Joubert Syndrome in French Canadians and Identification of Mutations in CEP104

Abstract

Joubert syndrome (JBTS) is a primarily autosomal-recessive disorder characterized by a distinctive mid-hindbrain and cerebellar malformation, oculomotor apraxia, irregular breathing, developmental delay, and ataxia. JBTS is a genetically heterogeneous ciliopathy. We sought to characterize the genetic landscape associated with JBTS in the French Canadian (FC) population. We studied 43 FC JBTS subjects from 35 families by combining targeted and exome sequencing. We identified pathogenic (n = 32 families) or possibly pathogenic (n = 2 families) variants in genes previously associated with JBTS in all of these subjects, except for one. In the latter case, we found a homozygous splice-site mutation (c.735+2T>C) in CEP104. Interestingly, we identified two additional non-FC JBTS subjects with mutations in CEP104; one of these subjects harbors a maternally inherited nonsense mutation (c.496C>T [p.Arg166*]) and a de novo splice-site mutation (c.2572-2A>G), whereas the other bears a homozygous frameshift mutation (c.1328_1329insT [p.Tyr444fs*3]) in CEP104. Previous studies have shown that CEP104 moves from the mother centriole to the tip of the primary cilium during ciliogenesis. Knockdown of CEP104 in retinal pigment epithelial (RPE1) cells resulted in severe defects in ciliogenesis. These observations suggest that CEP104 acts early during cilia formation by regulating the conversion of the mother centriole into the cilia basal body. We conclude that disruption of CEP104 causes JBTS. Our study also reveals that the cause of JBTS has been elucidated in the great majority of our FC subjects (33/35 [94%] families), even though JBTS shows substantial locus and allelic heterogeneity in this population.

The Ciliopathy-Associated Cep104 Protein Interacts with Tubulin and Nek1 Kinase

“…Nek1 locates to the basal body region and (when overexpressed as a kinase dead version) cilia (Shalom et al., 2008, White and Quarmby, 2008), is found in proximity to Cep104 (Gupta et al., 2015) and was recently also detected in Cep104 affinity-capture experiments in a high-throughput interactome study (Hein et al., 2015). Intriguingly, mutations in both Cep104 and Nek1 cause ciliopathies in humans: in the case of Cep104 Joubert syndrome (Srour et al., 2015) and in the case of Nek1 short-rib thoracic dystrophy or oral-facial-digital syndrome type II (El Hokayem et al., 2012, Monroe et al., 2016, Thiel et al., 2011). Both proteins play a role in cilia formation and/or length regulation (Jiang et al., 2012, Satish Tammana et al., 2013, Shalom et al., 2008, Thiel et al., 2011, Wang et al., 2014).…”

“…Several lines of evidence suggest that the Cep104-Nek1 interaction is physiologically relevant: Nek1 has been found in the proximity of Cep104 (Gupta et al., 2015 and our study), and was also detected in Cep104 affinity-capture experiments in a high-throughput interactome study (Hein et al., 2015). Furthermore, Nek1 and Cep104 are involved in the stabilization of the ciliary axoneme or in cilia formation and elongation (Jiang et al., 2012, Satish Tammana et al., 2013, Shalom et al., 2008, Thiel et al., 2011, Wang et al., 2014), and mutations in both cause ciliopathies (El Hokayem et al., 2012, Monroe et al., 2016, Srour et al., 2015, Thiel et al., 2011). …”

“…Depletion of Cep104 leads either to the absence of cilia or the formation of cilia with a reduced length (Jiang et al., 2012, Satish Tammana et al., 2013). Furthermore, Cep104 mutations are linked to Joubert syndrome, a genetically heterogeneous ciliopathy (Srour et al., 2015). Taken together, these in vivo data suggest that Cep104 has an important role in regulating microtubule length in cilia.…”

Tectonic Proteins Are Important Players in Non-Motile Ciliopathies

“…were reported in a female child with Joubert syndrome [26]. In addition, another compound heterozygous mutation of TCTN1 c.342_2A>C (a spicing mutation) and c.898C>T (p.Arg300*) was reported in one male fetus showing typical molar tooth sign [66].…”

Neurological disorders, genetic correlations, and the role of exome sequencing

“…Two examples in which exome sequencing has proven successful are the new neuroectodermal CHIME (C oloboma, H eart defects, Ichthyosiform dermatosis, Mental retardation, Ear anomalies with hearing loss) syndrome that is "presumed to be an autosomal recessive disorder because of recurrence in sibs" [26], and the Joubert syndrome, a disorder in which, for the majority of subjects, "the causal mutations have not yet been identified…" [27]. The recent 2015 French Canadian population study that utilized a combined targeted and exome sequencing approach successfully identified two non-French Canadian subjects with the CEP104 mutations of Joubert syndrome and demonstrated a 94% (n=43) prevalence of CEP104 disruption as conclusive evidence of causation "despite substantial locus and allelic [population] heterogeneity" [28].…”