Summary In order to study the synthesis and degradation processes of the photoreceptor membranes in the abalone, Nordotis discus, the localization of acyltransferase and acid hydrolase activities, respectively, were determined at the electron-microscopic level. Acyltransferase activity was localized on the cytoplasmic sides of thick (>10 nm) membranes of the following organelles: a few cisternae at the trans (or concave) side of Golgi apparatus, Golgi and probably related vesicles, short tubules, curved pentalaminar disks and limiting membranes of the phagosomal multivesicular bodies; all organelles were scattered in the peri- to supranuclear cytoplasm. The phospholipids, which are major components of the photoreceptor membrane, are considered to be synthesized by these membranes. Acid phosphatase activity was localized in the lumina of Golgi cisternae and vesicles, lysosomes, and smaller multivesicular and related bodies, but not in multilamellar bodies. The matrices of the larger multivesicular bodies and of the pigment granule complexes showed arylsulfatase activity. Vesiculated and autophagocytosed photoreceptor microvilli seemed to be degraded by acid hydrolases, forming multivesicular and related bodies. Supporting cells also showed acyltransferase and acid hydrolase activities.Abbreviations used in this Paper AcP
acid phosphatase
- ArS
arylsulfatase
- AT
acyltransferase
- ER
endoplasmic reticulum
- GERL
Golgi-endoplasmic reticulum-lysosomal complex
- MEB
meshwork body
- MLB
multilamellar body
- MVB
multivesicular body
- VLB
vesiculolamellar body 相似文献
Selective membrane transport pathways are essential for cells in situ to construct and maintain a polarized structure comprising multiple plasma membrane domains, which is essential for their specific cellular functions. Genetic screening in Drosophila photoreceptors harboring multiple plasma membrane domains enables the identification of genes involved in polarized transport pathways. Our genome-wide high-throughput screening identified a Rab6-null mutant with a rare phenotype characterized by a loss of 2 apical transport pathways with an intact basolateral transport. Although the functions of Rab6 in the Golgi apparatus are well known, its function in polarized transport is unexpected.
The mutant phenotype and localization of Rab6 strongly indicate that Rab6 regulates transport between the trans-Golgi network (TGN) and recycling endosomes (REs): basolateral cargos are segregated at the TGN before Rab6 functions, but cargos going to multiple apical domains are sorted at REs. Both the medial-Golgi resident protein Metallophosphoesterase (MPPE) and the TGN marker GalT::CFP exhibit diffused co-localized distributions in Rab6-deficient cells, suggesting they are trapped in the retrograde transport vesicles returning to trans-Golgi cisternae. Hence, we propose that Rab6 regulates the fusion of retrograde transport vesicles containing medial, trans-Golgi resident proteins to the Golgi cisternae, which causes Golgi maturation to REs. 相似文献
Pyrethroid insecticides are the front line vector control tools used in bed nets to reduce malaria transmission and its burden. However, resistance in major vectors such as Anopheles arabiensis is posing a serious challenge to the success of malaria control.Herein, we elucidated the molecular and biochemical basis of pyrethroid resistance in a knockdown resistance-free Anopheles arabiensis population from Chad, Central Africa. Using heterologous expression of P450s in Escherichia coli coupled with metabolism assays we established that the over-expressed P450 CYP6P4, located in the major pyrethroid resistance (rp1) quantitative trait locus (QTL), is responsible for resistance to Type I and Type II pyrethroid insecticides, with the exception of deltamethrin, in correlation with field resistance profile. However, CYP6P4 exhibited no metabolic activity towards non-pyrethroid insecticides, including DDT, bendiocarb, propoxur and malathion. Combining fluorescent probes inhibition assays with molecular docking simulation, we established that CYP6P4 can bind deltamethrin but cannot metabolise it. This is possibly due to steric hindrance because of the large vdW radius of bromine atoms of the dihalovinyl group of deltamethrin which docks into the heme catalytic centre.The establishment of CYP6P4 as a partial pyrethroid resistance gene explained the observed field resistance to permethrin, and its inability to metabolise deltamethrin probably explained the high mortality from deltamethrin exposure in the field populations of this Sudano-Sahelian An. arabiensis. These findings describe the heterogeneity in resistance towards insecticides, even from the same class, highlighting the need to thoroughly understand the molecular basis of resistance before implementing resistance management/control tools. 相似文献
Oxidative stress is a main factor responsible for key changes leading to the onset of age-related macular degeneration (ARMD) that occur in the retinal pigment epithelium (RPE), which is involved in phagocytosis of photoreceptor outer segments (POS). In this study, hydrogen peroxide (H2O2), H2O2 and iron ions (Fe) or rose Bengal (RB) in the presence of NADH and Fe were used to model free radical mediated oxidative stress to test if free radicals and singlet oxygen have different efficiency to inhibit phagocytosis of ARPE-19 cells. Free radical mediated oxidative stress was confirmed by HPLC-EC(Hg) measurements of cholesterol hydroperoxides in treated cells. Electron paramagnetic resonance (EPR) spin trapping was employed to detect superoxide anion. Cell survival was analyzed by the MTT assay. Specific phagocytosis of fluorescein-5-isothiocyanate-labeled POS and non-specific phagocytosis of fluorescent beads were measured by flow cytometry. HPLC analysis of cells photosensitized with RB in the presence of NADH and Fe indicated substantial increase in formation of free radical-dependent 7α/7β-hydroperoxides. EPR spin trapping confirmed the photogeneration of superoxide anion in samples enriched with RB, NADH and Fe. For all three protocols sub-lethal oxidative stress induced significant inhibition of the specific phagocytosis of POS. In contrast, non-specific phagocytosis was inhibited only by H2O2 or H2O2 and Fe treatment. Inhibition of phagocytosis was transient and recoverable by 24?h. These results suggest that free radicals may exert similar to singlet oxygen efficiency in inhibiting phagocytosis of RPE cells, and that the effect depends on the location where initial reactive species are formed. 相似文献
The 27-kDa outer membrane protein from eight strains of Coxiella burnetii was expressed in the pET-21c protein expression system. Two fusion proteins with molecular masses of 30 and 32 kDa were evident in all eight of the recombinants by SDS-PAGE and immunoblotting. A protein having an approximate size of 30 kDa was purified from the Escherichia coli lysates by one-step affinity purification. The utility of the purified recombinant protein in ELISA was also evaluated by testing its reactivity with human sera and comparing this reactivity with that of Nine Mile phase II antigen. All of the 40 IF-positive serum samples were ELISA-positive for both the Nine Mile phase II and recombinant antigens, and negative serum controls were negative for both antigens. These results suggest that ELISA with the 27-kDa recombinant antigen is a sensitive and specific method for detecting anti-C. burnetii antibodies in human sera. 相似文献
The intermediate filament keratin, K15, is present in variable abundance in stratified epithelia. In this study we have isolated and characterized the sheepK15gene, focusing on its expression in the follicles of sheep and mice. We show thatK15is expressed throughout the hair cycle in the basal layer of the outer root sheath that envelops the follicle. Strikingly, however, in large medullated wool follicles, a small group of basal outer root sheath cells located in the region thought to contain hair follicle stem cells areK15-negative. In the follicle bulbK15is expressed in cells situated next to the dermal papilla but not in the inner bulb cells. Elsewhere,K15is expressed at a low, variable level in the basal layer of the epidermis and sebaceous gland, often in a punctate pattern. In the esophagus of the sheepK15expression is restricted to the basal layer, in contrast to human esophagus where it is expressed throughout the epithelium. Transgenic mouse lines established with a 15-kb sheepK15gene construct exhibited faithful expression and showed no phenotypic consequences ofK15overexpression. An investigation of transgene expression showed thatK15is continuously expressed in outer root sheath cells during the hair cycle. Given its expression in the mitotically active basal cell layers of diverse epithelia and the follicle,K15expression appears to signal an early stage in the pathway of keratinocyte differentiation that precedes the decision of a cell to become epidermal or hair-like. 相似文献