Cell type-specific regulation of ciliary transition zone assembly in vertebrates.

Ciliopathies are life-threatening human diseases caused by defective cilia. They can often be traced back to mutations of genes encoding transition zone (TZ) proteins demonstrating that the understanding of TZ organisation is of paramount importance. The TZ consists of multimeric protein modules that are subject to a stringent assembly hierarchy. Previous reports place Rpgrip1l at the top of the TZ assembly hierarchy in Caenorhabditis elegans By performing quantitative immunofluorescence studies in RPGRIP1L-/- mouse embryos and human embryonic cells, we recognise a different situation in vertebrates in which Rpgrip1l deficiency affects TZ assembly in a cell type-specific manner. In cell types in which the loss of Rpgrip1l alone does not affect all modules, additional truncation or removal of vertebrate-specific Rpgrip1 results in an impairment of all modules. Consequently, Rpgrip1l and Rpgrip1 synergistically ensure the TZ composition in several vertebrate cell types, revealing a higher complexity of TZ assembly in vertebrates than in invertebrates.

The ciliary protein RPGRIP1L governs autophagy independently of its proteasome-regulating function at the ciliary base in mouse embryonic fibroblasts.

Previously, macroautophagy/autophagy was demonstrated to be regulated inter alia by the primary cilium. Mutations in RPGRIP1L cause ciliary dysfunctions resulting in severe human diseases summarized as ciliopathies. Recently, we showed that RPGRIP1L deficiency leads to a decreased proteasomal activity at the ciliary base in mice. Importantly, the drug-induced restoration of proteasomal activity does not rescue ciliary length alterations in the absence of RPGRIP1L indicating that RPGRIP1L affects ciliary function also via other mechanisms. Based on this knowledge, we analyzed autophagy in Rpgrip1l-negative mouse embryos. In these embryos, autophagic activity was decreased due to an increased activation of the MTOR complex 1 (MTORC1). Application of the MTORC1 inhibitor rapamycin rescued dysregulated MTORC1, autophagic activity and cilia length but not proteasomal activity in Rpgrip1l-deficient mouse embryonic fibroblasts demonstrating that RPGRIP1L seems to regulate autophagic and proteasomal activity independently from each other.

The transition zone protein Rpgrip1l regulates proteasomal activity at the primary cilium.

Mutations in RPGRIP1L result in severe human diseases called ciliopathies. To unravel the molecular function of RPGRIP1L, we analyzed Rpgrip1l(-/-) mouse embryos, which display a ciliopathy phenotype and die, at the latest, around birth. In these embryos, cilia-mediated signaling was severely disturbed. Defects in Shh signaling suggested that the Rpgrip1l deficiency causes an impairment of protein degradation and protein processing. Indeed, we detected a cilia-dependent decreased proteasomal activity in the absence of Rpgrip1l. We found different proteasomal components localized to cilia and identified Psmd2, a component of the regulatory proteasomal 19S subunit, as an interaction partner for Rpgrip1l. Quantifications of proteasomal substrates demonstrated that Rpgrip1l regulates proteasomal activity specifically at the basal body. Our study suggests that Rpgrip1l controls ciliary signaling by regulating the activity of the ciliary proteasome via Psmd2.