Nitrite Promotes the Growth and Decreases the Lignin Content of <i>indica</i> Rice Calli: A Comprehensive Transcriptome Analysis of Nitrite-Responsive Genes during <i>In Vitro</i> Culture of Rice

As both major macronutrients and signal molecules, nitrogen metabolites, such as nitrate and nitrite, play an important role in plant growth and development. In this study, the callus growth of indica rice cv. 9311 was significantly enhanced by nitrite, whereas the soluble protein content remained unchanged. The deep RNA sequencing technology (RNA-seq) showed that the transcriptional profiles of cv. 9311 calli were significantly changed after adding nitrite to the nitrate-free medium, and these nitrite-responsive genes were involved in a wide range of plant processes, particularly in the secondary metabolite pathways. Interestingly, most of the genes involved in phenylpropanoid-related pathways were coordinately down-regulated by nitrite, such as four cinnamoyl-CoA reductase, and these in turn resulted in the decrease of lignin content of indica calli. Furthermore, several candidate genes related to cell growth or stress responses were identified, such as genes coding for expansins, SMALL AUXIN UP RNA (SAUR) and HSP20s, and these suggested that nitrite could probably serve as a transcriptome signal to enhance the indica calli growth by regulation of various downstream genes expression. This study contributes to a better understanding of the function of nitrite during the process of plant tissue culture and could aid in the application of this technology to improved indica genetic transformation efficiency.作为主要的大量营养元素（macronutrients）与信号分子，氮代谢物（nitrogen metabolites，如硝酸盐（nitrate）、亚硝酸盐（nitrite））在植物生长发育过程中发挥关键作用。本研究发现，籼稻（indica rice）品种9311的愈伤组织（callus）生长可被亚硝酸盐显著促进，而可溶性蛋白含量未发生显著变化。深度RNA测序技术（RNA-seq）分析结果显示，在无硝酸盐培养基中添加亚硝酸盐后，9311愈伤组织的转录组谱（transcriptional profiles）发生显著改变；这些亚硝酸盐响应基因参与了广泛的植物生理过程，尤其富集于次生代谢物通路（secondary metabolite pathways）。值得注意的是，多数与苯丙烷类相关通路（phenylpropanoid-related pathways）相关的基因可被亚硝酸盐协同下调，例如4个肉桂酰辅酶A还原酶（cinnamoyl-CoA reductase）编码基因，这一变化进而导致籼稻愈伤组织的木质素（lignin）含量降低。此外，本研究还鉴定出若干与细胞生长或胁迫响应相关的候选基因，包括扩张蛋白（expansins）、生长素上调小RNA（SMALL AUXIN UP RNA，SAUR）以及热激蛋白20家族（HSP20s）的编码基因，这表明亚硝酸盐或可作为一类转录组信号，通过调控多种下游基因的表达来促进籼稻愈伤组织的生长。本研究有助于深化对亚硝酸盐在植物组织培养过程中功能的理解，可为该技术应用于提升籼稻遗传转化效率提供理论支撑。