Throughout the entire cultivation period, nutrient uptake from the soil is paramount to plants [5,6,18]. Typically, speciation and availability of nutrients present in the liquid phase of soils vary greatly in both space and time, especially in areas contaminated with various chemicals [5,6,18,24]. To tackle changes in their living environment efficiently, plants have evolved sophisticated sensor systems and signaling transition panels [9]. Nitrogen (N) fertilization is a limiting factor in determining the yield and quality of a crop [11] because it is essential for all physiological and biochemical processes in plants [3]. Nitrate (NO3−) and ammonium (NH4+) are the two major inorganic N sources for crops [33], with distinct uptake, transport, and assimilation mechanisms [8,25].

Free cyanide (CN−), an N-containing chemical, is regarded as the golden boy of gold extraction [40]. Cycanide levels in industrial effluents range from 5 to 600 mg L−1, depending on various activities, including electroplating, metal finishing and hardening, steel and printed circuit board manufacturing [19,26,27]. More than 100,000 tons of CN− are anticipated to enter environmental matrixes each year through these sources [35], polluting the ecosystem and poisoning living organisms. In fact, excessive CN− is detrimental to plants because it can inhibit the electron transfer from cytochrome c to oxygen in mitochondrion, deprive mitochondrial oxygenic respiration, and decrease the photosynthetic efficiency of chloroplasts [13]. During waste/waste-water treatment processes, the utilization of various N-containing pollutants as a nutrition source has attracted great attention during the past two decades [10]. Several research groups have attempted to investigate the incorporation of exogenous CN− assimilation into the N nutritional cycle in plants [7,20,35]. In comparison, non-toxic concentrations of CN− did not affect the N content in plant tissues in the presence of NH4+ or NO3− [32], providing intriguing evidence for co-assimilation of CN− and other N sources.

Amino acids (AAs) are chief elements in the N nutritional cycle in plants [15]. Among these, proline (Pro) and its associated species, such as glutamate (Glu), arginine (Arg), and ornithine (Orn), act as an N-cycle balancer, osmoregulation substance, ROS scavenger, and membrane stabilizer to alleviate stress-induced repercussions in plants [39]. Additionally, the level and transport of these AAs in subcellular organelles are crucial for adaptation to external N status [28,37], especially during exposure to N-containing toxins [35]. It is known that CN− can be converted into β-cyanoalanine in the presence of β-cyanoalanine synthase (β-CAS). Subsequently, β-cyanoalanine is prone to degradation into asparagine or aspartate with the production of NH4+ [7,20]. Hence, these degradation products and intermediates could enter into the N nutritional cycle in plants (Fig. 1), affecting AA composition in plant tissues. Until now, however, very little information is available on spatial-temporal variations of the innate pool of Pro and its synthesis-related amino acids (Pro-AAs, i.e., Glu, Arg, and Orn) in crop plants during CN− assimilation in the presence of other readily absorbed N compounds.

Rice (Oryza sativa L.), one of the leading crops, feeds more than half of the world's population [4]. It is also a promising candidate for ecological risk assessment and environmental monitoring investigations. Therefore, a hydroponic experiment was carried out to investigate biochemical and molecular responses in rice seedlings exposed to different KCN concentrations (0, 1.0, 2.0, and 3.0 mg CN/L) with fertilization of either NO3− or NH4+ following a 2-d and 4-d exposure. The objectives of our study were as follows: 1) to analyze the effects of different N sources on the relative growth rate (RGR) of rice seedlings after KCN exposure; 2) comparative analysis of the innate Pro, Glu, Arg and Orn levels in rice seedlings between KCN + NO3− and KCN + NH4+ treatments; 3) to clarify the differential expression of Pro metabolism-related genes in rice seedlings between KCN + NO3− and KCN + NH4+ treatments; 4) to uncover N utilization strategies in rice seedlings during detoxification of exogenous KCN.