An ovine hepatorenal fibrocystic model of a Meckel-like syndrome associated with dysmorphic primary cilia and TMEM67 mutations

Abstract

Meckel syndrome (MKS) is an inherited autosomal recessive hepatorenal fibrocystic syndrome, caused by mutations in TMEM67, characterized by occipital encephalocoele, renal cysts, hepatic fibrosis, and polydactyly. Here we describe an ovine model of MKS, with kidney and liver abnormalities, without polydactyly or occipital encephalocoele. Homozygous missense p.(Ile681Asn; Ile687Ser) mutations identified in ovine TMEM67 were pathogenic in zebrafish phenotype rescue assays. Meckelin protein was expressed in affected and unaffected kidney epithelial cells by immunoblotting, and in primary cilia of lamb kidney cyst epithelial cells by immunofluorescence. In contrast to primary cilia of relatively consistent length and morphology in unaffected kidney cells, those of affected cyst-lining cells displayed a range of short and extremely long cilia, as well as abnormal morphologies, such as bulbous regions along the axoneme. Putative cilia fragments were also consistently located within the cyst luminal contents. The abnormal ciliary phenotype was further confirmed in cultured interstitial fibroblasts from affected kidneys. These primary cilia dysmorphologies and length control defects were significantly greater in affected cells compared to unaffected controls. In conclusion, we describe abnormalities involving primary cilia length and morphology in the first reported example of a large animal model of MKS, in which we have identified TMEM67 mutations.

mtor Haploinsufficiency Ameliorates Renal Cysts and Cilia Abnormality in Adult Zebrafish tmem67 Mutants

“…For instance, bpck mice, wpk rats, and human MKS3 fetuses have elongated renal cilia; murine Tmem67 mutants exhibit shorter and fewer cilia; and ovine TMEM67 mutants display both very long and very short cilia in their kidneys. [9][10][11]13 Moreover, compared with those from wild-type mice, mouse embryonic fibroblasts derived from Tmem67-knockout mice present with longer, normal, or no cilia; similar findings have been obtained with cells expressing Tmem67 short hairpin RNA. 11,13,14,17 Therefore, additional studies with advanced imaging methods are needed to define ciliary defects and clarify the relationship between ciliary defects and cyst development.…”

“…[5][6][7] Spontaneous animal models have been used to study MKS3, including the wpk rat carrying a naturally occurring, single-point mutation in the Tmem67 gene; the bpck mouse with the causative Tmem67 gene encompassed in a large 245-kb deletion; and even a sheep model. [8][9][10] The murine models capture some characteristics of patients with MKS3, such as polycystic kidneys and hydrocephalus, but not encephalocele, biliary abnormalities, or polydactyly; whereas the sheep model has the characteristic hepatorenal disease. 8 -12 A targeted knockout mouse (Tmem67 tm1[Dgen/H] ) has also been generated that exhibits pulmonary hypoplasia, cardiac malformation, kidney cysts, and some encephalocele, and that dies by postnatal day 1 due to pulmonary and/or cardiac defects.…”

A human patient-derived cellular model of Joubert syndrome reveals ciliary defects which can be rescued with targeted therapies

“…Cells depleted of KIF7 (JBTSF12) and KIAA0556 (JBTS26) also have extended axonemes ( 34 , 35 ). The KIF7 deficient cilia were both long and had irregular bends along their length and cilia from fibroblasts derived from ovine kidney with mutations in TMEM67 (JBTS6) have abnormally long cilia, with bulbous defects along the axonemes and sharp kinks ( 36 ).…”

Histomorphology and Immunohistochemistry of a Congenital Nephromegaly Demonstrate Concurrent Features of Heritable and Acquired Cystic Nephropathies in a Girgentana Goat (Capra falconeri)

“…Ruminant models provide several advantages in terms of genetics, metabolism, surgical procedures, and pharmacologic treatments of perinatal diseases [ 38 ]. TMEM67-gene was figured out to host the causative mutation of Meckel's syndrome in humans using an ovine research model [ 39 ]. These studies emphasize the relevance of comprehensive and comparative investigations of PKD.…”