Involvement of nuclear factor-kB and Nurr-1 in cytokine-induced transcription of proopiomelanocortin gene in AtT20 corticotroph cells.
OBJECTIVE
The precise mechanism whereby proinflammatory cytokines activate the hypothalamo-pituitary-adrenal axis is still unclear. We examined whether transcription factors nuclear factor (NF)-kappaB and Nurr-1 are involved in the cytokine-induced proopiomelanocortin (POMC) gene expression.


METHODS
The mouse corticotropinoma cell line AtT20 was treated with tumor necrosis factor-alpha (TNF-alpha) or interleukin-1beta (IL-1beta). Real-time PCR, luciferase assay and Western blotting were conducted to assess the gene expression, promoter activity and protein expression in various conditions.


RESULTS
Intrinsic expression of NF-kappaB was confirmed by RT-PCR. An active component of NF-kappaB (p65) was upregulated in the nuclear fraction by both TNF-alpha and IL-1beta treatment in a dose- and time-related manner. These cytokines potently stimulated the promoter activity of NF-kappaB and Nurr-1. We also found rapid upregulation of the Nurr-1 gene and protein after treatment with these cytokines. Cotreatment of the cells with either of the cytokines and corticotropin-releasing hormone resulted in additive effects. Cytokine-induced Nurr-1 transcription and Nurr-1 transcription induced by overexpression of NF-kappaB were both blunted by mutagenesis within the NF-kappaB responsive element, which implies that Nurr-1 upregulation specifically requires NF-kappaB binding to its own DNA-binding site. Proinflammatory cytokines exert positive effects on POMC gene expression, which were inhibited by pretreatment with a specific NF-kappaB inhibitor.


CONCLUSIONS
These results together imply that Nurr-1 expression is a connecting point between neuroendocrine and immune systems in mediating cytokine-induced POMC gene expression.
Takayasu
Shinobu
S
Department of Endocrinology and Metabolism, Hirosaki University School of Medicine, Hirosaki, Japan. stakayas@cc.hirosaki-u.ac.jp
Iwasaki
Yasumasa
Y
Nigawara
Takeshi
T
Asai
Masato
M
Yoshida
Masanori
M
Kageyama
Kazunori
K
Suda
Toshihiro
T
D000324:Adrenocorticotropic Hormone / Q000378:metabolism; D000818:Animals; D001665:Binding Sites / Q000235:genetics; D002460:Cell Line; D016207:Cytokines / Q000378:metabolism* / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D005786:Gene Expression Regulation / Q000502:physiology; D007030:Hypothalamo-Hypophyseal System / Q000166:cytology / Q000378:metabolism*; D007107:Immune System / Q000166:cytology / Q000378:metabolism; D018836:Inflammation Mediators / Q000378:metabolism / Q000494:pharmacology; D053583:Interleukin-1beta / Q000378:metabolism / Q000494:pharmacology; D051379:Mice; D016328:NF-kappa B / Q000235:genetics* / Q000378:metabolism; D009490:Neurosecretory Systems / Q000166:cytology / Q000378:metabolism; D057126:Nuclear Receptor Subfamily 4, Group A, Member 2 / Q000235:genetics* / Q000378:metabolism; D010903:Pituitary Gland, Anterior / Q000166:cytology / Q000378:metabolism*; D010913:Pituitary-Adrenal System / Q000166:cytology / Q000378:metabolism; D011333:Pro-Opiomelanocortin / Q000096:biosynthesis / Q000235:genetics*; D012333:RNA, Messenger / Q000187:drug effects / Q000378:metabolism; D013315:Stress, Psychological / Q000235:genetics / Q000378:metabolism / Q000503:physiopathology; D015533:Transcriptional Activation / Q000502:physiology; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism / Q000494:pharmacology; D015854:Up-Regulation / Q000235:genetics
D016207:Cytokines; D018836:Inflammation Mediators; D053583:Interleukin-1beta; D016328:NF-kappa B; D057126:Nuclear Receptor Subfamily 4, Group A, Member 2; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; D011333:Pro-Opiomelanocortin; D000324:Adrenocorticotropic Hormone