Nitrate Modulates Fruit Lignification by Regulating CgLAC3 Expression in Pomelo

Lignification of juice sacs is a primary contributor to reductions in fruit quality, with impacts on taste and economic value of pomelo (<i>Citrus grandis</i>). To date, information on the regulation of fruit lignification remains fragmentary. In this study, we first analyzed the relationship between lignification and nutrient status of pomelo juice sacs, which revealed a significant positive correlation between nitrate (NO₃^-) concentration and lignin concentration, with over 60% of lignin accumulation explained by NO₃^- levels in three models of machine learning-based regression. Results from field trails in 11 pomelo orchards, as well as in pear fruits and soybean roots exposed to low or high NO₃^- supplies, further demonstrated that nitrate plays an important role in lignification. Transcriptomic analysis further showed that pomelo <i>laccases</i> (<i>CgLACs</i>) were more intensively up-regulated upon addition of NO₃^- than any of the genes encoding one of the other 12 enzymes involved in lignin biosynthesis. Among the nine identified <i>CgLACs</i>, <i>CgLAC3</i> was the most significantly up-regulated <i>CgLAC</i> in high nitrate treated plants. Over-expressing <i>CgLAC3</i> increased lignin concentrations in both pomelo albedo and soybean hairy roots. Taken together, we conclude that nitrate modulates fruit lignification in pomelo through regulation of <i>CgLAC3</i> expression, which suggests that NO₃^--N fertilization may affect fruit lignification, and thereby can be managed to improve fruit quality.