Multiple domains in the Crumbs Homolog 2a (Crb2a) protein are required for regulating rod photoreceptor size

Background Vertebrate retinal photoreceptors are morphologically complex cells that have two apical regions, the inner segment and the outer segment. The outer segment is a modified cilium and is continuously regenerated throughout life. The molecular and cellular mechanisms that underlie vertebrate photoreceptor morphogenesis and the maintenance of the outer segment are largely unknown. The Crumbs (Crb) complex is a key regulator of apical membrane identity and size in epithelia and in Drosophila photoreceptors. Mutations in the human gene CRUMBS HOMOLOG 1 (CRB1) are associated with early and severe vision loss. Drosophila Crumbs and vertebrate Crb1 and Crumbs homolog 2 (Crb2) proteins are structurally similar, all are single pass transmembrane proteins with a large extracellular domain containing multiple laminin- and EGF-like repeats and a small intracellular domain containing a FERM-binding domain and a PDZ-binding domain. In order to begin to understand the role of the Crb family of proteins in vertebrate photoreceptors we generated stable transgenic zebrafish in which rod photoreceptors overexpress full-length Crb2a protein and several other Crb2a constructs engineered to lack specific domains. Results We examined the localization of Crb2a constructs and their effects on rod morphology. We found that only the full-length Crb2a protein approximated the normal localization of Crb2a protein apical to adherens junctions in the photoreceptor inner segment. Several Crb2a construct proteins localized abnormally to the outer segment and one construct localized abnormally to the cell body. Overexpression of full-length Crb2a greatly increased inner segment size while expression of several other constructs increased outer segment size. Conclusions Our observations suggest that particular domains in Crb2a regulate its localization and thus may regulate its regionalized function. Our results also suggest that the PDZ-binding domain in Crb2a might bring a protein(s) into the Crb complex that alters the function of the FERM-binding domain

Genetic Interaction of Centrosomin and Bazooka in Apical Domain Regulation in Drosophila Photoreceptor

Cell polarity genes including Crumbs (Crb) and Par complexes are essential for controlling photoreceptor morphogenesis. Among the Crb and Par complexes, Bazooka (Baz, Par-3 homolog) acts as a nodal component for other cell polarity proteins. Therefore, finding other genes interacting with Baz will help us to understand the cell polarity genes' role in photoreceptor morphogenesis. mutation on developing eyes to determine its role in photoreceptor morphogenesis. We found that Cnn is dispensable for retinal differentiation in eye imaginal discs during the larval stage. However, photoreceptors deficient in Cnn display dramatic morphogenesis defects including the mislocalization of Crumbs (Crb) and Bazooka (Baz) during mid-stage pupal eye development, suggesting that Cnn is specifically required for photoreceptor morphogenesis during pupal eye development. This role of Cnn in apical domain modulation was further supported by Cnn's gain-of-function phenotype. Cnn overexpression in photoreceptors caused the expansion of the apical Crb membrane domain, Baz and adherens junctions (AJs). photoreceptor

Activation of JNK Signaling Mediates Amyloid-ß-Dependent Cell Death

Alzheimer's disease (AD) is an age related progressive neurodegenerative disorder. One of the reasons for Alzheimer's neuropathology is the generation of large aggregates of Aß42 that are toxic in nature and induce oxidative stress, aberrant signaling and many other cellular alterations that trigger neuronal cell death. However, the exact mechanisms leading to cell death are not clearly understood.We employed a Drosophila eye model of AD to study how Aß42 causes cell death. Misexpression of higher levels of Aß42 in the differentiating photoreceptors of fly retina rapidly induced aberrant cellular phenotypes and cell death. We found that blocking caspase-dependent cell death initially blocked cell death but did not lead to a significant rescue in the adult eye. However, blocking the levels of c-Jun NH(2)-terminal kinase (JNK) signaling pathway significantly rescued the neurodegeneration phenotype of Aß42 misexpression both in eye imaginal disc as well as the adult eye. Misexpression of Aß42 induced transcriptional upregulation of puckered (puc), a downstream target and functional read out of JNK signaling. Moreover, a three-fold increase in phospho-Jun (activated Jun) protein levels was seen in Aß42 retina as compared to the wild-type retina. When we blocked both caspases and JNK signaling simultaneously in the fly retina, the rescue of the neurodegenerative phenotype is comparable to that caused by blocking JNK signaling pathway alone.Our data suggests that (i) accumulation of Aß42 plaques induces JNK signaling in neurons and (ii) induction of JNK contributes to Aß42 mediated cell death. Therefore, inappropriate JNK activation may indeed be relevant to the AD neuropathology, thus making JNK a key target for AD therapies

Role of Kinesin Heavy Chain in Crumbs Localization along the Rhabdomere Elongation in Drosophila Photoreceptor

BACKGROUND:Crumbs (Crb), a cell polarity gene, has been shown to provide a positional cue for the extension of the apical membrane domain, adherens junction (AJ), and rhabdomere along the growing proximal-distal axis during Drosophila photoreceptor morphogenesis. In developing Drosophila photoreceptors, a stabilized microtubule structure was discovered and its presence was linked to polarity protein localization. It was therefore hypothesized that the microtubules may provide trafficking routes for the polarity proteins during photoreceptor morphogenesis. This study has examined whether Kinesin heavy chain (Khc), a subunit of the microtubule-based motor Kinesin-1, is essential in polarity protein localization in developing photoreceptors. METHODOLOGY/PRINCIPAL FINDINGS:Because a genetic interaction was found between crb and khc, Crb localization was examined in the developing photoreceptors of khc mutants. khc was dispensable during early eye differentiation and development. However, khc mutant photoreceptors showed a range of abnormalities in the apical membrane domain depending on the position along the proximal-distal axis in pupal photoreceptors. The khc mutant showed a progressive mislocalization in the apical domain along the distal-proximal axis during rhabdomere elongation. The khc mutation also led to a similar progressive defect in the stabilized microtubule structures, strongly suggesting that Khc is essential for microtubule structure and Crb localization during distal to proximal rhabdomere elongation in pupal morphogenesis. This role of Khc in apical domain control was further supported by khc's gain-of-function phenotype. Khc overexpression in photoreceptors caused disruption of the apical membrane domain and the stabilized microtubules in the developing photoreceptors. CONCLUSIONS/SIGNIFICANCE:In summary, we examined the role of khc in the regulation of the apical Crb domain in developing photoreceptors. Since the rhabdomeres in developing pupal eyes grow along the distal-proximal axis, these phenotypes suggest that Khc is essential for the microtubule structures and apical membrane domains during the distal-proximal elongation of photoreceptors, but is dispensable for early eye development

Compartment syndrome in all four extremities: a rare case associated with systemic capillary leak syndrome

While isolated limb injuries often lead to suspicion for and treatment of compartment syndrome, multi-limb compartment syndrome induced by a systemic disease is a rare entity, in which lack of immediate diagnosis can have catastrophic consequences to the patient. We present a patient with idiopathic systemic capillary leak syndrome (SCLS) resulting in compartment syndrome in all four extremities. Treatment required bilateral hand, forearm, calf, and thigh fasciotomies. Presenting symptoms included pain in all four extremities, malaise, and confusion. Laboratory data included polycythemia, hypoalbuminemia, leukocytosis, and elevated creatinine, but not a monoclonal spike on serum electrophoresis as is common in SCLS. Thus, when faced with clinical evidence of multiple extremity compartment syndrome, the surgeon should consider systemic disorders, such as SCLS, and aggressively treat the compartment syndrome as well as the underlying disease