MicroRNA-192 in diabetic kidney glomeruli and its function in TGF-β-induced collagen expression via inhibition of E-box repressors

• Published in: Proceedings of the National Academy of Sciences - PNAS 2007-02, Vol.104 (9), p.3432-3437
• Main Authors: Kato, Mitsuo, Zhang, Jane, Wang, Mei, Lanting, Linda, Yuan, Hang, Rossi, John J, Natarajan, Rama
• Format: Article
• Language: English
• Subjects: Animals; Biological Sciences; Blotting, Western; Cell Line; Chromatin Immunoprecipitation; Collagen - metabolism; Collagen Type I; Collagens; Diabetes complications; Diabetic nephropathies; Diabetic Nephropathies - genetics; Diabetic Nephropathies - metabolism; DNA Primers; Down regulation; E-Box Elements - genetics; Gene Expression Regulation; Homeodomain Proteins - metabolism; Kidneys; Mesangial Cells - metabolism; Mice; MicroRNAs - metabolism; Nerve Tissue Proteins - metabolism; Oligonucleotide Array Sequence Analysis; Repressor Proteins - antagonists & inhibitors; Repressor Proteins - genetics; Reverse Transcriptase Polymerase Chain Reaction; RNA; Transcription Factors - metabolism; Transforming Growth Factor beta - metabolism; Type 1 diabetes mellitus; Type 2 diabetes mellitus; Untranslated regions
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.0611192104

Key features of diabetic nephropathy (DN) include the accumulation of extracellular matrix proteins such as collagen 1-α 1 and -2 (Col1a1 and -2). Transforming growth factor β1 (TGF-β), a key regulator of these extracellular matrix genes, is increased in mesangial cells (MC) in DN. By microarray profiling, we noted that TGF-β increased Col1a2 mRNA in mouse MC (MMC) but also decreased mRNA levels of an E-box repressor, δEF1. TGF-β treatment or short hairpin RNAs targeting δEF1 increased enhancer activity of upstream E-box elements in the Col1a2 gene. TGF-β also decreased the expression of Smad-interacting protein 1 (SIP1), another E-box repressor similar to δEF1. Interestingly, we noted that SIP1 is a target of microRNA-192 (miR-192), a key miR highly expressed in the kidney. miR-192 levels also were increased by TGF-β in MMC. TGF-β treatment or transfection with miR-192 decreased endogenous SIP1 expression as well as reporter activity of a SIP1 3' UTR-containing luciferase construct in MMC. Conversely, a miR-192 inhibitor enhanced the luciferase activity, confirming SIP1 to be a miR-192 target. Furthermore, miR-192 synergized with δEF1 short hairpin RNAs to increase Col1a2 E-box-luc activity. Importantly, the in vivo relevance was noted by the observation that miR-192 levels were enhanced significantly in glomeruli isolated from streptozotocin-injected diabetic mice as well as diabetic db/db mice relative to corresponding nondiabetic controls, in parallel with increased TGF-β and Col1a2 levels. These results uncover a role for miRs in the kidney and DN in controlling TGF-β-induced Col1a2 expression by down-regulating E-box repressors.