Nicotinamide N-methyltransferase (NNMT) regulates the glucocorticoid signaling pathway during the early phase of adipogenesis

Obesity is associated with adipose tissue dysfunction through the differentiation and expansion of pre-adipocytes to adipocytes (hyperplasia) and/or increases in size of pre-existing adipocytes (hypertrophy). A cascade of transcriptional events coordinates the differentiation of pre-adipocytes into fully differentiated adipocytes; the process of adipogenesis. Although nicotinamide N-methyltransferase (NNMT) has been associated with obesity, how NNMT is regulated during adipogenesis, and the underlying regulatory mechanisms, remain undefined. In present study we used genetic and pharmacological approaches to elucidate the molecular signals driving NNMT activation and its role during adipogenesis. Firstly, we demonstrated that during the early phase of adipocyte differentiation NNMT is transactivated by CCAAT/Enhancer Binding Protein beta (CEBPB) in response to glucocorticoid (GC) induction. We found that Nnmt knockout, using CRISPR/Cas9 approach, impaired terminal adipogenesis by influencing the timing of cellular commitment and cell cycle exit during mitotic clonal expansion, as demonstrated by cell cycle analysis and RNA sequencing experiments. Biochemical and computational methods showed that a novel small molecule, called CC-410, stably binds to and highly specifically inhibits NNMT. CC-410 was, therefore, used to modulate protein activity during pre-adipocyte differentiation stages, demonstrating that, in line with the genetic approach, chemical inhibition of NNMT at the early stages of adipogenesis impairs terminal differentiation by deregulating the GC network. These congruent results conclusively demonstrate that NNMT is a key component of the GC-CEBP axis during the early stages of adipogenesis and could be a potential therapeutic target for both early-onset obesity and glucocorticoid-induced obesity.This work was supported by the Spanish Association Against Cancer (PROYE18061FERN to M.F.F.), the Asturias Government (PCTI) co-funding 2018-2023/FEDER (IDI/2018/146 and IDI/2021/000077 to M.F.F.), the Health Institute Carlos III (Plan Nacional de I+D+I) co-funding FEDER (PI18/01527 and PI21/01067 to M.F.F and A.F.F.), CIBERER Acciones Cooperativas y Complementarias Intramurales (ACCI20-35 to M.F.F.), the Fundación General CSIC (0348_CIE_6_E to M.F.F.), the ISCIII (COV00624 to J.R.T. and M.F.F.), ISPA and the Asociación Galbán (2021-052-INTRAMUR GALBAN-GOURR to R.G.U.), ISPA-Jannsen (2021-048-INTRAMURAL NOV-TEVAR to J.R.T.), CSIC (202020E092 to M.F.F), and the European Commission NextGenerationEU, through CSIC’s Global Health Platform (PTI Salud Global) and the Spanish Ministry of Science and Innovation through the Recovery, Transformation and Resilience Plan (SGL2021-03-39 and SGL2021-03-040). The authors are also grateful to the Spanish Ministry of Science and Innovation (PID2021-126663NB-100; CTQ2016-76829-R; PID2020-113473GB-I00), the Regional Government of Andalusia (BIO198 and P18-FR-3487) and the Ramón Areces Foundation (2021-2024). A.R. is supported by CSIC (SOLAUT_00038505 SGL2103040). J.R.T. is supported by a Juan de la Cierva fellowship from the Spanish Ministry of Science and Innovation MCIN/AEI/https://doi.org/10.13039/501100011033 (IJC2018-036825-I). R.F.P. (BP17-114) and P.S.M. (BP17-165) are supported by the Severo Ochoa program. R.G.U. is supported by the Centro de Investigación Biomédica en Red de Enfermedades Raras (CIBERER). M.L.M.-C is supported by grants from Ministerio de Ciencia, Innovación y Universidades (MICINN) PID2020-117116RB-I00 integrated in Plan Estatal de Investigación Científica y Técnica y Innovación (to M.L.M.-C), and co-funded with Fondos FEDER (to M.L.M.-C), Subprograma Retos Colaboración RTC2019-007125-1 (to M.L.M.-C), La Caixa Foundation Program HR17-00601 (to MLM-C), Proyectos Investigación en Salud DTS20/00138 (to M.L.M.-C), Departamento de Industria del Gobierno Vasco (to M.L.M.-C), Elkartek (KK-2020/00008) (to M.L.M.-C). We also acknowledge support from the Institute of Oncology of Asturias (IUOPA, supported by Obra Social Cajastur Liberbank, Spain), the Health Research Institute of Asturias (ISPA-FINBA) and Consorcio Centro de Investigación Biomédica en Red (CIBERER-ISCIII).Peer reviewe