Cell cycle-related kinase regulates mammalian eye development through positive and negative regulation of the Hedgehog pathway

Abstract

Cell cycle-related kinase (CCRK) is a conserved regulator of ciliogenesis whose loss in mice leads to a wide range of developmental defects, including exencephaly, preaxial polydactyly, skeletal abnormalities, and microphthalmia. Here, we investigate the role of CCRK in mouse eye development. Ccrk mutants show dramatic patterning defects, with an expansion of the optic stalk domain into the optic cup, as well as an expansion of the retinal pigment epithelium (RPE) into neural retina (NR) territory. In addition, Ccrk mutants display a shortened optic stalk. These defects are associated with bimodal changes in Hedgehog (Hh) pathway activity within the eye, including the loss of proximal, high level responses but a gain in distal, low level responses. We simultaneously removed the Hh activator GLI2 in Ccrk mutants (Ccrk-/-;Gli2-/-), which resulted in rescue of optic cup patterning and exacerbation of optic stalk length defects. Next, we disrupted the Hh pathway antagonist GLI3 in mutants lacking CCRK (Ccrk-/-;Gli3-/-), which lead to even greater expansion of the RPE markers into the NR domain and a complete loss of NR specification within the optic cup. These results indicate that CCRK functions in eye development by both positively and negatively regulating the Hh pathway, and they reveal distinct requirements for Hh signaling in patterning and morphogenesis of the eyes.

Patterning the Vertebrate Retina with Morphogenetic Signaling Pathways

“…In both Xenopus and chick embryos, Hh activity promotes RPE development (Perron and others 2003; Zhang and Yang 2001). Studies in mouse embryos instead suggest that RPE and neural retina specification occurs only at intermediate and low levels of Hh activity, respectively (Lupu and others 2018). These conclusions are based on the use of mutant mice in which the primary cilium—an essential organelle for Hh signaling reception—does not form properly.…”

The hedgehog pathway and ocular developmental anomalies

“…The analysis of both Ccrk −/− ; Gli2 −/− double mutants showed compensation for the excess distal Shh signaling activity mediated by the loss of Ccrk in terms of ocular vesicle morphology, although at the expense of ventral and midline tissues with normally highest exposure to Shh. Likewise, Ccrk −/− ; Gli3 −/− double mutants show exacerbation of the distal RPE phenotype at the expense of neural retina fate, and total loss of lens specification (Lupu et al 2018), supporting the idea that such effects result from loss of the Gli3-R TF form and expansion of midline fates at the expense of distal tissues.…”

“…Recently published work demonstrates that Ccrk (cell cycle-related kinase)-null mouse mutants show abnormal, shorter cilia, and display a number of corresponding developmental defects, amongst which microphthalmia due to fully penetrant coloboma and frequent loss of the lens primordium (Lupu et al 2018). A result of disrupting Shh-dependent proximo-distal patterning at optic cup stages, malformations and effects on target gene transcription induced by mutant Ccrk confirm the hypothesis that different Shh activity readout levels lead to different ocular fates.…”

Loss-of-function variants in TBC1D32 underlie syndromic hypopituitarism

“…4). Furthermore, the loss of mouse cell cycle-related kinase (CCRK) function, a protein known to interact with TBC1D32 (10), has been shown to result in a highly similar embryonic phenotype to that of the Bromi null mouse, resulting in disrupted optic cup and lens formation, shortened optic stalk, reduced neural retina specification, and ectopic retinal pigment epithelium formation (10,38,39). Although our experiment using a human cell line did not find an association with CCRK, five TBC1D32-interacting proteins (CI114, UHRF1, AURKB, CENPV, SVIL) were associated with positive regulation of cytokinesis and the cell cycle (Fig.…”