TY - JOUR

T1 - Equilibrative nucleoside transporters mediate the import of nicotinamide riboside and nicotinic acid riboside into human cells

AU - Kropotov, Andrey

AU - Kulikova, Veronika

AU - Nerinovski, Kirill

AU - Yakimov, Alexander

AU - Svetlova, Maria

AU - Solovjeva, Ljudmila

AU - Sudnitsyna, Julia

AU - Migaud, Marie E.

AU - Khodorkovskiy, Mikhail

AU - Ziegler, Mathias

AU - Nikiforov, Andrey

N1 - Funding Information: Funding: A.N. was supported by the Program for Molecular and Cell Biology and Post-Genomic Technologies of the Russian Academy of Sciences. V.K. was supported by the Russian Foundation for Basic Research (grant No. 19-34-60039). M.Z. was supported by the Research Council of Norway (RCN), No. 250395. M.Z., M.E.M., M.K., and A.N. are supported by a network grant (VitaCross) from the RCN. Publisher Copyright: © 2021 by the authors. Licensee MDPI, Basel, Switzerland. Copyright: Copyright 2021 Elsevier B.V., All rights reserved.

PY - 2021/1/30

Y1 - 2021/1/30

N2 - Nicotinamide riboside (NR), a new form of vitamin B3, is an effective precursor of nicoti-namide 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 mem-bers 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.

AB - Nicotinamide riboside (NR), a new form of vitamin B3, is an effective precursor of nicoti-namide 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 mem-bers 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.

KW - Concentrative nucleoside transporters

KW - Equilibrative nucleo-side transporters

KW - Human cells

KW - NAD metabolism

KW - Nicotinamide riboside

KW - Nicotinic acid riboside

UR - http://www.scopus.com/inward/record.url?scp=85100056816&partnerID=8YFLogxK

U2 - 10.3390/ijms22031391

DO - 10.3390/ijms22031391

M3 - Article

AN - SCOPUS:85100056816

VL - 22

SP - 1

EP - 13

JO - International Journal of Molecular Sciences

JF - International Journal of Molecular Sciences

SN - 1422-0067

IS - 3

M1 - 1391

ER -