Kropotov Andrey, Kulikova Veronika, Nerinovski Kirill, Yakimov Alexander, Svetlova Maria, Solovjeva Ljudmila, Sudnitsyna Julia, Migaud Marie E, Khodorkovskiy Mikhail, Ziegler Mathias, Nikiforov Andrey
Institute of Cytology, Russian Academy of Sciences, St. Petersburg 194064, Russia.
Sechenov Institute of Evolutionary Physiology and Biochemistry, Russian Academy of Sciences, St. Petersburg 194223, Russia.
Int J Mol Sci. 2021 Jan 30;22(3):1391. doi: 10.3390/ijms22031391.
Nicotinamide riboside (NR), a new form of vitamin B3, is an effective precursor of nicotinamide adenine dinucleotide (NAD) in human and animal cells. The introduction of NR into the body effectively increases the level of intracellular NAD and thereby restores physiological functions that are weakened or lost in experimental models of aging and various pathologies. Despite the active use of NR in applied biomedicine, the mechanism of its transport into mammalian cells is currently not understood. In this study, we used overexpression of proteins in HEK293 cells, and metabolite detection by NMR, to show that extracellular NR can be imported into cells by members of the equilibrative nucleoside transporter (ENT) family ENT1, ENT2, and ENT4. After being imported into cells, NR is readily metabolized resulting in Nam generation. Moreover, the same ENT-dependent mechanism can be used to import the deamidated form of NR, nicotinic acid riboside (NAR). However, NAR uptake into HEK293 cells required the stimulation of its active utilization in the cytosol such as phosphorylation by NR kinase. On the other hand, we did not detect any NR uptake mediated by the concentrative nucleoside transporters (CNT) CNT1, CNT2, or CNT3, while overexpression of CNT3, but not CNT1 or CNT2, moderately stimulated NAR utilization by HEK293 cells.
烟酰胺核糖（NR）是维生素B3的一种新形式，是人和动物细胞中烟酰胺腺嘌呤二核苷酸（NAD）的有效前体。将NR引入体内可有效提高细胞内NAD水平，从而恢复在衰老实验模型和各种病理状态下减弱或丧失的生理功能。尽管NR在应用生物医药中得到了积极应用，但其进入哺乳动物细胞的机制目前尚不清楚。在本研究中，我们通过在HEK293细胞中过表达蛋白质，并利用核磁共振检测代谢物，表明细胞外NR可通过平衡核苷转运体（ENT）家族的ENT1、ENT2和ENT4成员导入细胞。导入细胞后，NR很容易被代谢生成烟酰胺（Nam）。此外，相同的ENT依赖机制可用于导入NR的脱酰胺形式——烟酸核糖（NAR）。然而，NAR进入HEK293细胞需要刺激其在胞质溶胶中的活性利用，如通过NR激酶磷酸化。另一方面，我们未检测到由集中核苷转运体（CNT）CNT1、CNT2或CNT3介导的任何NR摄取，而CNT3（而非CNT1或CNT2）的过表达适度刺激了HEK293细胞对NAR的利用。
