NtABCG6L contributes to root-to-leaf nicotine translocation and leaf nicotine accumulation, and interacts with NtWBC in tobacco

Tobacco ( Nicotiana tabacum L.) is a globally important cash crop and the predominant natural source of nicotine. Previously, integrated transcriptomic and proteomic analyses identified NtABCG6L , an ABCG-family transporter, as a candidate mediator of nicotine transport. However, its specific function and underlying mechanism of action remain unresolved. In the present study, NtABCG6L co-localised with a plasma membrane marker and is preferentially expressed in roots. Root NtABCG6L transcript abundance was positively correlated with the nicotine translocation coefficient across developmental stages. Overexpression markedly increased nicotine translocation efficiency, leading to elevated nicotine accumulation in leaves. Yeast assays supported the involvement of NtABCG6L in nicotine transport. Transcriptomic profiling revealed that NtABCG6L overexpression triggered the co-expression of additional transport-related genes, thereby synergistically enhancing nicotine movement. Reduced root nicotine levels relieved feedback repression of negative regulators of nicotine biosynthesis ( ASNA , speD and E TR ), stimulated de novo nicotine synthesis, and promoted further transfer to leaves. Together, these results indicate that NtABCG6L contributes to root-to-leaf nicotine translocation and interacts with NtWBC in tobacco. • NtABCG6L resides in the plasma membrane, shows root-preferential expression, and its transcript abundance tracks closely with tissue nicotine levels. • Transgenic overexpression of NtABCG6L significantly increases root-to-leaf nicotine translocation efficiency, elevating nicotine accumulation in leaves. • NtABCG6L overexpression co-induces additional transport genes and relieves feedback repression on biosynthetic regulators (ASNA, speD, ETR), stimulating de novo nicotine synthesis to sustain the heightened flux.