AN INDIVIDUAL OF THE VOLINTSEVO PERIOD FROM KURILOVKA: THE FIRST ARCHAEOGENETIC DATA Индивид волынцевского времени из Куриловки: первые археогенетические данные

Nitrogen (N) availability strongly influences tea growth and the accumulation of quality-related metabolites, but long-term responses to withdrawal of external N can vary among cultivars. In this study, we characterized phenotypic, spectral, biochemical, and transcriptomic responses of four Camellia sinensis accessions—‘Kolkhida’, ‘Karatum’, and γ-irradiation-derived mutant forms #582 and #619—during two, four, or six months of complete withdrawal of exogenous NH₄NO₃ in greenhouse sand culture. Phenotypic, spectral, and biochemical measurements included all four accessions at all three time points; RNA-seq data were available for all four accessions were represented at four and six months, whereas two-month point were available only for ‘Kolkhida’ and ‘Karatum’. Nitrogen withdrawal decreased L-theanine and caffeine and generally increased simple and gallated catechins, consistent with a change in carbon/nitrogen balance towards phenolic metabolism. The four cultivars exhibited markedly distinct responses. ‘Karatum’ showed the optimum resilience, with minimal phenotypic alteration and a neutral transcriptional response, while #582 demonstrated a stronger stress-associated phenotype and metabolic response. Within the available RNA-seq comparisons, DEG counts were lower at four months than at six months, while the two-month datasets for ‘Kolkhida’ and ‘Karatum’ contained the largest DEG sets for those two cultivars, suggesting a phased acclimation process. CsELIP1, CsSRG1, CsHHO2/4, CsNRT2.4, and CsTAT2 were identified as potential candidate genes associated with nitrogen limitation, flavonoid regulation, and amino acid metabolism. Our findings show that N-deficiency reactions in tea are highly cultivar- and time-dependent, making single-time-point evaluations insufficient. We propose ‘Karatum’ as a promising candidate for further evaluation in low-nitrogen dose–response and field trials based on our combined biochemical and transcriptional findings.

Nitrogen deficiency is one of the key factors limiting the growth and raw-material quality of tea plants; however, cell-autonomous mechanisms underlying the response of Camellia sinensis (L.) Kuntze to reduced nitrogen availability remain insufficiently studied. This study characterized biochemical and transcriptomic changes in callus cultures of three tea cultivars (‘Kolkhida’, ‘Karatum’, and cultivar #582) after two months of cultivation on Murashige–Skoog medium lacking nitrogen-containing components. Nitrogen withdrawal caused visible stress symptoms, reduced total nitrogen in ‘Kolkhida’ and ‘Karatum’ (total nitrogen was not determined in cultivar #582), and altered the accumulation of L-theanine, caffeine, and catechins in a cultivar-dependent manner. ‘Kolkhida’ showed decreases in simple catechins, L-theanine, and caffeine; ‘Karatum’ showed increases in simple and gallated catechins together with a decrease in L-theanine; and cultivar #582 showed smaller changes in most measured biochemical traits. RNA-seq identified 1898 differentially expressed genes (DEGs) in ‘Kolkhida’, 4551 in ‘Karatum’, and 9869 in cultivar #582. The enriched Gene Ontology term “response to karrikin” (GO:0080167) was common to all three cultivars, whereas cultivar-specific profiles differed substantially. In the flavonoid biosynthesis pathway, key phenylpropanoid genes were predominantly downregulated in cultivar #582 but induced in ‘Kolkhida’ and ‘Karatum’. RT-qPCR confirmed the direction of change for selected genes. Under the tested in vitro conditions, ‘Kolkhida’ showed the strongest biochemical deterioration, whereas cultivar #582 combined the greatest biochemical stability with the most extensive transcriptomic remodeling. Because total nitrogen was not measured in cultivar #582, its apparent tolerance remains provisional. Tea callus cultures therefore provide a controlled discovery system for cell-autonomous candidate responses, but cultivar rankings and candidate markers require validation in independent callus lines and whole plants.