Bardet–Biedl syndrome proteins control the cilia length through regulation of actin polymerization

Abstract

Primary cilia are cellular appendages important for signal transduction and sensing the environment. Bardet–Biedl syndrome proteins form a complex that is important for several cytoskeleton-related processes such as ciliogenesis, cell migration and division. However, the mechanisms by which BBS proteins may regulate the cytoskeleton remain unclear. We discovered that Bbs4- and Bbs6-deficient renal medullary cells display a characteristic behaviour comprising poor migration, adhesion and division with an inability to form lamellipodial and filopodial extensions. Moreover, fewer mutant cells were ciliated [48% ± 6 for wild-type (WT) cells versus 23% ± 7 for Bbs4 null cells; P < 0.0001] and their cilia were shorter (2.55 μm ± 0.41 for WT cells versus 2.16 μm ± 0.23 for Bbs4 null cells; P < 0.0001). While the microtubular cytoskeleton and cortical actin were intact, actin stress fibre formation was severely disrupted, forming abnormal apical stress fibre aggregates. Furthermore, we observed over-abundant focal adhesions (FAs) in Bbs4-, Bbs6- and Bbs8-deficient cells. In view of these findings and the role of RhoA in regulation of actin filament polymerization, we showed that RhoA-GTP levels were highly upregulated in the absence of Bbs proteins. Upon treatment of Bbs4-deficient cells with chemical inhibitors of RhoA, we were able to restore the cilia length and number as well as the integrity of the actin cytoskeleton. Together these findings indicate that Bbs proteins play a central role in the regulation of the actin cytoskeleton and control the cilia length through alteration of RhoA levels.

STAR syndrome-associated CDK10/Cyclin M regulates actin network architecture and ciliogenesis

“…40 Second, and directly relevant to the regulation of actin dynamics, PKN2 interacts with RhoA and it makes a positive contribution to various RhoA-regulated processes, including stress fiber formation. 39 Moreover, RhoA inhibition is known to promote ciliogenesis, 22 a result we have confirmed in our assay system (Fig. S3C, D).…”

“…13,22,23 PKN2 0 s N-terminal domain is known to mediate its interaction with RhoA. 39 Thus, we hypothesized that CDK10/CycM might target this region, presumably in manner that would enhance RhoA and PKN2 0 s function in actin polymerization.…”

“…20,21 In contrast, in monociliated cells, activation of RhoA and its downstream signaling pathway has been shown to suppress primary ciliogenesis by promoting the formation of actin stress fiber networks. 13,22,23 The presence of abnormal ciliogenesis and/ or abnormal primary cilium length during development has been linked to severe developmental syndromes, collectively dubbed ciliopathies. 11,24 Notably, hyperactivation of RhoA and consequent changes in actin polymerization have been observed in several prototypical ciliopathies, including Bardet-Biedl, Meckel and Joubert syndromes.…”

“…11,24 Notably, hyperactivation of RhoA and consequent changes in actin polymerization have been observed in several prototypical ciliopathies, including Bardet-Biedl, Meckel and Joubert syndromes. 22,23,25 STAR syndrome is an X-linked dominant disorder caused by mutations in family with sequence similarity 58, member A (FAM58A). 26 We recently reported that Cyclin M (CycM), encoded by FAM58A, interacts with Cyclin-Dependent Kinase 10 (CDK10) to form an active protein kinase.…”

CDK10 Mutations in Humans and Mice Cause Severe Growth Retardation, Spine Malformations, and Developmental Delays

“…Immunostaining, using ARL13B and gamma-tubulin antibodies, revealed significantly longer cilia in Cdk10KO MEFs than in WT cells ( Figures 3F and 3G), corroborating results that were reported after the silencing of Cdk10 or cyclin M by siRNA 10 (for a review, see Guen et al 33 ). BBS4, which interacts with CEP290, 34 is a BBSome component that is essential for cilia formation, 35 providing a rationale for our observation of longer cilia in Cdk10KO MEFs.…”