Gene batteries are sets of coregulated genes with common cis-regulatory elements that define the differentiated state of a cell. The nature of gene batteries for individual neuronal cellular subtypes and their linked cis-regulatory elements is poorly defined. Through molecular dissection of the highly modular cis-regulatory architecture of individual neuronally expressed genes, we have defined a conserved 16 bp cis-regulatory motif that drives gene expression in a single interneuron subtype, termed AIY, in the nematode Caenorhabditis elegans. This motif is bound and activated by the Paired- and LIM-type homeodomain proteins CEH-10 and TTX-3. Using genome-wide phylogenetic footprinting, we delineated the location, distribution, and evolution of AIY-specific cis-regulatory elements throughout the genome and thereby defined a large battery of AIY-expressed genes, all of which represent direct Paired/LIM homeodomain target genes. The identity of these homeodomain targets provides novel insights into the biology of the AIY interneuron.

Biogenic amines regulate a variety of behaviors. Their functions are predominantly mediated through G-protein-coupled 7-transmembrane domain receptors (GPCR), 16 of which are predicted to exist in the genome sequence of the nematode Caenorhabditis elegans. We describe here the expression pattern of several of these aminergic receptors, including two serotonin receptors (ser-1 and ser-4), one tyramine receptor (ser-2), and two dopamine receptors (dop-1 and dop-2). Moreover, we describe distinct but partially overlapping expression patterns of different splice forms of the ser-2 tyramine receptor locus. We find that each of the aminergic receptor genes is expressed in restricted regions of the nervous system and that many of them reveal significant overlap with the expression of regulatory factors of the LIM homeobox (Lhx) gene family. We demonstrate that the expression of several of the biogenic amine receptors is abrogated in specific cell types in Lhx gene mutants, thus establishing a role for these Lhx genes in regulating aspects of neurotransmission. We extend these findings with other cell fate markers and show that the lim-4 Lhx gene is required for several but not all aspects of RID motor neuron differentiation and that the lim-6 Lhx gene is required for specific aspects of RIS interneuron differentiation. We also use aminergic receptor gfp reporter fusions as tools to visualize the anatomy of specific neurons in Lhx mutant backgrounds and find that the development of the elaborate dendritic branching pattern of the PVD harsh touch sensory neuron requires the mec-3 Lhx gene. Lastly, we analyze a mutant allele of the ser-2 tyramine receptor, a target of the ttx-3 Lhx gene in the AIY interneuron class. ser-2 mutants display none of the defects previously shown to be associated with loss of AIY function.

IMPORTANCE-Age-related macular degeneration (AMD) is a multifactorial disease with genetic and environmental factors contributing to risk. Histopathologic changes underlying AMD are not fully understood, particularly the relationship between choriocapillaris (CC) dysfunction and phenotypic variability of this disease.OBJECTIVE-To examine histopathologic changes in the CC of eyes with clinically documented AMD. DESIGN, SETTING, AND PARTICIPANTS-The study was designed in 2011. Tissues were collected post mortem (2012)(2013)(2014)(2015)(2016), and histopathological images were obtained from participants enrolled in AMD studies since 1988. Clinical records and images were collected from participants as standard protocol. Eyes without AMD (n = 4) and eyes with early (n = 9), intermediate (n = 5), and advanced stages of AMD (geographic atrophy, n = 5; neovascular disease, n = 13) were evaluated. Choroidal vasculature was labeled using Ulex europaeus agglutinin lectin and examined using confocal microscopy.MAIN OUTCOMES AND MEASURES-A standardized classification system was applied to determine AMD stage. Ocular records and images were reviewed and histopathologic analyses performed. Viability of the choroidal vasculature was analyzed for each AMD stage.RESULTS-All participants were white. Fourteen were male, and 16 were female. The mean age was 90.5 years among AMD patients and 88.5 years among control participants. Submacular CC dropout without retinal pigment eipthelial (RPE) loss was observed in all cases with early stages of AMD. Higher vascular area loss for each AMD stage was observed compared with control participants: 20.5% in early AMD (95% CI, 11.2%-40.2%; P < .001), 12.5% in intermediate AMD (95% CI, 2.9%-21.4%; P = .01), 39.0% loss in GA (95% CI, 32.1%-45.4%; P < .001), and 38.2% loss in neovascular disease where RPE remained intact (95% CI, 27.7%-47.9%; P < .001).Hypercellular, apparent neovascular buds were adjacent to areas of CC loss in 22.2% of eyes with early AMD and 40% of eyes with intermediate AMD.CONCLUSIONS AND RELEVANCE-Retinal pigment epithelial atrophy preceded CC loss in geographic atrophy, but CC loss occurred in the absence of RPE atrophy in 2 of 9 eyes with earlystage AMD. Given the cross-sectional nature of this study and the small number of eyes evaluated, definitive conclusions regarding this progression cannot be determined with certainty. We speculate that neovascular buds may be a precursor to neovascular disease. Hypoxic RPE resulting from reduced blood supply might upregulate production of vascular endothelial growth factor, providing the stimulus for neovascular disease.Age-related macular degeneration (AMD) is a chronic and degenerative disease affecting the central part of the retina. It is the leading cause of irreversible visual loss in adults older than 60 years. 1,2 The etiology of AMD is multifactorial, and contributions of both genetic and environmental risk factors are well established [2][3][4] ; however, histopathologic changes Seddon et al.

SignificanceIn proliferative diabetic retinopathy (PDR), the most vision-threatening sequela of diabetic eye disease, retinal ischemia leads to increased expression of angiogenic factors that promote neovascularization. Although therapies targeting the potent angiogenic mediator vascular endothelial growth factor have been remarkably successful for the treatment of diabetic macular edema, this approach has not proven sufficient to prevent the development of retinal neovascularization, implicating additional angiogenic factor(s) in PDR pathogenesis. We demonstrate here that angiopoietin-like 4 is a potent angiogenic mediator with markedly increased expression in the eyes of PDR patients. Our studies identify a novel therapeutic target for the treatment of ocular neovascular disease and may have broad implications for the treatment of other diseases dependent on pathologic angiogenesis.