The MAL protein is crucial for proper membrane condensation at the ciliary base, which is required for primary cilium elongation

Abstract

The base of the primary cilium contains a zone of condensed membranes whose importance is not known. Here, we have studied the involvement of MAL, a tetraspanning protein that exclusively partitions into condensed membrane fractions, in the condensation of membranes at the ciliary base and investigated the importance of these membranes in primary cilium formation. We show that MAL accumulates at the ciliary base of epithelial MDCK cells. Knockdown of MAL expression resulted in a drastic reduction in the condensation of membranes at the ciliary base, the percentage of ciliated cells and the length of the cilia, but did not affect the docking of the centrosome to the plasma membrane or produce missorting of proteins to the pericentriolar zone or to the membrane of the remaining cilia. Rab8 (for which there are two isoforms, Rab8A and Rab8b), IFT88 and IFT20, which are important components of the machinery of ciliary growth, were recruited normally to the ciliary base of MAL-knockdown cells but were unable to elongate the primary cilium correctly. MAL, therefore, is crucial for the proper condensation of membranes at the ciliary base, which is required for efficient primary cilium extension.

The ciliary membrane of polarized epithelial cells stems from a midbody remnant-associated membrane patch with condensed nanodomains

“…4A and Movie 2). This result is consistent with a previous report showing that proper membrane condensation at the centrosome zone is necessary for efficient primary cilium assembly (Reales, et al, 2015). It is of note that the CAMP decreased in size as the cilium elongated, as seen in 3D reconstructions ( Fig.…”

Novel role for the midbody in primary ciliogenesis by polarized epithelial cells

“…MDCK cells form a PC at the center of the apical membrane when they are grown at high cell density, but unlike other cells, they do not form a PC in response to serum deprivation. The PC of MDCK cells does not contain a ciliary pocket ( Zuo et al, 2009 ; Reales et al, 2015 ), and the centrosome of nonciliated cells localizes close to the apical membrane and is devoid of a large ciliary vesicle ( Fig. 1, A and B ).…”

“…This connection can facilitate transport of material or can be itself the means by which the midbody remnant enables the centrosome for PC formation. A second possibility for the role of the remnant in ciliogenesis is that, because the PC has compact membranes at the ciliary base ( Vieira et al, 2006 ) and these specialized membranes are required for PC formation ( Reales et al, 2015 ), the stalk zone constitutes a compact membrane domain used to form the ciliary base. A third possibility is that the presence of the remnant proximal to the centrosome promotes the signaling required for primary ciliogenesis.…”

Permeability barriers for generating a unique ciliary protein and lipid composition

“…This region of condensed lipid was identified using Laurdan [77], a probe whose fluorescence emission is sensitive to the degree of condensation (ordering) of its lipid bilayer environment. This ring of condensed lipid does not appear to regulate the passage of membrane proteins as knockdown of MAL, a membrane protein that partitions into condensed membranes, disrupted the formation of the ring of condensed lipid but did not cause the missorting of non-ciliary proteins to the ciliary compartment [78]. The region of condensed lipid may thus contribute to establishing and/or maintaining unique lipid environments for the ciliary and plasma membranes.…”