Figure 2

Generation and characterization of podocyte-specific Med22 mouse line. (A) A mouse line carrying a cassette with lacZ and neomycin-resistance genes, as well as FRT and LoxP sites targeting the exon 3 of the Med22 gene, was crossed with ThIRES-Cre line to generate a conventional knockout allele, and with a FLP-deleter line to generate a conditional knockout allele. Floxed mouse line was crossed with a NPHS2-cre line to generate podocyte-specific knockout line. (B) Genotyping of conditional allele generated a 580 bp product, whereas wild type produced a 479 bp band. (C) In control mice, Med22 mRNA spanning exon 2–4 and exon 1–2 were amplified in glomerular, lung and spleen tissue. In pod-Med22 mice, the same products were amplified but also an additional shorter variant in glomerular tissue was detected with primers targeting exons 2 and 4. This corresponded in size to the skipping of exon 3. (D) No significant albuminuria is detected in pod-Med22 mice at 8 weeks of age as detected by measuring urine albumin/creatinine ratios. At 12 weeks, pod-Med22 mice show massive albuminuria that further progresses by 16 weeks of age. Blood urea nitrogen levels were unchanged in pod-Med22 mice at 8 weeks of age but were significantly elevated at 16 weeks of age. Pod-Med22 mice start to die prematurely at 15 weeks of age and no mice survive past 20 weeks. ns not significant, **p < 0.01, ***p < 0.001.