotech. Sustain. Dev.

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Rhodanese enzyme was partially purified and characterized from seed and mesocarp of snake tomato (Trichosanthes cucumerina) using 80% ammonium sulphate precipitation. The enzyme isolated from T. cucumerina mesocarp had a specific activity of 8.8 RU/mg with a percentage yield of 87.8%, while that from T. cucumerina seed had a specific activity of 19.7 RU/mg with a percentage yield of 14.6%. The K m of rhodanese from T. cucumerina mesocarp and seed for sodium thiosulphate (Na 2 S 2 O 3 ) and potassium cyanide (KCN) were 12.5 and 10 mM, respectively. The enzyme from both mesocarp and seed was not inhibited by ammonium persulphate and sodium metabisulphite. The optimum temperature for both the seed and the mesocarp rhodanese was 50°C, while the optimum pH was 8.0 and 7.0, respectively. Rhodanese from T. cucumerina mesocarp was not inhibited by the metal ions at concentrations of 0.01 and 0.001 M, while rhodanese from the seed was inhibited by the metal ions at the two concentrations of each of the salts (KCl, MnCl 2 , NaCl, NiCl 2 , ZnCl 2 and BaCl 2 ) .

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“…This mechanism can be regarded in the context of the molecular evolution of xenobiotic metabolism and resistance in mites (19). The CAS activity found in the gut of some herbivore insects can be the sum of different isoenzymes (20). In addition to its role as a weapon, the cyanide molecule has been proposed to play a regulatory role in plants under certain circumstances (21).…”

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confidence: 99%

A Cyanide-Induced 3-Cyanoalanine Nitrilase in the Cyanide-Assimilating Bacterium Pseudomonas pseudoalcaligenes Strain CECT 5344

Acera

Carmona

Castillo

et al. 2017

Appl Environ Microbiol

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Pseudomonas pseudoalcaligenes CECT 5344 is a bacterium able to assimilate cyanide as a sole nitrogen source. Under this growth condition, a 3-cyanoalanine nitrilase enzymatic activity was induced. This activity was encoded by nit4, one of the four nitrilase genes detected in the genome of this bacterium, and its expression in Escherichia coli enabled the recombinant strain to fully assimilate 3-cyanoalanine. P. pseudoalcaligenes CECT 5344 showed a weak growth level with 3-cyanoalanine as the N source, unless KCN was also added. Moreover, a nit4 knockout mutant of P. pseudoalcaligenes CECT 5344 became severely impaired in its ability to grow with 3-cyanoalanine and cyanide as nitrogen sources. The native enzyme expressed in E. coli was purified up to electrophoretic homogeneity and biochemically characterized. Nit4 seems to be specific for 3-cyanoalanine, and the amount of ammonium derived from the enzymatic activity doubled in the presence of exogenously added asparaginase activity, which demonstrated that the Nit4 enzyme had both 3-cyanoalanine nitrilase and hydratase activities. The nit4 gene is located downstream of the cyanide resistance transcriptional unit containing cio1 genes, whose expression levels are under the positive control of cyanide. Real-time PCR experiments revealed that nit4 expression was also positively regulated by cyanide in both minimal and LB media. These results suggest that this gene cluster including cio1 and nit4 could be involved both in cyanide resistance and in its assimilation by P. pseudoalcaligenes CECT 5344.IMPORTANCE Cyanide is a highly toxic molecule present in some industrial wastes due to its application in several manufacturing processes, such as gold mining and the electroplating industry. The biodegradation of cyanide from contaminated wastes could be an attractive alternative to physicochemical treatment. P. pseudoalcaligenes CECT 5344 is a bacterial strain able to assimilate cyanide under alkaline conditions, thus avoiding its volatilization as HCN. This paper describes and characterizes an enzyme (Nit4) induced by cyanide that is probably involved in cyanide assimilation. The biochemical characterization of Nit4 provides a segment for building a cyanide assimilation pathway in P. pseudoalcaligenes. This information could be useful for understanding, and hopefully improving, the mechanisms involved in bacterial cyanide biodegradation and its application in the treatment of cyanide-containing wastes.

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Cyanide detoxification in an insect herbivore: Molecular identification of β-cyanoalanine synthases from Pieris rapae

Cited by 42 publications

References 52 publications

β-Cyanoalanine Synthases and Their Possible Role in Pierid Host Plant Adaptation

β-Cyanoalanine Synthases and Their Possible Role in Pierid Host Plant Adaptation

Comparative studies on the partial purification and characterization of rhodanese from seed and mesocarp of snake tomatoes (Trichosanthes cucumerina Linn.)

A Cyanide-Induced 3-Cyanoalanine Nitrilase in the Cyanide-Assimilating Bacterium Pseudomonas pseudoalcaligenes Strain CECT 5344

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