Fig. 6

NUCB2/Nesfatin-1 upregulates SREBP2 and HMGCR expression via the mTORC1 pathway. A The mRNA expression levels of HMGCR and SREBP2 were measured by RT-qPCR in NUCB2 silencing and control cell lines treated with 50 nM rapamycin for 24 h. B Western blot analysis of SREBP2 expression in breast cancer cells treated with rapamycin (0, 20, or 50 nM) for 24 h. C RT-qPCR analysis of HMGCR and SREBP2 mRNA expression in shNUCB2-BT-549 and shNUCB2-MDA-MB-231 stable cell lines treated with exogenous Nesfatin-1 and rapamycin. The NUCB2 knockdown cell lines were starved for 24 h, followed by incubation with exogenous Nesfatin-1 (0, 400, and 1000 pg/mL) for 24 h. The RNA was extracted and used as templates for RT-qPCR. Another group of cells was starved for 24 h, treated with 50 nM rapamycin for 1 h, followed by incubation with 1000 pg/mL Nesfatin-1 for 24 h, and then the RNA was extracted and used as templates for RT-qPCR. D Western blot analysis of SREBP2 protein expression in NUCB2 knockdown cell lines treated with Nesfatin-1 combined with or without rapamycin. The NUCB2 knockdown cell lines were starved for 24 h, followed by incubation with 1000 pg/mL Nesfatin-1 for 24 h. Another group of cells was starved for 24 h, treated with 50 nM rapamycin for 1 h, followed by incubation with 1,000 pg/mL Nesfatin-1 for 24 h, and then the lysates were collected and analyzed by Western blot. The statistical significance was verified using a non-paired t-test. *P < 0.05, **P < 0.01, ***P < 0.001