APC7 mediates ubiquitin signaling in constitutive heterochromatin in the developing mammalian brain.

Neurodevelopmental cognitive disorders provide insights into mechanisms of human brain development. Here, we report an intellectual disability syndrome caused by the loss of APC7, a core component of the E3 ubiquitin ligase anaphase promoting complex (APC). In mechanistic studies, we uncover a critical role for APC7 during the recruitment and ubiquitination of APC substrates. In proteomics analyses of the brain from mice harboring the patient-specific APC7 mutation, we identify the chromatin-associated protein Ki-67 as an APC7-dependent substrate of the APC in neurons. Conditional knockout of the APC coactivator protein Cdh1, but not Cdc20, leads to the accumulation of Ki-67 protein in neurons in vivo, suggesting that APC7 is required for the function of Cdh1-APC in the brain. Deregulated neuronal Ki-67 upon APC7 loss localizes predominantly to constitutive heterochromatin. Our findings define an essential function for APC7 and Cdh1-APC in neuronal heterochromatin regulation, with implications for understanding human brain development and disease.

Homozygosity for a mutation affecting the catalytic domain of tyrosyl-tRNA synthetase (YARS) causes multisystem disease.

Aminoacyl-tRNA synthetases (ARSs) are critical for protein translation. Pathogenic variants of ARSs have been previously associated with peripheral neuropathy and multisystem disease in heterozygotes and homozygotes, respectively. We report seven related children homozygous for a novel mutation in tyrosyl-tRNA synthetase (YARS, c.499C > A, p.Pro167Thr) identified by whole exome sequencing. This variant lies within a highly conserved interface required for protein homodimerization, an essential step in YARS catalytic function. Affected children expressed a more severe phenotype than previously reported, including poor growth, developmental delay, brain dysmyelination, sensorineural hearing loss, nystagmus, progressive cholestatic liver disease, pancreatic insufficiency, hypoglycemia, anemia, intermittent proteinuria, recurrent bloodstream infections and chronic pulmonary disease. Related adults heterozygous for YARS p.Pro167Thr showed no evidence of peripheral neuropathy on electromyography, in contrast to previous reports for other YARS variants. Analysis of YARS p.Pro167Thr in yeast complementation assays revealed a loss-of-function, hypomorphic allele that significantly impaired growth. Recombinant YARS p.Pro167Thr demonstrated normal subcellular localization, but greatly diminished ability to homodimerize in human embryonic kidney cells. This work adds to a rapidly growing body of research emphasizing the importance of ARSs in multisystem disease and significantly expands the allelic and clinical heterogeneity of YARS-associated human disease. A deeper understanding of the role of YARS in human disease may inspire innovative therapies and improve care of affected patients.

Genomic diagnostics within a medically underserved population: efficacy and implications.

PurposeWe integrated whole-exome sequencing (WES) and chromosomal microarray analysis (CMA) into a clinical workflow to serve an endogamous, uninsured, agrarian community.MethodsSeventy-nine probands (newborn to 49.8 years) who presented between 1998 and 2015 remained undiagnosed after biochemical and molecular investigations. We generated WES data for probands and family members and vetted variants through rephenotyping, segregation analyses, and population studies.ResultsThe most common presentation was neurological disease (64%). Seven (9%) probands were diagnosed by CMA. Family WES data were informative for 37 (51%) of the 72 remaining individuals, yielding a specific genetic diagnosis (n = 32) or revealing a novel molecular etiology (n = 5). For five (7%) additional subjects, negative WES decreased the likelihood of genetic disease. Compared to trio analysis, "family" WES (average seven exomes per proband) reduced filtered candidate variants from 22 ± 6 to 5 ± 3 per proband. Nineteen (51%) alleles were de novo and 17 (46%) inherited; the latter added to a population-based diagnostic panel. We found actionable secondary variants in 21 (4.2%) of 502 subjects, all of whom opted to be informed.ConclusionCMA and family-based WES streamline and economize diagnosis of rare genetic disorders, accelerate novel gene discovery, and create new opportunities for community-based screening and prevention in underserved populations.

An Amish founder mutation disrupts a PI(3)P-WHAMM-Arp2/3 complex-driven autophagosomal remodeling pathway.

Actin nucleation factors function to organize, shape, and move membrane-bound organelles, yet they remain poorly defined in relation to disease. Galloway-Mowat syndrome (GMS) is an inherited disorder characterized by microcephaly and nephrosis resulting from mutations in the WDR73 gene. This core clinical phenotype appears frequently in the Amish, where virtually all affected individuals harbor homozygous founder mutations in WDR73 as well as the closely linked WHAMM gene, which encodes a nucleation factor. Here we show that patient cells with both mutations exhibit cytoskeletal irregularities and severe defects in autophagy. Reintroduction of wild-type WHAMM restored autophagosomal biogenesis to patient cells, while inactivation of WHAMM in healthy cell lines inhibited lipidation of the autophagosomal protein LC3 and clearance of ubiquitinated protein aggregates. Normal WHAMM function involved binding to the phospholipid PI(3)P and promoting actin nucleation at nascent autophagosomes. These results reveal a cytoskeletal pathway controlling autophagosomal remodeling and illustrate several molecular processes that are perturbed in Amish GMS patients.

Mutations in CRADD Result in Reduced Caspase-2-Mediated Neuronal Apoptosis and Cause Megalencephaly with a Rare Lissencephaly Variant.

Lissencephaly is a malformation of cortical development typically caused by deficient neuronal migration resulting in cortical thickening and reduced gyration. Here we describe a "thin" lissencephaly (TLIS) variant characterized by megalencephaly, frontal predominant pachygyria, intellectual disability, and seizures. Trio-based whole-exome sequencing and targeted re-sequencing identified recessive mutations of CRADD in six individuals with TLIS from four unrelated families of diverse ethnic backgrounds. CRADD (also known as RAIDD) is a death-domain-containing adaptor protein that oligomerizes with PIDD and caspase-2 to initiate apoptosis. TLIS variants cluster in the CRADD death domain, a platform for interaction with other death-domain-containing proteins including PIDD. Although caspase-2 is expressed in the developing mammalian brain, little is known about its role in cortical development. CRADD/caspase-2 signaling is implicated in neurotrophic factor withdrawal- and amyloid-ß-induced dendritic spine collapse and neuronal apoptosis, suggesting a role in cortical sculpting and plasticity. TLIS-associated CRADD variants do not disrupt interactions with caspase-2 or PIDD in co-immunoprecipitation assays, but still abolish CRADD's ability to activate caspase-2, resulting in reduced neuronal apoptosis in vitro. Homozygous Cradd knockout mice display megalencephaly and seizures without obvious defects in cortical lamination, supporting a role for CRADD/caspase-2 signaling in mammalian brain development. Megalencephaly and lissencephaly associated with defective programmed cell death from loss of CRADD function in humans implicate reduced apoptosis as an important pathophysiological mechanism of cortical malformation. Our data suggest that CRADD/caspase-2 signaling is critical for normal gyration of the developing human neocortex and for normal cognitive ability.

Recessive nephrocerebellar syndrome on the Galloway-Mowat syndrome spectrum is caused by homozygous protein-truncating mutations of WDR73.

We describe a novel nephrocerebellar syndrome on the Galloway-Mowat syndrome spectrum among 30 children (ages 1.0 to 28 years) from diverse Amish demes. Children with nephrocerebellar syndrome had progressive microcephaly, visual impairment, stagnant psychomotor development, abnormal extrapyramidal movements and nephrosis. Fourteen died between ages 2.7 and 28 years, typically from renal failure. Post-mortem studies revealed (i) micrencephaly without polymicrogyria or heterotopia; (ii) atrophic cerebellar hemispheres with stunted folia, profound granule cell depletion, Bergmann gliosis, and signs of Purkinje cell deafferentation; (iii) selective striatal cholinergic interneuron loss; and (iv) optic atrophy with delamination of the lateral geniculate nuclei. Renal tissue showed focal and segmental glomerulosclerosis and extensive effacement and microvillus transformation of podocyte foot processes. Nephrocerebellar syndrome mapped to 700 kb on chromosome 15, which contained a single novel homozygous frameshift variant (WDR73 c.888delT; p.Phe296Leufs*26). WDR73 protein is expressed in human cerebral cortex, hippocampus, and cultured embryonic kidney cells. It is concentrated at mitotic microtubules and interacts with α-, ß-, and γ-tubulin, heat shock proteins 70 and 90 (HSP-70; HSP-90), and the carbamoyl phosphate synthetase 2/aspartate transcarbamylase/dihydroorotase multi-enzyme complex. Recombinant WDR73 p.Phe296Leufs*26 and p.Arg256Profs*18 proteins are truncated, unstable, and show increased interaction with α- and ß-tubulin and HSP-70/HSP-90. Fibroblasts from patients homozygous for WDR73 p.Phe296Leufs*26 proliferate poorly in primary culture and senesce early. Our data suggest that in humans, WDR73 interacts with mitotic microtubules to regulate cell cycle progression, proliferation and survival in brain and kidney. We extend the Galloway-Mowat syndrome spectrum with the first description of diencephalic and striatal neuropathology.

CODAS syndrome is associated with mutations of LONP1, encoding mitochondrial AAA+ Lon protease.

CODAS syndrome is a multi-system developmental disorder characterized by cerebral, ocular, dental, auricular, and skeletal anomalies. Using whole-exome and Sanger sequencing, we identified four LONP1 mutations inherited as homozygous or compound-heterozygous combinations among ten individuals with CODAS syndrome. The individuals come from three different ancestral backgrounds (Amish-Swiss from United States, n = 8; Mennonite-German from Canada, n = 1; mixed European from Canada, n = 1). LONP1 encodes Lon protease, a homohexameric enzyme that mediates protein quality control, respiratory-complex assembly, gene expression, and stress responses in mitochondria. All four pathogenic amino acid substitutions cluster within the AAA(+) domain at residues near the ATP-binding pocket. In biochemical assays, pathogenic Lon proteins show substrate-specific defects in ATP-dependent proteolysis. When expressed recombinantly in cells, all altered Lon proteins localize to mitochondria. The Old Order Amish Lon variant (LONP1 c.2161C>G[p.Arg721Gly]) homo-oligomerizes poorly in vitro. Lymphoblastoid cell lines generated from affected children have (1) swollen mitochondria with electron-dense inclusions and abnormal inner-membrane morphology; (2) aggregated MT-CO2, the mtDNA-encoded subunit II of cytochrome c oxidase; and (3) reduced spare respiratory capacity, leading to impaired mitochondrial proteostasis and function. CODAS syndrome is a distinct, autosomal-recessive, developmental disorder associated with dysfunction of the mitochondrial Lon protease.

A population-based study of KCNH7 p.Arg394His and bipolar spectrum disorder.

We conducted blinded psychiatric assessments of 26 Amish subjects (52 ± 11 years) from four families with prevalent bipolar spectrum disorder, identified 10 potentially pathogenic alleles by exome sequencing, tested association of these alleles with clinical diagnoses in the larger Amish Study of Major Affective Disorder (ASMAD) cohort, and studied mutant potassium channels in neurons. Fourteen of 26 Amish had bipolar spectrum disorder. The only candidate allele shared among them was rs78247304, a non-synonymous variant of KCNH7 (c.1181G>A, p.Arg394His). KCNH7 c.1181G>A and nine other potentially pathogenic variants were subsequently tested within the ASMAD cohort, which consisted of 340 subjects grouped into controls subjects and affected subjects from overlapping clinical categories (bipolar 1 disorder, bipolar spectrum disorder and any major affective disorder). KCNH7 c.1181G>A had the highest enrichment among individuals with bipolar spectrum disorder (χ(2) = 7.3) and the strongest family-based association with bipolar 1 (P = 0.021), bipolar spectrum (P = 0.031) and any major affective disorder (P = 0.016). In vitro, the p.Arg394His substitution allowed normal expression, trafficking, assembly and localization of HERG3/Kv11.3 channels, but altered the steady-state voltage dependence and kinetics of activation in neuronal cells. Although our genome-wide statistical results do not alone prove association, cumulative evidence from multiple independent sources (parallel genome-wide study cohorts, pharmacological studies of HERG-type potassium channels, electrophysiological data) implicates neuronal HERG3/Kv11.3 potassium channels in the pathophysiology of bipolar spectrum disorder. Such a finding, if corroborated by future studies, has implications for mental health services among the Amish, as well as development of drugs that specifically target HERG3/Kv11.3.

A homozygous SLITRK6 nonsense mutation is associated with progressive auditory neuropathy in humans.

OBJECTIVES/HYPOTHESIS: SLITRK family proteins control neurite outgrowth and regulate synaptic development. In mice, Slitrk6 plays a role in the survival and innervation of sensory neurons in the inner ear, vestibular apparatus, and retina, and also influences axial eye length. We provide the first detailed description of the auditory phenotype in humans with recessive SLITRK6 deficiency. STUDY DESIGN: Prospective observational case study. METHODS: Nine closely related Amish subjects from an endogamous Amish community of Pennsylvania underwent audiologic and vestibular testing. Single nucleotide polymorphism microarrays were used to map the chromosome locus, and Sanger sequencing or high-resolution melt analysis were used to confirm the allelic variant. RESULTS: All nine subjects were homozygous for a novel nonsense variant of SLITRK6 (c.1240C>T, p.Gln414Ter). Adult patients had high myopia. The 4 oldest SLITRK6 c.1240C>T homozygotes had absent ipsilateral middle ear muscle reflexes (MEMRs). Distortion product otoacoustic emissions (DPOAEs) were absent in all ears tested and the cochlear microphonic (CM) was increased in amplitude and duration in young patients and absent in the two oldest subjects. Auditory brainstem responses (ABRs) were dys-synchronised bilaterally with no reproducible waves I, III, or V at high intensities. Hearing loss and speech reception thresholds deteriorated symmetrically with age, which resulted in severe-to-profound hearing impairment by early adulthood. Vestibular evoked myogenic potentials were normal in three ears and absent in one. CONCLUSION: Homozygous SLITRK6 c.1240C>T (p.Gln414Ter) nonsense mutations are associated with high myopia, cochlear dysfunction attributed to outer hair cell disease, and progressive auditory neuropathy.

A homozygous missense mutation in HERC2 associated with global developmental delay and autism spectrum disorder.

We studied a unique phenotype of cognitive delay, autistic behavior, and gait instability segregating in three separate sibships. We initiated genome-wide mapping in two sibships using Affymetrix 10K SNP Mapping Arrays and identified a homozygous 8.2 Mb region on chromosome 15 common to five affected children. We used exome sequencing of two affected children to assess coding sequence variants within the mapped interval. Four novel homozygous exome variants were shared between the two patients; however, only two variants localized to the mapped interval on chromosome 15. A third sibship in an Ohio Amish deme narrowed the mapped interval to 2.6 Mb and excluded one of the two novel homozygous exome variants. The remaining variant, a missense change in HERC2 (c.1781C>T, p.Pro594Leu), occurs in a highly conserved proline residue within an RCC1-like functional domain. Functional studies of truncated HERC2 in adherent retinal pigment epithelium cells suggest that the p.Pro594Leu variant induces protein aggregation and leads to decreased HERC2 abundance. The phenotypic correlation with the mouse Herc1 and Herc2 mutants as well as the phenotypic overlap with Angelman syndrome provide further evidence that pathogenic changes in HERC2 are associated with nonsyndromic intellectual disability, autism, and gait disturbance. Hum Mutat 33:1639-1646, 2012. © 2012 Wiley Periodicals, Inc.

Genetic mapping and exome sequencing identify variants associated with five novel diseases.

The Clinic for Special Children (CSC) has integrated biochemical and molecular methods into a rural pediatric practice serving Old Order Amish and Mennonite (Plain) children. Among the Plain people, we have used single nucleotide polymorphism (SNP) microarrays to genetically map recessive disorders to large autozygous haplotype blocks (mean = 4.4 Mb) that contain many genes (mean = 79). For some, uninformative mapping or large gene lists preclude disease-gene identification by Sanger sequencing. Seven such conditions were selected for exome sequencing at the Broad Institute; all had been previously mapped at the CSC using low density SNP microarrays coupled with autozygosity and linkage analyses. Using between 1 and 5 patient samples per disorder, we identified sequence variants in the known disease-causing genes SLC6A3 and FLVCR1, and present evidence to strongly support the pathogenicity of variants identified in TUBGCP6, BRAT1, SNIP1, CRADD, and HARS. Our results reveal the power of coupling new genotyping technologies to population-specific genetic knowledge and robust clinical data.

Central regulation of photosensitive membrane turnover in the lateral eye of Limulus, II: octopamine acts via adenylate cyclase/cAMP-dependent protein kinase to prime the retina for transient rhabdom shedding.

Why photoreceptors turn over a portion of their photoreceptive membrane daily is not clear; however, failure to do so properly leads to retinal degeneration in vertebrates and invertebrates. Little is known about the molecular mechanisms that regulate shedding and renewal of photoreceptive membrane. Photoreceptive cells in the lateral eye of the horseshoe crab Limulus turn over their photoreceptive membrane (rhabdom) in brief, synchronous burst in response to dawn each morning. Transient rhabdom shedding (TRS), the first phase of rhabdom turnover in Limulus, is triggered by dawn, but requires a minimum of 3-5 h of overnight priming from the central circadian clock (Chamberlain & Barlow, 1984). We determined previously that the clock primes the lateral eye for TRS using the neurotransmitter octopamine (OA) (Khadilkar et al., 2002), and report here that OA primes the eye for TRS through a G(s)-coupled, adenylate cyclase (AC)/cyclic adenosine 3',5'-monophosphate (cAMP)/cAMP-dependent protein kinase (PKA) signaling cascade. Long-term intraretinol injections (6-7 h @ 1.4 microl/min) of the AC activator forskolin, or the cAMP analogs Sp-cAMP[s] and 8-Br-cAmp primed the retina for TRS in eyes disconnected from the circadian clock, and/or in intact eyes during the day when the clock is quiescent. This suggests that OA primes the eye for TRS by stimulating an AC-mediated rise in intracellular cAMP concentration ([cAMP]i). Co-injection of SQ 22,536, an AC inhibitor, or the PKA inhibitors H-89 and PKI (14-22) with OA effectively antagonized octopaminergic priming by reducing the number of photoreceptors primed for TRS and the amount of rhabdom shed by those photoreceptors compared with eyes treated with OA alone. Our data suggest that OA primes the lateral eye for TRS in part through long-term phosphorylation of a PKA substrate.

Desensitization of the photoresponse in Limulus ventral photoreceptors by protein kinase C precedes rhabdomere disorganization and endocytosis.

Limulus photoreceptors utilize the phosphoinositide pathway to generate light-induced single photon events (quantum bumps) that sum to form the depolarizing receptor potential. The protein kinase C (PKC) activator, (-)-indolactam V (ILV) rapidly desensitizes the light response in Limulus ventral nerve photoreceptors. Within 10 min of extracellular application, 100 nM (-)-ILV caused a decrease in the mean amplitude of quantum bumps to 38% of control values. PKC activation by (-)-ILV also causes photosensitive membrane disorganization and endocytosis. To investigate whether this precedes desensitization of the electrical response, we fixed cells after 10-min incubation with 25 microM (-)-ILV, a concentration sufficient to cause a 1000-fold desensitization of the receptor potential. The photosensitive microvilli of these photoreceptors remained narrow, densely packed, and well organized. Increasing the incubation time to 60 min did, however, induce disorganization and swelling of the microvilli and endocytosis of the photosensitive membrane, as previously reported. Measurement of membrane capacitance did not indicate a significant reduction in membrane area accompanying desensitization by (-)-ILV. PKC-induced reduction in light sensitivity therefore precedes the detection of ultrastructural changes in the rhabdomeral membrane and is not due to a net loss of membrane.

Limulus opsins: diurnal regulation of expression.

Much has been learned from studies of Limulus photoreceptors about the role of the circadian clock and light in the removal of photosensitive membrane. However, little is known in this animal about mechanisms regulating photosensitive membrane renewal, including the synthesis of proteins in, and associated with, the photosensitive membrane. To begin to understand renewal, this study examines diurnal changes in the levels of mRNAs encoding opsin, the integral membrane protein component of visual pigment, and the relative roles of light and the circadian clock in producing these changes. We show that at least two distinct opsin genes encoding very similar proteins are expressed in both the lateral and ventral eyes, and that during the day and night in the lateral eye, the average level of mRNA encoding opsinl is consistently higher than that encoding opsin2. Northern blot assays showed further that total opsin mRNA in the lateral eyes of animals maintained under natural illumination increases during the afternoon (9 & 12 h after sunrise) in the light and falls at night in the dark. This diurnal change occurs whether or not the eyes receive input from the circadian clock, but it is eliminated in eyes maintained in the dark. Thus, it is regulated by light and darkness, not by the circadian clock, with light stimulating an increase in opsin mRNA levels. The rise in opsin mRNA levels observed under natural illumination was seasonal; it occurred during the summer but not the spring and fall. However, a significant increase in opsin mRNA levels could be achieved in the fall by exposing lateral eyes to 3 h of natural illumination followed by 9 h of artificial light. The diurnal regulation of opsin mRNA levels contrasts sharply with the circadian regulation of visual arrestin mRNA levels (Battelle et al., 2000). Thus, in Limulus, distinctly different mechanisms regulate the levels of mRNA encoding two proteins critical for the photoresponse.

Adaptive visual metamorphosis in a deep-sea hydrothermal vent crab.

Hydrothermal vents along the mid-ocean ridges host ephemeral ecosystems of diverse endemic fauna including several crustacean species, some of which undergo planktonic development as larvae up to 1,000 m above and 100 km away from the vents. Little is known about the role of vision in the life history of vent fauna. Here we report that planktonic zoea larvae of the vent crab Bythograea thermydron possess image-forming compound eyes with a visual pigment sensitive to the blue light of mesopelagic waters. As they metamorphose and begin to descend to and settle at the vents, they lose their image-forming optics and develop high-sensitivity naked-retina eyes. The spectral absorbance of the visual pigment in these eyes shifts towards longer wavelengths from larva to postlarva to adult. This progressive visual metamorphosis trades imaging for increased sensitivity, and changes spectral sensitivity from the blue wavelengths of the larval environment towards the dim, longer wavelengths produced in the deeper bathypelagic vent environment of the adults. As hydrothermal vents produce light, vision may supplement thermal and chemical senses to orient postlarval settlement at vent sites.

Central regulation of photosensitive membrane turnover in the lateral eye of Limulus. I. Octopamine primes the retina for daily transient rhabdom shedding.

Limulus lateral eyes shed and renew a portion of their photosensitive membrane (rhabdom) daily. Shedding, in many species including Limulus, is regulated by complex interactions between circadian rhythms and light. Little is known about how circadian clocks and photoreceptors communicate to regulate shedding. Limulus photoreceptors do not contain an endogenous circadian oscillator, but rely upon efferent outflow from a central clock for circadian timing. To investigate whether the putative efferent neurotransmitter octopamine (OA) communicates circadian rhythms that prime the lateral eye for transient rhabdom shedding, we decoupled photoreceptors from the clock by transecting the lateral optic nerve (contains the retinal efferent fibers). Overnight (6 h) intraretinal injections of 40 microM OA restored transient shedding to lateral eyes with transected nerves to levels comparable to those of intact internal control eyes. To determine whether OA acts alone in communicating circadian rhythms that prime the lateral eye for transient shedding, we "primed" eyes with intact nerves for transient shedding with exogenous OA during subjective day. In nature, lateral eyes shed their rhabdoms only once a day at dawn following overnight efferent priming. Eyes in animals placed in darkness during subjective day, when the retinal efferents are quiescent, and injected for 6 h with 40 microM OA shed their rhabdoms in response to a second introduction to light. Untreated control eyes of the same animals did not. The same results were observed in vitro in lateral eyes treated similarly. Octopamine is the only efferent neurotransmitter/messenger required to make lateral eyes competent for transient shedding. Phentolamine, an OA receptor antagonist, reduced the number of photoreceptors primed for transient shedding and the amount of rhabdom shed in those photoreceptors suggesting that OA acts via a specific OA receptor.

Protein kinase C-induced disorganization and endocytosis of photosensitive membrane in Limulus ventral photoreceptors.

Protein kinase C (PKC) desensitizes the light response in photoreceptors from the ventral optic nerve of the horseshoe crab Limulus. Photoisomerization of Limulus rhodopsin leads to phosphoinositide hydrolysis, resulting in the production of inositol trisphosphate and diacylglycerol (DAG). Inositol trisphosphate mobilizes intracellular stores of Ca(2+), resulting in photoreceptor excitation in Limulus, while DAG may activate PKC. We investigated whether PKC-mediated desensitization of the photoresponse is accompanied by ultrastructural changes in the rhodopsin-bearing photosensitive membrane (rhabdom) in Limulus ventral photoreceptors. PKC activation by (-)-indolactam V in darkness induces disorganization and swelling of the rhodopsin-containing microvilli and endocytosis of rhabdomeral membrane. The effects of (-)-indolactam V on dark-adapted photoreceptor ultrastructure are reversible, are stereospecific, are blocked by coapplication of PKC inhibitors, and closely match those induced by continuous, bright light. Rhabdom disorganization and endocytosis via PKC activation may, therefore, contribute to desensitization of the light-adapted photoreceptor.