Photoreceptors are polarized neurons, with very specific subcellular compartmentalization and unique requirements for protein expression and trafficking. Each photoreceptor contains an outer segment, the site of photon capture that initiates vision, an inner segment that houses the biosynthetic machinery and a synaptic terminal for signal transmission to downstream neurons. Outer segments and inner segments are connected by a connecting cilium (CC), the equivalent of a transition zone (TZ) of primary cilia. The connecting cilium is part of the basal body/axoneme backbone that stabilizes the outer segment. This report will update the reader on late developments in photoreceptor ciliogenesis and transition zone formation, specifically in mouse photoreceptors, focusing on early events in photoreceptor ciliogenesis. The connecting cilium, an elongated and narrow structure through which all outer segment proteins and membrane components must traffic, functions as a gate that controls access to the outer segment. Here we will review genes and their protein products essential for basal body maturation and for CC/TZ genesis, sorted by phenotype. Emphasis is given to naturally occurring mouse mutants and gene knockouts that interfere with CC/TZ formation and ciliogenesis.

Modulation of connectivity formation in the developing brain in response to external stimuli is poorly understood. Here, we show that the raphe nucleus and its serotonergic projections regulate pathfinding of commissural axons in zebrafish. We found that the raphe neurons extend projections toward midline-crossing axons and that when serotonergic signaling is blocked by pharmacological inhibition or by raphe neuron ablation, commissural pathfinding is disrupted. We demonstrate that the serotonin receptor htr2a is expressed on these commissural axons and that genetic knock-down of htr2a disrupts crossing. We further show that knock-down of htr2a or ablation of the raphe neurons increases ephrinB2a protein levels in commissural axons. An ephrinB2a mutant can rescue midline crossing when serotonergic signaling is blocked. Furthermore, we found that regulation of serotonin expression in the raphe neurons is modulated in response to the developmental environment. Hypoxia causes the raphe to decrease serotonin levels, leading to a reduction in midline crossing. Increasing serotonin in the setting of hypoxia restored midline crossing. Our findings demonstrate an instructive role for serotonin in axon guidance acting through ephrinB2a and reveal a novel mechanism for developmental interpretation of the environmental milieu in the generation of mature neural circuitry.

Cytoplasmic dynein (dynein 1), a major retrograde motor of eukaryotic cells, is a 1.4 MDa protein complex consisting of a pair of heavy chains (DYNC1H1) and a set of heterodimeric noncatalytic accessory components termed intermediate, light intermediate and light chains. DYNC1H1 (4644 amino acids) is the dynein backbone encoded by a gene consisting of 77 exons. We generated a floxed Dync1h1 allele that excises exons 24 and 25 and truncates DYNC1H1 during Six3Cre-induced homologous recombination. Truncation results in loss of the motor and microtubule-binding domain. Dync1h1F/F;Six3Cre photoreceptors degenerated rapidly within two postnatal weeks. In the postnatal day 6 (P6) Dync1h1F/F;Six3Cre central retina, outer and inner nuclear layers were severely disorganized and lacked a recognizable outer plexiform layer (OPL). Although the gene was effectively silenced by P6, DYNC1H1 remnants persisted and aggregated together with rhodopsin, PDE6 and centrin-2-positive centrosomes in the outer nuclear layer. As photoreceptor degeneration is delayed in the Dync1h1F/F;Six3Cre retina periphery, retinal lamination and outer segment elongation are in part preserved. DYNC1H1 strongly persisted in the inner plexiform layer (IPL) beyond P16 suggesting lack of clearance of the DYNC1H1 polypeptide. This persistence of DYNC1H1 allows horizontal, rod bipolar, amacrine and ganglion cells to survive past P12. The results show that cytoplasmic dynein is essential for retina lamination, nuclear positioning, vesicular trafficking of photoreceptor membrane proteins and inner/outer segment elaboration.

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