Murine Joubert syndrome reveals Hedgehog signaling defects as a potential therapeutic target for nephronophthisis

Abstract

Significance The molecular mechanisms underlying the juvenile onset cystic kidney disease nephronophthisis, remain incompletely understood. Our mutant mouse model identifies abnormal Hedgehog signaling as the primary lesion in nephronophthisis, although currently the perceived knowledge is that aberrant wingless-int signaling is responsible. Primary kidney collecting duct cells isolated from mutant mice with nephronophthisis are morphologically and functionally rescued when Hedgehog signaling is stimulated. This finding was replicated in ex vivo cultured urine-derived renal epithelial cells from a patient with Joubert syndrome and nephronophthisis. Understanding the initial molecular mechanisms of nephronophthisis is a significant advancement of the wider field of ciliopathies and identifies Hedgehog signaling as a potential therapeutic target for these conditions.

DNA replication stress underlies renal phenotypes in CEP290-associated Joubert syndrome

“…LacZ/LacZ H-2Kb-tsA58 +/− and Cep290 +/+ H-2Kb-tsA58 +/− collecting duct cells were isolated from kidneys of 1-month-old transgenic mice (19). Cells were incubated at 33°C in 5% CO 2 .…”

“…URECs were obtained from a patient with JS and a healthy gender-and age-matched control and from additional healthy individuals. URECs were derived as previously described (19,29). All cells were tested for mycoplasma every 2 weeks and were negative.…”

“…After polymerization for 30 minutes at 33°C, warm medium was dripped over the matrix until just covered, and cells were cultured at 33°C, 5% CO 2 . The cells formed spheroids with cleared lumens 3 to 4 days later (19). During the last 18 hours, spheroids were cultured with medium without serum containing 400 nM APH, with or without 200 nM CDK1/2i for 18 hours.…”

“…Cells were lysed in high-salt buffer (300 mM NaCl, 50 mM HEPES, pH 7.5, 0.8% Triton X-100, 8% glycerol, 1 mM EDTA, 1 mM DTT, protease inhibitors, and phosphatase inhibitors). LacZ/LacZ hypomorphic mutation bred on a pure 129/Ola genetic background (F 1 heterozygotes were backcrossed with C57/b6 for 6 generations) were used as previously described (19). Male and female mice were sacrificed at various ages and genotyped.…”

Genetic Disturbances in Patients with Bodily Isomerism from a Single Center: Clinical Implications of Affected Genes and Potential Impact of Ciliary Dyskinesia

“…[19][20][21] Hepatic, renal, and central nervous system malformations and dysfunction may also be a result of ciliary dyskinesia. 20,[22][23][24][25][26][27][28][29][30][31][32] Ciliary dyskinesia and isomerism are now known to be associated with specific genetic mutations, some being common to both. Genes such as Lefty1 and Pitx2 were initially demonstrated to be associated with isomerism, with additional genes subsequently identified.…”