title: Interleukin-8 is regulated by miR-203 at the posttranscriptional level in primary human keratinocytes.
abstract: Background: MicroRNAs are important posttranscriptional regulators of gene expression. MiR-203 is a miRNA preferentially expressed in the skin, and an important regulator of keratinocyte differentiation. MiR-203 has been implicated in skin diseases, in particular in psoriasis in which it is overexpressed, and in basal cell carcinoma where it acts as a tumor suppressor miRNA. Objectives: To identify novel targets for miR-203 that may be relevant in skin physiology and diseases. Materials & Methods: Bioinformatics was used to identify putative miR-203 targets among genes expressed in keratinocytes. Interleukin-8 (IL-8) gene expression and concentration in keratinocyte medium was measured by quantitative real-time PCR and ELISA, respectively. For miRNA overexpression, resting or TNF-α-treated primary human keratinocytes were transfected with synthetic precursor of miR-203, or scramble miRNA precursors using Lipofectamine 2000. 3'UTR luciferase reporter experiments were performed to prove the direct miRNA:mRNA interaction. Site-specific mutagenesis was used to mutate the predicted miR-203 binding sites in the 3'UTR of IL-8 gene. Results: Bioinformatic analysis indentified two putative miR-203 binding sites in the 3'UTR of IL-8. MiR-203 suppressed IL-8 mRNA and protein expression in primary human keratinocytes both under resting conditions and after TNF-α treatment. Overexpression of miR-203 suppressed the luciferase activity of a reporter gene fused with the IL-8 3'UTR. The suppressive effect was abolished when the predicted binding sites of miR-203 on IL-8 3'UTR were mutated. Conclusion: We identify IL-8 as a novel target of miR-203 for posttranscriptional suppression. These findings may have relevance in diseases in which miR-203 and IL-8 expression are deregulated.
authors.affiliation: Unit of Dermatology and Venereology, Department of Medicine, Karolinska Institutet, Stockholm, Sweden.
authors.forename: Tianling
authors.initials: T
authors.lastname: Wei
authors.forename: Ning
authors.initials: N
authors.lastname: Xu
authors.forename: Florian
authors.initials: F
authors.lastname: Meisgen
authors.forename: Mona
authors.initials: M
authors.lastname: Ståhle
authors.forename: Enikö
authors.initials: E
authors.lastname: Sonkoly
authors.forename: Andor
authors.initials: A
authors.lastname: Pivarcsi
keywords: IL-8; chemokine; cytokines; microRNAs; non-melanoma skin cancer; psoriasis; skin