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Mutations in LRBA, a BEACH domain protein, cause severe immune deficiency in humans. LRBA is expressed in many tissues and organs according to biochemical analysis, but little is known about its cellular and subcellular localization, and its deficiency phenotype outside the immune system. By LacZ histochemistry of Lrba gene-trap mice, we performed a comprehensive survey of LRBA expression in numerous tissues, detecting it in many if not all epithelia, in exocrine and endocrine cells, and in subpopulations of neurons. Immunofluorescence microscopy of the exocrine and endocrine pancreas, salivary glands, and intestinal segments, confirmed these patterns of cellular expression and provided information on the subcellular localizations of the LRBA protein. Immuno-electron microscopy demonstrated that in neurons and endocrine cells, which co-express LRBA and its closest relative, neurobeachin, both proteins display partial association with endomembranes in complementary, rather than overlapping, subcellular distributions. Prominent manifestations of human LRBA deficiency, such as inflammatory bowel disease or endocrinopathies, are believed to be primarily due to immune dysregulation. However, as essentially all affected tissues also express LRBA, it is possible that LRBA deficiency enhances their vulnerability and contributes to the pathogenesis.

Mutations in LRBA, a BEACH domain protein, cause severe immune deficiency in humans. LRBA is expressed in many tissues and organs according to biochemical analysis, but little is known about its cellular and subcellular localization, and its deficiency phenotype outside the immune system. By LacZ histochemistry of Lrba gene-trap mice, we performed a comprehensive survey of LRBA expression in numerous tissues, detecting it in many if not all epithelia, in exocrine and endocrine cells, and in subpopulations of neurons. Immunofluorescence microscopy of the exocrine and endocrine pancreas, salivary glands, and intestinal segments, confirmed these patterns of cellular expression and provided information on the subcellular localizations of the LRBA protein. Immuno-electron microscopy demonstrated that in neurons and endocrine cells, which co-express LRBA and its closest relative, neurobeachin, both proteins display partial association with endomembranes in complementary, rather than overlapping, subcellular distributions. Prominent manifestations of human LRBA deficiency, such as inflammatory bowel disease or endocrinopathies, are believed to be primarily due to immune dysregulation. However, as essentially all affected tissues also express LRBA, it is possible that LRBA deficiency enhances their vulnerability and contributes to the pathogenesis.

Mutations in LRBA, a BEACH domain protein, cause severe immune deficiency in humans. LRBA is expressed in many tissues and organs according to biochemical analysis, but little is known about its cellular and subcellular localization, and its deficiency phenotype outside the immune system. By LacZ histochemistry of Lrba gene-trap mice, we performed a comprehensive survey of LRBA expression in numerous tissues, detecting it in many if not all epithelia, in exocrine and endocrine cells, and in subpopulations of neurons. Immunofluorescence microscopy of the exocrine and endocrine pancreas, salivary glands, and intestinal segments, confirmed these patterns of cellular expression and provided information on the subcellular localizations of the LRBA protein. Immuno-electron microscopy demonstrated that in neurons and endocrine cells, which co-express LRBA and its closest relative, neurobeachin, both proteins display partial association with endomembranes in complementary, rather than overlapping, subcellular distributions. Prominent manifestations of human LRBA deficiency, such as inflammatory bowel disease or endocrinopathies, are believed to be primarily due to immune dysregulation. However, as essentially all affected tissues also express LRBA, it is possible that LRBA deficiency enhances their vulnerability and contributes to the pathogenesis.

Mutations in LRBA, a BEACH domain protein, cause severe immune deficiency in humans. LRBA is expressed in many tissues and organs according to biochemical analysis, but little is known about its cellular and subcellular localization, and its deficiency phenotype outside the immune system. By LacZ histochemistry of Lrba gene-trap mice, we performed a comprehensive survey of LRBA expression in numerous tissues, detecting it in many if not all epithelia, in exocrine and endocrine cells, and in subpopulations of neurons. Immunofluorescence microscopy of the exocrine and endocrine pancreas, salivary glands, and intestinal segments, confirmed these patterns of cellular expression and provided information on the subcellular localizations of the LRBA protein. Immuno-electron microscopy demonstrated that in neurons and endocrine cells, which co-express LRBA and its closest relative, neurobeachin, both proteins display partial association with endomembranes in complementary, rather than overlapping, subcellular distributions. Prominent manifestations of human LRBA deficiency, such as inflammatory bowel disease or endocrinopathies, are believed to be primarily due to immune dysregulation. However, as essentially all affected tissues also express LRBA, it is possible that LRBA deficiency enhances their vulnerability and contributes to the pathogenesis.

Mutations in LRBA, a BEACH domain protein, cause severe immune deficiency in humans. LRBA is expressed in many tissues and organs according to biochemical analysis, but little is known about its cellular and subcellular localization, and its deficiency phenotype outside the immune system. By LacZ histochemistry of Lrba gene-trap mice, we performed a comprehensive survey of LRBA expression in numerous tissues, detecting it in many if not all epithelia, in exocrine and endocrine cells, and in subpopulations of neurons. Immunofluorescence microscopy of the exocrine and endocrine pancreas, salivary glands, and intestinal segments, confirmed these patterns of cellular expression and provided information on the subcellular localizations of the LRBA protein. Immuno-electron microscopy demonstrated that in neurons and endocrine cells, which co-express LRBA and its closest relative, neurobeachin, both proteins display partial association with endomembranes in complementary, rather than overlapping, subcellular distributions. Prominent manifestations of human LRBA deficiency, such as inflammatory bowel disease or endocrinopathies, are believed to be primarily due to immune dysregulation. However, as essentially all affected tissues also express LRBA, it is possible that LRBA deficiency enhances their vulnerability and contributes to the pathogenesis.

The protein machinery of neurotransmitter exocytosis requires efficient orchestration in space and time, for speed and precision of neurotransmission and also for synaptic ontogeny and plasticity. However, its spatial organization in situ is virtually unknown. Aczonin/Piccolo is a putative organizer protein of mammalian active zones. We determined by immunogold electron microscopy (EM) (i) the spatial arrangement (i. e., topology) of 11 segments of the Aczonin polypeptide in situ, and correlated it to (ii) the positioning of Aczonin-interacting domains of Bassoon, CAST/ELKS, Munc13, and RIM and (iii) the ultrastructurally defined presynaptic macromolecular aggregates known as dense projections and synaptic ribbons. At conventional synapses, Aczonin assumes a compact molecular topology within a layer 35 to 80 nm parallel to the plasma membrane (PM), with a "trunk" sitting on the dense projection top and a C-terminal "arm" extending down toward the PM and sideward to the dense projection periphery. At ribbon synapses, Aczonin occupies the whole ribbon area. Bassoon colocalizes with Aczonin at conventional synapses but not at ribbon synapses. At both conventional and ribbon synapses, CAST, Munc13, and RIM are segregated from Aczonin, closer to the PM, and Aczonin is positioned such that it may control the access of neurotransmitter vesicles to the fusion site.

Iron sequestration by host iron-binding proteins is an important mechanism of resistance to microbial infections. Inside oral epithelial cells, iron is stored within ferritin, and is therefore not usually accessible to pathogenic microbes. We observed that the ferritin concentration within oral epithelial cells was directly related to their susceptibility to damage by the human pathogenic fungus, Candida albicans. Thus, we hypothesized that host ferritin is used as an iron source by this organism. We found that C. albicans was able to grow on agar at physiological pH with ferritin as the sole source of iron, while the baker's yeast Saccharomyces cerevisiae could not. A screen of C. albicans mutants lacking components of each of the three known iron acquisition systems revealed that only the reductive pathway is involved in iron utilization from ferritin by this fungus. Additionally, C. albicans hyphae, but not yeast cells, bound ferritin, and this binding was crucial for iron acquisition from ferritin. Transcriptional profiling of wild-type and hyphal-defective C. albicans strains suggested that the C. albicans invasin-like protein Als3 is required for ferritin binding. Hyphae of an Deltaals3 null mutant had a strongly reduced ability to bind ferritin and these mutant cells grew poorly on agar plates with ferritin as the sole source of iron. Heterologous expression of Als3, but not Als1 or Als5, two closely related members of the Als protein family, allowed S. cerevisiae to bind ferritin. Immunocytochemical localization of ferritin in epithelial cells infected with C. albicans showed ferritin surrounding invading hyphae of the wild-type, but not the Deltaals3 mutant strain. This mutant was also unable to damage epithelial cells in vitro. Therefore, C. albicans can exploit iron from ferritin via morphology dependent binding through Als3, suggesting that this single protein has multiple virulence attributes.

Iron sequestration by host iron-binding proteins is an important mechanism of resistance to microbial infections. Inside oral epithelial cells, iron is stored within ferritin, and is therefore not usually accessible to pathogenic microbes. We observed that the ferritin concentration within oral epithelial cells was directly related to their susceptibility to damage by the human pathogenic fungus, Candida albicans. Thus, we hypothesized that host ferritin is used as an iron source by this organism. We found that C. albicans was able to grow on agar at physiological pH with ferritin as the sole source of iron, while the baker's yeast Saccharomyces cerevisiae could not. A screen of C. albicans mutants lacking components of each of the three known iron acquisition systems revealed that only the reductive pathway is involved in iron utilization from ferritin by this fungus. Additionally, C. albicans hyphae, but not yeast cells, bound ferritin, and this binding was crucial for iron acquisition from ferritin. Transcriptional profiling of wild-type and hyphal-defective C. albicans strains suggested that the C. albicans invasin-like protein Als3 is required for ferritin binding. Hyphae of an Deltaals3 null mutant had a strongly reduced ability to bind ferritin and these mutant cells grew poorly on agar plates with ferritin as the sole source of iron. Heterologous expression of Als3, but not Als1 or Als5, two closely related members of the Als protein family, allowed S. cerevisiae to bind ferritin. Immunocytochemical localization of ferritin in epithelial cells infected with C. albicans showed ferritin surrounding invading hyphae of the wild-type, but not the Deltaals3 mutant strain. This mutant was also unable to damage epithelial cells in vitro. Therefore, C. albicans can exploit iron from ferritin via morphology dependent binding through Als3, suggesting that this single protein has multiple virulence attributes.

Palmdelphin is a newly identified cytosolic isoform of paratemmin-1, a lipid raft-associated protein implicated in cell shape control. Like paralemmin- 1,palmdelphin is phosphorylated, giving rise to electrophoretic band heterogeneity that is most pronounced in the brain. In ultracentrifugation and get filtration palmdelphin behaves as a non-globular monomer. Its C-terminal region binds glutamine synthetase. Immunohistochemical analysis of the rat brain shows a prominent localization of palmdelphin in the cerebral cortex, hippocampus, amygdala, septum, indusium griseum, piriform cortex, nucleus supraopticus, and nucleus of the lateral olfactory tract. Many of the circumscript palmdelphin-positive areas are related to the olfactory system. Immunoperoxidase electron microscopy reveals a discontinuous distribution of palmdelphin immunoreactivity, in the form of spots scattered throughout the cytoplasm of selected neuronal perikarya and dendrites, including dendritic spines, often in association with endomembranes, and in a pattern which is similar to that of the cytoplasmic fraction of paralemmin-1. In subcellular fractionation experiments palmdelphin behaves as a cytosolic protein which, however, can be partially recruited from cytosol to the detergent-resistant fraction of a membrane/cytoskeletal cell ghost preparation. These observations suggest that palmdelphin may peripherally associate with endomembranes or cytoskeleton-linked structures.