title: Recombinant R-spondin2 and Wnt3a up- and down-regulate novel target genes in C57MG mouse mammary epithelial cells.
abstract: R-spondins (Rspos) comprise a family of four secreted proteins that have important roles in cell proliferation, cell fate determination and organogenesis. Rspos typically exert their effects by potentiating the Wnt/β-catenin signaling pathway. To systematically investigate the impact of Rspo/Wnt on gene expression, we performed a microarray analysis using C57MG mouse mammary epithelial cells treated with recombinant Rspo2 and/or Wnt3a. We observed the up- and down-regulation of several previously unidentified target genes, including ones that encode proteins involved in immune responses, effectors of other growth factor signaling pathways and transcription factors. Dozens of these changes were validated by quantitative real time RT-PCR. Time course experiments showed that Rspo2 typically had little or no effect on Wnt-dependent gene expression at 3 or 6 h, but enhanced expression at 24 h, consistent with biochemical data indicating that Rspo2 acts primarily to sustain rather than acutely increase Wnt pathway activation. Up-regulation of gene expression was inhibited by pre-treatment with Dickkopf1, a Wnt/β-catenin pathway antagonist, and by siRNA knockdown of β-catenin expression. While Dickkopf1 blocked Rspo2/Wnt3a-dependent down-regulation, a number of down-regulated genes were not affected by β-catenin knockdown, suggesting that in these instances down-regulation was mediated by a β-catenin-independent mechanism.
authors.affiliation: Laboratory of Cellular and Molecular Biology, National Cancer Institute, Bethesda, Maryland, United States of America.
authors.forename: Bolormaa
authors.initials: B
authors.lastname: Baljinnyam
authors.forename: Malgorzata
authors.initials: M
authors.lastname: Klauzinska
authors.forename: Saad
authors.initials: S
authors.lastname: Saffo
authors.forename: Robert
authors.initials: R
authors.lastname: Callahan
authors.forename: Jeffrey S
authors.initials: JS
authors.lastname: Rubin
mesh_terms: D000818:Animals; D002460:Cell Line; D015536:Down-Regulation / Q000187:drug effects; D004847:Epithelial Cells / Q000187:drug effects* / Q000378:metabolism; D005260:Female; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation / Q000187:drug effects*; D008321:Mammary Glands, Animal / Q000187:drug effects* / Q000378:metabolism; D051379:Mice; D046228:Microarray Analysis; D034741:RNA, Small Interfering / Q000494:pharmacology; D011994:Recombinant Proteins / Q000494:pharmacology; D019699:Thrombospondins / Q000494:pharmacology* / Q000502:physiology; D015854:Up-Regulation / Q000187:drug effects; D054928:Validation Studies as Topic; D060509:Wnt3A Protein / Q000494:pharmacology* / Q000502:physiology; D051176:beta Catenin / Q000037:antagonists & inhibitors / Q000235:genetics; D000098907:R-Spondins
chemical_list: D034741:RNA, Small Interfering; D011994:Recombinant Proteins; D019699:Thrombospondins; D060509:Wnt3A Protein; D051176:beta Catenin; C518904:RSPO2 protein, mouse; D000098907:R-Spondins; C555511:Wnt3a protein, mouse