The striated ciliary rootlet is a prominent cytoskeleton originating from basal bodies of ciliated cells. Although a familiar structure in cell biology, its function has remained unresolved. In this study, we carried out targeted disruption in mice of the gene for rootletin, a component of the rootlet. In the mutant, ciliated cells are devoid of rootlets. Phototransduction and ciliary beating in sensory and motile cilia initially exhibit no apparent functional deficits. However, photoreceptors degenerate over time, and mutant lungs appear prone to pathological changes consistent with insufficient mucociliary clearance. Further analyses revealed a striking fragility at the ciliary base in photoreceptors lacking rootlets. In vitro assays suggest that the rootlet is among the least dynamic of all cytoskeletons and interacts with actin filaments. Thus, a primary function of the rootlet is to provide structural support for the cilium. Inasmuch as photoreceptors elaborate an exceptionally enlarged sensory cilium, they are especially dependent on the rootlet for structural integrity and long-term survival.The ciliary rootlet originates from the proximal ends of basal bodies, the centriole-related structure that anchors the cilia, and extends proximally toward the cell nuclei (5, 25). Prominent rootlets are associated with sensory cilia of photoreceptor cells and motile cilia, such as those lining the respiratory tract, oviduct, and brain ventricles. The retinal photoreceptor elaborates a single enlarged distal cilium known as the outer segment. The outer segment is packed with photosensitive membranous disks and specializes in phototransduction. It is among the largest of all mammalian cilia, spanning approximately 25 m in length and more than 1 m in diameter. The outer segment is linked to the cell body, the inner segment, through a thin bridge called the connecting cilium (3). In a photoreceptor, the rootlet appears as a very thick striated filament that traverses the entire cell body all the way to the synaptic terminal (17,19,26). In epithelial cells bearing motile cilia, rootlets appear as robust subapical filamentous networks.

Aryl hydrocarbon receptor-interacting protein-like 1 (AIPL1) is a member of the FK-506-binding protein family expressed specifically in retinal photoreceptors. Mutations in AIPL1 cause Leber congenital amaurosis, a severe early-onset retinopathy that leads to visual impairment in infants. Here we show that knockdown of AIPL1 expression in mice also produces a retinopathy but over a more extended time course. Before any noticeable pathology, there was a reduction in the level of rod cGMP phosphodiesterase (PDE) proportional to the decrease in AIPL1 expression, whereas other photoreceptor proteins were unaffected. Consistent with less PDE in rods, flash responses had a delayed onset, a reduced gain, and a slower recovery of flash responses. We suggest that AIPL1 is a specialized chaperone required for rod PDE biosynthesis. Thus loss of AIPL1 would result in a condition that phenocopies retinal degenerations in the rd mouse and in a subgroup of human patients.

Membrane palmitoylated protein 4 (Mpp4) is a member of the membrane-associated guanylate kinase family. We show that Mpp4 localizes specifically to the plasma membrane of photoreceptor synaptic terminals. Plasma membrane Ca(2+) ATPases (PMCAs), the Ca(2+) extrusion pumps, interact with an Mpp4-dependent presynaptic membrane protein complex that includes Veli3 and PSD95. In mice lacking Mpp4, PMCAs were lost from rod photoreceptor presynaptic membranes. Synaptic ribbons were enlarged, a phenomenon known to correlate with higher Ca(2+). SERCA2 (sarcoplasmic-endoplasmic reticulum Ca(2+) ATPase, type 2), which pumps cytosolic Ca(2+) into intracellular Ca(2+) stores and localizes next to the ribbons, was increased. The distribution of IP(3)RII (InsP(3) receptor, type 2), which releases Ca(2+) from the stores, was shifted away from the synaptic terminals. Synaptic transmission to second-order neurons was maintained but was reduced in amplitude. These data suggest that loss of Mpp4 disrupts a Ca(2+) extrusion mechanism at the presynaptic membranes, with ensuing adaptive responses by the photoreceptor to restore Ca(2+) homeostasis. We propose that Mpp4 organizes a presynaptic protein complex that includes PMCAs and has a role in modulating Ca(2+) homeostasis and synaptic transmission in rod photoreceptors.