Co-regulation of intragenic microRNA miR-153 and its host gene Ia-2 ß: identification of miR-153 target genes with functions related to IA-2ß in pancreas and brain.

ABSTRACT

AIMS/HYPOTHESIS: We analysed the genomic organisation of miR-153, a microRNA embedded in genes that encode two of the major type 1 diabetes autoantigens, islet-associated protein (IA)-2 and IA-2ß. We also identified miR-153 target genes that correlated with IA-2ß localisation and function. METHODS: A bioinformatics approach was used to identify miR-153's genomic organisation. To analyse the co-regulation of miR-153 and IA-2ß, quantitative PCR analysis of miR-153 and Ia-2ß (also known as Ptprn2) was performed after a glucose stimulation assay in MIN6B cells and isolated murine pancreatic islets, and also in wild-type Ia-2 (also known as Ptprn), Ia-2ß single knockout and Ia-2/Ia-2ß double knockout mouse brain and pancreatic islets. Bioinformatics identification of miR-153 target genes and validation via luciferase reporter assays, western blotting and quantitative PCR were also carried out. RESULTS: Two copies of miR-153, miR-153-1 and miR-153-2, are localised in intron 19 of Ia-2 and Ia-2ß, respectively. In rodents, only miR-153-2 is conserved. We demonstrated that expression of miR-153-2 and Ia-2ß in rodents is partially co-regulated as demonstrated by a strong reduction of miR-153 expression levels in Ia-2ß knockout and Ia-2/Ia-2ß double knockout mice. miR-153 levels were unaffected in Ia-2 knockout mice. In addition, glucose stimulation, which increases Ia-2 and Ia-2ß expression, also significantly increased expression of miR-153. Several predicted targets of miR-153 were reduced after glucose stimulation in vitro, correlating with the increase in miR-153 levels. CONCLUSIONS/INTERPRETATION: This study suggests the involvement of miR-153, IA-2ß and miR-153 target genes in a regulatory network, which is potentially relevant to insulin and neurotransmitter release.