title: NF-kappaB p65 subunit mediates lipopolysaccharide-induced Na(+)/I(-) symporter gene expression by involving functional interaction with the paired domain transcription factor Pax8.
abstract: The Gram-negative bacterial endotoxin lipopolysaccharide (LPS) elicits a variety of biological responses. Na(+)/I(-) symporter (NIS)-mediated iodide uptake is the main rate-limiting step in thyroid hormonogenesis. We have recently reported that LPS stimulates TSH-induced iodide uptake. Here, we further analyzed the molecular mechanism involved in the LPS-induced NIS expression in Fisher rat thyroid cell line 5 (FRTL-5) thyroid cells. We observed an increase in TSH-induced NIS mRNA expression in a dose-dependent manner upon LPS treatment. LPS enhanced the TSH-stimulated NIS promoter activity denoting the NIS-upstream enhancer region (NUE) as responsible for the stimulatory effects. We characterized a novel putative conserved kappaB site for the transcription factor nuclear factor-kappaB (NF-kappaB) within the NUE region. NUE contains two binding sites for the transcription factor paired box 8 (Pax8), main regulator of NIS transcription. A physical interaction was observed between the NF-kappaB p65 subunit and paired box 8 (Pax8), which appears to be responsible for the synergic effect displayed by these transcription factors on NIS gene transcription. Moreover, functional blockage of NF-kappaB signaling and site-directed mutagenesis of the kappaB cis-acting element abrogated LPS stimulation. Silencing expression of p65 confirmed its participation as an effector of LPS-induced NIS stimulation. Furthermore, chromatin immunoprecipitation corroborated that NIS is a novel target gene for p65 transactivation in response to LPS. Moreover, we were able to corroborate the LPS-stimulatory effect on thyroid cells in vivo in LPS-treated rats, supporting that thyrocytes are capable of responding to systemic infections. In conclusion, our results reveal a new mechanism involving p65 in the LPS-induced NIS expression, denoting a novel aspect in thyroid cell differentiation.
authors.affiliation: Centro de Investigaciones en Bioquímica Clínica e Inmunología-Consejo Nacional de Investigaciones Científicas y Técnicas (CIBICI-CONICET), Departamento de Bioquímica Clínica, Facultad de Ciencias Químicas, Universidad Nacional de Córdoba, Haya de la Torre y Medina Allende, 5000 Córdoba, Argentina.
authors.forename: Juan Pablo
authors.initials: JP
authors.lastname: Nicola
authors.forename: Magalí
authors.initials: M
authors.lastname: Nazar
authors.forename: Iván Darío
authors.initials: ID
authors.lastname: Mascanfroni
authors.forename: Claudia Gabriela
authors.initials: CG
authors.lastname: Pellizas
authors.forename: Ana María
authors.initials: AM
authors.lastname: Masini-Repiso
mesh_terms: D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D004742:Enhancer Elements, Genetic / Q000235:genetics; D005786:Gene Expression Regulation / Q000187:drug effects*; D020868:Gene Silencing / Q000187:drug effects; D006801:Humans; D008070:Lipopolysaccharides / Q000494:pharmacology*; D008969:Molecular Sequence Data; D000072636:PAX8 Transcription Factor; D051761:Paired Box Transcription Factors / Q000378:metabolism*; D011485:Protein Binding / Q000187:drug effects; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D051381:Rats; D027981:Symporters / Q000235:genetics* / Q000378:metabolism; D013961:Thyroid Gland / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D013972:Thyrotropin / Q000494:pharmacology; D051996:Transcription Factor RelA / Q000378:metabolism*; D014158:Transcription, Genetic / Q000187:drug effects; D015533:Transcriptional Activation / Q000187:drug effects; D015854:Up-Regulation / Q000187:drug effects; D000099209:Sodium-Iodide Symporters
chemical_list: D008070:Lipopolysaccharides; D000072636:PAX8 Transcription Factor; D051761:Paired Box Transcription Factors; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D027981:Symporters; D013972:Thyrotropin; D051996:Transcription Factor RelA; D000099209:Sodium-Iodide Symporters; C083325:PAX8 protein, human