Nitrogen (N) is associated with amino acid metabolism in higher plants. Theanine is an important amino acid in tea plants. To explore the relationship between theanine metabolism and N conditions, we examined the differentially expressed genes (DEGs), proteins (DEPs), and microRNAs (DEMs) involved in theanine metabolism in tea plant shoots and roots under N sufficiency and deficiency conditions. Transcriptome, proteome, and microRNA analyses were performed on tea plant shoots and roots under N sufficiency and deficiency conditions. The contents of theanine, expression levels of genes involved in theanine metabolism, contents of proteinogenic amino acids, and activity of enzymes were analyzed. The DEP–DEG correlation pairs and negative DEM–DEG interactions related to theanine metabolism were identified based on correlation analyses. The expression profiles of DEGs and negative DEM–DEG pairs related to theanine biosynthesis were consistent with the sequencing results. Our results suggest that the molecular and physiological mechanism of theanine accumulation is significantly affected by N sufficiency and deficiency conditions. The DEGs, DEPs, and DEMs and the activity of the enzymes involved in theanine biosynthesis might play vital roles in theanine accumulation under N sufficiency and deficiency conditions in the shoots and roots of tea plants.

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