title: A phosphomimetic mutant of RelA/p65 at Ser536 induces apoptosis and senescence: An implication for tumor-suppressive role of Ser536 phosphorylation.
abstract: Hundreds of NF-κB inhibitors have been developed for cancer therapy, but their clinical efficacy is unsatisfactory. Here we show that the phosphorylation activation at Ser536 of RelA/p65 protein, a main subunit in the NF-κB family, may play a tumor-suppressive role. In normal colon mucosa, RelA/p65 phosphorylation at Ser536 was increasingly increased with the maturation and apoptotic shedding of epithelial cells, but the phosphorylation at Ser536 was decreased in colon cancer. In colon (HCT116 p53 wt and p53 -/-), breast (MCF7), and prostate (LNCaP and DU145) cancer cells, a phosphomimetic mutation of RelA/p65 at Ser536 (named p65/S536D) triggered dramatic apoptosis through affecting expression of a wide range of cell death/survival genes, such as Bim, Puma, Noxa, Bcl-2 and survivin. In HCT116 cells, p65/S536D mutant upregulated Fas, insulted mitochondrial membrane potential, and triggered cleavage and activation of caspase-3, 7, 8 and 9. A FasL neutralizing antibody (NOK1) prevented cell death induced by the p65/S536D. A pan inhibitor of caspases, Z-VAD-FMK (20 μM), blocked caspase-mediated mitochondrial membrane depolarization. This p65/S536D also triggered senescence in HCT116 cells through a p16-dependent pathway, but not in MFC7 due to lack of p16. Intratumoral delivery of the p65/S536D effectively suppressed tumor growth in nude mice. Together our data suggest that the phosphorylation of RelA/p65 at Ser536 may confer it a tumor-suppressive role by inducing apoptosis and senescence, highlighting the importance of discriminating the function and active status of individual active sites in RelA/p65 when NF-κB inhibitors are considered for targeted therapy of cancer.
authors.affiliation: Department of Medical Microbiology, Immunology & Cell Biology, Simmons Cancer Institute, Southern Illinois University School of Medicine, 913 N. Rutledge Street, Springfield, IL, 62794.
authors.forename: Yiwen
authors.initials: Y
authors.lastname: Bu
authors.affiliation: Department of Medical Microbiology, Immunology & Cell Biology, Simmons Cancer Institute, Southern Illinois University School of Medicine, 913 N. Rutledge Street, Springfield, IL, 62794.
authors.forename: Xiaoning
authors.initials: X
authors.lastname: Li
authors.affiliation: Department of Medical Microbiology, Immunology & Cell Biology, Simmons Cancer Institute, Southern Illinois University School of Medicine, 913 N. Rutledge Street, Springfield, IL, 62794.|Division of Stem Cell Regulation and Application, State Key Laboratory of Chinese Medicine Powder and Medicine Innovation in Hunan (Incubation), Hunan University of Chinese Medicine, Changsha, Hunan, 410208, China.
authors.forename: Yingchun
authors.initials: Y
authors.lastname: He
authors.affiliation: Department of Medical Microbiology, Immunology & Cell Biology, Simmons Cancer Institute, Southern Illinois University School of Medicine, 913 N. Rutledge Street, Springfield, IL, 62794.
authors.forename: Chenfei
authors.initials: C
authors.lastname: Huang
authors.affiliation: Department of Medical Microbiology, Immunology & Cell Biology, Simmons Cancer Institute, Southern Illinois University School of Medicine, 913 N. Rutledge Street, Springfield, IL, 62794.
authors.forename: Yi
authors.initials: Y
authors.lastname: Shen
authors.affiliation: Department of Medical Microbiology, Immunology & Cell Biology, Simmons Cancer Institute, Southern Illinois University School of Medicine, 913 N. Rutledge Street, Springfield, IL, 62794.
authors.forename: Yu
authors.initials: Y
authors.lastname: Cao
authors.affiliation: Division of Stem Cell Regulation and Application, State Key Laboratory of Chinese Medicine Powder and Medicine Innovation in Hunan (Incubation), Hunan University of Chinese Medicine, Changsha, Hunan, 410208, China.
authors.forename: Dan
authors.initials: D
authors.lastname: Huang
authors.affiliation: Division of Stem Cell Regulation and Application, State Key Laboratory of Chinese Medicine Powder and Medicine Innovation in Hunan (Incubation), Hunan University of Chinese Medicine, Changsha, Hunan, 410208, China.
authors.forename: Chuan
authors.initials: C
authors.lastname: Cai
authors.affiliation: Division of Stem Cell Regulation and Application, State Key Laboratory of Chinese Medicine Powder and Medicine Innovation in Hunan (Incubation), Hunan University of Chinese Medicine, Changsha, Hunan, 410208, China.
authors.forename: Yuhong
authors.initials: Y
authors.lastname: Wang
authors.affiliation: Division of Stem Cell Regulation and Application, State Key Laboratory of Chinese Medicine Powder and Medicine Innovation in Hunan (Incubation), Hunan University of Chinese Medicine, Changsha, Hunan, 410208, China.
authors.forename: Ziqi
authors.initials: Z
authors.lastname: Wang
authors.affiliation: Division of Stem Cell Regulation and Application, State Key Laboratory of Chinese Medicine Powder and Medicine Innovation in Hunan (Incubation), Hunan University of Chinese Medicine, Changsha, Hunan, 410208, China.
authors.forename: Duan-Fang
authors.initials: DF
authors.lastname: Liao
authors.affiliation: Department of Medical Microbiology, Immunology & Cell Biology, Simmons Cancer Institute, Southern Illinois University School of Medicine, 913 N. Rutledge Street, Springfield, IL, 62794.|Division of Stem Cell Regulation and Application, State Key Laboratory of Chinese Medicine Powder and Medicine Innovation in Hunan (Incubation), Hunan University of Chinese Medicine, Changsha, Hunan, 410208, China.
authors.forename: Deliang
authors.initials: D
authors.lastname: Cao
mesh_terms: D000818:Animals; D017209:Apoptosis; D045744:Cell Line, Tumor; D016922:Cellular Senescence; D003110:Colonic Neoplasms / Q000473:pathology*; D006801:Humans; D051379:Mice; D009154:Mutation; D010766:Phosphorylation; D012694:Serine; D051996:Transcription Factor RelA / Q000502:physiology*; D025521:Tumor Suppressor Proteins / Q000502:physiology*
chemical_list: C498143:RELA protein, human; D051996:Transcription Factor RelA; D025521:Tumor Suppressor Proteins; D012694:Serine
keywords: NF-κB; RelA/p65; Ser536 phosphorylation; apoptosis; phosphomimetic mutation; senescence; tumor suppressor