The β-cyanoalanine pathway is involved in the response to water deficit in Arabidopsis thaliana

Abstract

The β-cyanoalanine pathway is primarily responsible for detoxification of excess cyanide produced by plants. Recent evidence suggests that cyanide detoxification via this pathway may be involved in the response and tolerance to water deficit in plants. The aim of this study was to explore this role in Arabidopsis thaliana in greater detail. The first objective was to establish responsiveness of the pathway to the magnitude and duration of water deficit. The second objective was to examine how interruption of single genes (AtCysA1, AtCysC1 and AtNIT4) encoding enzymes of the pathway influenced the ability to metabolize cyanide and withstand water deficit. Arabidopsis plants were exposed to conditions which emulated acute and chronic water deficit, followed by measurement of tissue cyanide concentration, activity of enzymes, and physiological parameters. The results for wild-type Arabidopsis demonstrated a transient increase in cyanide concentration and β-cyanoalanine synthase activity, followed by a decrease in both. The increase in enzyme activity was localized to the tissue in direct proximity to the stress. The knockdown AtCysA1 mutant did not differ from wild-type while AtCysC1 mutants were slightly more sensitive to water deficit. The AtNIT4 mutant was the most sensitive showing decreased growth along with altered chlorophyll content under water deficit as compared to wild-type. Collectively, the results indicated that the pathway is responsive to water deficit although the severity of stress did not alter the nature of the response, implying that the capacity to remove cyanide generated during water deficit may contribute to tolerance to this stress in Arabidopsis.

Determination of the Michaelis–Menten kinetics and the genes expression involved in phyto-degradation of cyanide and ferri-cyanide

“…NIT is another key enzyme responsible for assimilating b-cyanoalanine into either asparagine or aspartate (Piotrowski et al 2001;Machingura et al 2013). Indeed, altered expression of NIT genes encoding enzymes that metabolize b-cyanoalanine was established in two Arabidopsis lines (Howden et al 2009).…”

“…In our current investigation, remarkable up-regulation of genes associated with NIT was observed, suggesting positive response of NIT gene to cyanide exposure. ASPNASE (asparaginase) has been considered as an additional enzymes involved in the conversion of asparagine into aspartate and ammonium (Piotrowski et al 2001;Machingura et al 2013). Evidently, an increase of ASPNASE activity in wheat has been reported during cyanide exposure, suggesting that there may be shared regulatory elements of the b-CAS pathway and ASPNASE .…”

“…The two most prevalent sources of endogenous cyanide are the turnover of cyanogenic glycosides or cyanolipids and ethylene biosynthesis (Machingura et al 2013). Naturally, cyanide in plant tissues is at concentrations below these that cause adverse effects on plants (Whankaew et al 2014) because phytodegradation capacity of cyanide has several orders of magnitude higher than that of production (Manning 1988).…”

“…The principal pathway for functioning cyanide degradation is the b-cyanoalanine pathway, which is initiated by bcyanoalanine synthase (CAS,EC 4.4.1.9) to form intermediates b-cyanoalanine and sulfide from cysteine (Miller and Conn 1980;Maruyama et al 2001;Siegien and Bogatek 2006). The nitrile group of b-cyanoalanine is then converted into either asparagine or aspartate by nitrile hydratase/nitrilase (EC 3.5.5.1) designated in Arabidopsis as nitrilase 4 (NIT4), in conjunction with ammonium (Piotrowski et al 2001;Machingura et al 2013;O'Leary et al 2014). Here, interests have been generated using endogenous source of ammonium stemmed from degradation of exogenous cyanide as nitrogen source for plant nutrition.…”

Resistance to cyanide by salicylate pretreatment in Salix babylonica L.

“…For instance, uptake and translocation of CN -in free and other forms have been observed in different plant species such as Hordeum vulgar, Sorghum bicolor, Eucalyptus spp., Avena sativa and Salix eriocephala (Ebbs et al 2003, 2008, Samiotakis and Ebbs 2004. It has also shown that some of the physiological conditions, such as nitrogen nutrition status and environmental stresses like water deficiency, induce the assimilation pathways of CN - , Machingura et al 2013.…”