DC-SIGN and influenza hemagglutinin dynamics in plasma membrane microdomains are markedly different

DC-SIGN, a Ca²⁺-dependent transmembrane lectin, is found assembled in microdomains on the plasma membranes of dendritic cells. These microdomains bind a large variety of pathogens and facilitate their uptake for subsequent antigen presentation. In this study, DC-SIGN dynamics in microdomains were explored with several fluorescence microscopy methods and compared with dynamics for influenza hemagglutinin (HA), which is also found in plasma membrane microdomains. Fluorescence imaging indicated that DC-SIGN microdomains may contain other C-type lectins and that the DC-SIGN cytoplasmic region is not required for microdomain formation. Fluorescence recovery after photobleaching measurements showed that neither full-length nor cytoplasmically truncated DC-SIGN in microdomains appreciably exchanged with like molecules in other microdomains and the membrane surround, whereas HA in microdomains exchanged almost completely. Line-scan fluorescence correlation spectroscopy indicated an essentially undetectable lateral mobility for DC-SIGN but an appreciable mobility for HA within their respective domains. Single-particle tracking with defined-valency quantum dots confirmed that HA has significant mobility within microdomains, whereas DC-SIGN does not. By contrast, fluorescence recovery after photobleaching indicated that inner leaflet lipids are able to move through DC-SIGN microdomains. The surprising stability of DC-SIGN microdomains may reflect structural features that enhance pathogen uptake either by providing high-avidity platforms and/or by protecting against rapid microdomain endocytosis.     

Functional diversity of isoamylase oligomers: the ISA1 homo-oligomer is essential for amylopectin biosynthesis in rice endosperm

Rice (Oryza sativa) endosperm has two isoamylase (ISA) oligomers, ISA1 homo-oligomer and ISA1-ISA2 hetero-oligomer. To examine their contribution to starch synthesis, expression of the ISA1 or ISA2 gene was differently regulated in various transgenic plants. Although suppression of ISA2 gene expression caused the endosperm to have only the homo-oligomer, no significant effects were detected on the starch phenotypes. In contrast, ISA2 overexpression led to endosperm having only the hetero-oligomer, and starch synthesis in the endosperm was drastically impaired, both quantitatively and qualitatively, because the starch was devoid of typical starch features, such as thermal and x-ray diffraction properties, and water-soluble highly branched maltodextrins were accumulated. In the ISA2 overexpressed line, about 60% to 70% of the ISA1-ISA2 hetero-oligomer was bound to starch, while the ISA homo- and hetero-oligomers from the wild type were mostly present in the soluble form at the early milking stage of the endosperm. Detailed analysis of the relative amounts of homo- and hetero-oligomers in various lines also led us to the conclusion that the ISA1 homo-oligomer is essential, but not the ISA1-ISA2 oligomer, for starch production in rice endosperm. The relative amounts of ISA1 and ISA2 proteins were shown to determine the ratio of both oligomers and the stoichiometry of both ISAs in the hetero-oligomer. It was noted when compared with the homo-oligomer that all the hetero-oligomers from rice endosperm and leaf and potato (Solanum tuberosum) tuber were much more stable at 40°C. This study provides substantial data on the structural and functional diversity of ISA oligomers between plant tissues and species.     

Increased expression of lipocalin-type-prostaglandin D synthase in ulcerative colitis and exacerbating role in murine colitis

The pathogenesis of ulcerative colitis (UC) is unclear, but enhancement of disease activity by usage of nonsteroidal anti-inflammatory drugs suggests involvement of prostanoid in its pathophysiology. However, biological effect of prostaglandin (PG) D(2) on intestinal inflammation remains unknown. We investigated the expression of enzymes for PGD(2) synthesis, prostaglandin D synthase (PGDS), and its relation to the activity of colitis in UC patients. The role of lipocalin-type PGDS (L-PGDS) using a murine colitis model was also assessed. Tissue samples were obtained by colonic biopsies from patients with UC. Expression levels of mRNAs for L-PGDS and hematopoietic-type PGDS were investigated by quantitative RT-PCR. COX-2 and L-PGDS expression was investigated by immunohistochemistry. Localization of L-PGDS expression was also determined by in situ hybridization. In experimental study, mice were treated with dextran sodium sulfate in the drinking water to induce colitis. The degree of colonic inflammation was compared with L-PGDS(-/-) mice and control mice. The level of L-PGDS mRNA expression was increased in UC patients in parallel with disease activity. Colocalization of L-PGDS and cyclooxygenase (COX) 2 was observed in lamina proprial infiltrating cells and muscularis mucosa in UC patients. The level of hematopoietic PGDS mRNA expression did not differ from control mucosa. Dextran sodium sulfate treatment to L-PGDS(-/-) mice showed lower disease activity than control mice. We reported for the first time the presence of L-PGDS in the COX-2-expressing cells in the mucosa of active UC patients and that only L-PGDS increased with disease activity. An animal model study suggests that PGD(2) derived from L-PGDS-expressing cells plays proinflammatory roles in colitis.     

Seroprevalence of Antibodies Against Paracoccidioides Spp. in Captive Dolphins from Three Aquaria in Japan

Mycopathologia - The skin disease paracoccidioidomycosis ceti occurs in several dolphin species globally. Infection by the unculturable fungi Paracoccidioides brasilensis or other Paracoccidioides...     

Quantification of laminarialean zoospores in seawater by real‐time PCR

Laminarialean plants undergo heteromorphic alternation of generations between the macroscopic diploid sporophyte and the microscopic haploid gametophyte. The generations change through the formation and release of asexual or sexual reproductive cells. It is difficult to monitor the release and diffusion of zoospores into the environment. Furthermore, even if zoospores can be detected, species identification remains difficult. This study attempted to develop a sensitive and fast identification method for laminarialean zoospores using quantitative PCR. In addition, we aimed to estimate the density of zoospores in natural seawater. Specific primers for Saccharina japonica and Undaria pinnatifida were designed and used to estimate the quantity of zoospores in seawater. DNA samples (environmental DNA) were collected from seawater once or twice each month for 2 years at the same area, and seasonal variations in the release of zoospores was monitored. The estimated maturation period based on the number of released zoospores in this study was comparable with those of the previously reported maturation periods of S. japonica and U. pinnatifida sporophytes. This supports the validity of our method in estimating zoospore release from laminarialean plants. The method will be a useful tool for ecological studies on these commercially relevant species.     

Aldosterone: a mediator of retinal ganglion cell death and the potential role in the pathogenesis in normal-tension glaucoma

Glaucoma is conventionally defined as a chronic optic neuropathy characterized by progressive loss of retinal ganglion cells (RGCs) and optic nerve fibers. Although glaucoma is often associated with elevated intraocular pressure (IOP), significant IOP reduction does not prevent progression of the disease in some glaucoma patients. Thus, exploring IOP-independent mechanisms of RGC loss is important. We describe chronic systemic administration of aldosterone and evaluate its effect on RGCs in rat. Aldosterone was administered via an osmotic minipump that was implanted subcutaneously into the mid-scapular region. Although systemic administration of aldosterone caused RGC loss associated with thinning of the retinal nerve fiber layer without elevated IOP, the other cell layers appeared to be unaffected. After chronic administration of aldosterone, RGC loss was observed at 2 weeks in the peripheral retina and at 4 weeks in the central retina. However, administration of mineralocorticoid receptor blocker prevented RGC loss. These results demonstrate aldosterone is a critical mediator of RGC loss that is independent of IOP. We believe this rat normal-tension glaucoma (NTG) animal model not only offers a powerful system for investigating the mechanism of neurodegeneration in NTG, but can also be used to develop therapies directed at IOP-independent mechanisms of RGC loss.