Chloroplast biogenesis of photosystem II cores involves a series of assembly-controlled steps that regulate translation

The biogenesis of photosystem II, one of the major photosynthetic protein complexes, involves a cascade of assembly-governed regulation of translation of its major chloroplast-encoded subunits. In Chlamydomonas reinhardtii, the presence of the reaction center subunit D2 is required for the expression of the other reaction center subunit D1, while the presence of D1 is required for the expression of the core antenna subunit apoCP47. Using chimeric genes expressed in the chloroplast, we demonstrate that the decreased synthesis of D1 or apoCP47 in the absence of protein assembly is due to a genuine downregulation of translation. This regulation is mediated by the 5' untranslated region of the corresponding mRNA and originates from negative feedback exerted by the unassembled D1 or apoCP47 polypeptide. However, autoregulation of translation of subunit D1 is not implicated in the recovery from photoinhibition, which involves an increased translation of psbA mRNA in response to the degradation of photodamaged D1. De novo synthesis and repair of photosystem II complexes are independently controlled.     

Stable isotope evidence of diverse species-specific and individual wintering strategies in seabirds

Although there is increasing evidence that climatic variations during the non-breeding season shape population dynamics of seabirds, most aspects of their winter distribution and ecology remain essentially unknown. We used stable isotope signatures in feathers to infer and compare the moulting (wintering) habitat of subantarctic petrels breeding at two distant localities (South Georgia and Kerguelen). Petrels showed species-specific wintering habitat preferences, with a similar pattern of latitudinal segregation for all but one taxon. At both localities, delta13C values indicated that blue petrels (Halobaena caerulea) moult in Antarctic waters, South Georgian diving petrels (Pelecanoides georgicus) in the vicinity of the archipelagos and/or in the Polar Frontal Zone and Antarctic prions (Pachyptila desolata) in warmer waters. In contrast, common diving petrels (Pelecanoides urinatrix) showed divergent strategies, with low and high intrapopulation variation at South Georgia and Kerguelen, respectively. Birds from Kerguelen dispersed over a much wider range of habitats, from coastal to oceanic waters and from Antarctica to the subtropics, whereas those from South Georgia wintered mainly in waters around the archipelago. This study is the first to show such striking between-population heterogeneity in individual wintering strategies, which could have important implications for likely demographic responses to environmental perturbation.     

Systemic dysregulation of CEACAM1 in melanoma patients

It was previously shown that CEACAM1 on melanoma cells strongly predicts poor outcome. Here, we show a statistically significant increase of serum CEACAM1 in 64 active melanoma patients, as compared to 48 patients with no evidence of disease and 37 healthy donors. Among active patients, higher serum CEACAM1 correlated with LDH values and with decreased survival. Multivariate analysis with neutralization of LDH showed that increased serum CEACAM1 carries a hazard ratio of 2.40. In vitro, soluble CEACAM1 was derived from CEACAM1(+), but neither from CEACAM1(?) melanoma cells nor from CEACAM1(+) lymphocytes, and directly correlated with the number of CEACAM1(+) melanoma cells. Production of soluble CEACAM1 depended on intact de novo protein synthesis and secretion machineries, but not on metalloproteinase function. An unusually high percentage of CEACAM1(+) circulating NK and T lymphocytes was demonstrated in melanoma patients. CEACAM1 inhibited killing activity in functional assays. CEACAM1 expression could not be induced on lymphocytes by serum from patients with high CEACAM1 expression. Further, expression of other NK receptors was impaired, which collectively indicate on a general abnormality. In conclusion, the systemic dysregulation of CEACAM1 in melanoma patients further denotes the role of CEACAM1 in melanoma and may provide a basis for new tumor monitoring and prognostic platforms.     

Glycosylation of PrPC Determines Timing of Neuroinvasion and Targeting in the Brain following Transmissible Spongiform Encephalopathy Infection by a Peripheral Route

Transmissible spongiform encephalopathy (TSE) infectivity naturally spreads from site of entry in the periphery to the central nervous system where pathological lesions are formed. Several routes and cells within the host have been identified as important for facilitating the infectious process. Expression of the glycoprotein cellular PrP (PrP^C) is considered a key factor for replication of infectivity in the central nervous system (CNS) and its transport to the brain, and it has been suggested that the infectious agent propagates from cell to cell via a domino-like effect. However, precisely how this is achieved and what involvement the different glycoforms of PrP have in these processes remain to be determined. To address this issue, we have used our unique models of gene-targeted transgenic mice expressing different glycosylated forms of PrP. Two TSE strains were inoculated intraperitoneally into these mice to assess the contribution of diglycosylated, monoglycosylated, and unglycosylated PrP in spreading of infectivity to the brain. This study demonstrates that glycosylation of host PrP has a profound effect in determining the outcome of disease. Lack of diglycosylated PrP slowed or prevented disease onset after peripheral challenge, suggesting an important role for fully glycosylated PrP in either the replication of the infectious agent in the periphery or its transport to the CNS. Moreover, mice expressing unglycosylated PrP did not develop clinical disease, and mice expressing monoglycosylated PrP showed strikingly different neuropathologic features compared to those expressing diglycosylated PrP. This demonstrates that targeting in the brain following peripheral inoculation is profoundly influenced by the glycosylation status of host PrP.Transmissible spongiform encephalopathies (TSE) or prion diseases are a group of fatal neurodegenerative diseases which include Creutzfeldt-Jakob disease (CJD) in humans, scrapie in sheep and goats, bovine spongiform encephalopathies (BSE) in cattle, and chronic wasting disease (CWD) in deer and elk (30). These diseases can be sporadic, familial, or acquired by infection, and the common hallmark is a distinct pathology in the central nervous system (CNS) characterized by neuronal loss, spongiform degeneration, and gliosis (38, 46).Expression of the host-encoded cellular PrP (PrP^C) is fundamental for the onset of disease since PrP-deficient mice are refractory to TSE infection (11, 31). PrP^C is a glycoprotein with two consensus sites for attachment of N-linked glycans (at codons 180 and 196 in the mouse) which are variably occupied, producing di-, mono-, and unglycosylated PrP (43). The diversity in glycosylation, combined with the complexity of added sugars, results in a large number of glycosylated forms of PrP (41). A central event associated with TSE infection is the conformational conversion of PrP^C into an abnormal protease-resistant form, PrP^Sc (39). PrP^Sc is deposited in brain and, in some but not all cases, in peripheral organs of individuals affected by TSE (21).Although the pathology associated with TSE is found in the brain, the periphery is the most natural route of acquiring infection. Evidence suggests that oral transmission via contaminated food is linked with transmission of BSE to humans, resulting in variant CJD (vCJD) (10, 47), and blood transfusion has been identified as a probable route of human-to-human transmission of vCJD (23, 27, 36). Moreover, parenteral administration of contaminated human tissue-derived therapeutics has been shown to facilitate iatrogenic spread of these diseases (8, 46). It is therefore important to understand the mechanisms that allow the infectious agent to propagate in the periphery and be transported to the CNS prior to the onset of neurodegeneration in the brain.Many studies have been conducted to understand routes of transmission (for a review see references 1 and 29). Lymphoid tissues such as the spleen have been shown to play a fundamental role in agent replication and propagation in the very early stages of disease. Indeed, studies of splenectomized and asplenic mice have shown the lymphoreticular system (LRS) to be an important site for TSE agent replication (14, 26). The periphery also appears to have a role in processing the infectious agent following intracerebral (i.c.) inoculation as PrP^Sc accumulates in the spleen shortly after inoculation and before accumulation of the abnormal protein in the brain (15, 17). Within the LRS, follicular dendritic cells (FDC) have been shown to be important for the uptake of infectivity and subsequent spreading toward the CNS (7, 28, 33, 35). Several studies have also suggested the peripheral nervous systems (PNS) as a potential route of infectivity to the brain, implicating the vagus and sciatic nerves in this process (5, 20, 25, 34).Expression of PrP^C in the peripheral tissues appears to be an important prerequisite for the transport of infectivity to the CNS following peripheral routes of inoculation. Indeed, it has been proposed that a continuous chain of cells expressing PrP^C is fundamental for TSE neuroinvasion (6, 40), with overexpression of endogenous PrP in the PNS greatly facilitating the spread of infectivity (19). Thus, host PrP appears to have a fundamental role in the uptake, transport, and replication of the infectious agent (6). Moreover, it has been suggested that the different PrP^C glycoforms may influence the timing of neuroinvasion by directly influencing the interaction with the infectious agent (19). However, the mechanism by which the different glycoforms are involved in these processes remains to be determined.In order to investigate the role of PrP^C glycosylation in TSE disease after peripheral infection with different TSE strains, we have used our inbred gene-targeted transgenic mice expressing different glycosylated forms of PrP. These mice expressed PrP with no sugars at the first (designated G1/G1 in homozygous mice) or the second glycosylation site (G2/G2) or both (G3/G3) under the control of the endogenous PrP promoter (13). We have previously shown that following intracerebral inoculation, all glycotypes are susceptible to infection with at least one TSE strain and that the type of PrP glycosylation in the host influenced the incubation period but not the distribution of pathological lesions in the brain (45). Here, we examine the influence of host PrP glycosylation on the peripheral acquisition of infection and demonstrate that, unlike the intracerebral route, mice without PrP glycosylation were resistant to disease and that the different glycoforms had a profound influence on not only the timing of disease but also the type and distribution of the PrP^Sc deposits in the brain.     

Pulse stimulation with odors or IBMX/forskolin potentiates responses in isolated olfactory neurons

Many odor responses are mediated by the adenosine 3',5'-cyclic monophosphate (cAMP) pathway in which the cAMP-gated current is amplified by Ca2+-dependent Cl- current. In olfactory neurons, prolonged exposure to odors decreases the odor response and is an adaptive effect. Several studies suggest that odor adaptation is linked to elevated intracellular Ca2+. In the present study, using the perforated configuration of the patch clamp technique, we found that repetitive odor stimulation elicits a potentiation of the subsequent responses in olfactory neurons. This potentiation is mimicked by stimulating the cAMP pathway and does not appear to be related to phosphorylation of ion channels since protein kinase inhibitors could not block it. Our data suggest that local increases in [Ca2+]i via activation of the cAMP pathway mediate the pulse-elicited potentiation. In the first odor application, entry of Ca2+ through cyclic nucleotide-gated channels appears to be buffered. Repetitive stimulation allows local increases in [Ca2+]i, recruiting more Ca2+-dependent Cl- channels with each subsequent odor pulse.     

Transepithelial pathogen uptake into the small intestinal lamina propria

The lamina propria that underlies and stabilizes the gut lining epithelium is densely populated with strategically located mononuclear phagocytes. Collectively, these lamina propria macrophages and dendritic cells (DC) are believed to be crucial for tissue homeostasis as well as the innate and adaptive host defense. Lamina propria DC were recently shown to gain direct access to the intestinal lumen by virtue of epithelium-penetrating dendrites. However, the role of these structures in pathogen uptake remains under debate. In this study, we report that entry of a noninvasive model pathogen (Aspergillus fumigatus conidia) into the murine small intestinal lamina propria persists in the absence of either transepithelial dendrites or lamina propria DC and macrophages. Our results suggest the existence of multiple pathogen entry pathways and point at the importance of villus M cells in the uptake of gut lumen Ags. Interestingly, transepithelial dendrites seem altogether absent from the small intestine of BALB/c mice suggesting that the function of lamina propria DC extensions resides in their potential selectivity for luminal Ags, rather than in general uptake or gut homeostasis.     

Overexpression of inducible heat shock protein 70 and its mutants in astrocytes is associated with maintenance of mitochondrial physiology during glucose deprivation stress

Wild-type inducible Hsp70 (WT) and 2 folding deficient mutants protect the brain against focal cerebral ischemia in vivo and brain cells from oxygen-glucose deprivation (OGD) in vitro, but the protective mechanisms remain unclear. Mitochondria are central to both normal physiological function and the regulation of cell death. We tested the effect of overexpressing Hsp70 and 2 mutants, Hsp70-K71 E, an adenosine triphosphatase (ATPase)-deficient point mutant, and Hsp70-381-640, a deletion mutant lacking the ATPase domain on mitochondrial physiology under glucose deprivation (GD) stress in primary cultured astrocytes. Mitochondrial membrane potential was assessed using a potentiometric fluorescent dye tetramethylrhodamine ethyl ester (TMRE). By 5 hours of GD, the mitochondria in the LXSN control transfected astrocytes had markedly reduced membrane potential. However, in the Hsp70-WT, -K71E, and -381-640 groups, there was no apparent change in TMRE signal during 5 hours of GD. Oxygen consumption was measured to assess oxidative respiration. Overexpression of Hsp70-K71 E and -381-640 prevented the decrease in state III respiration observed at 5 hours, and all 3 prevented the increase in state IV respiration found in LXSN controls after 5 hours of GD. Reactive oxygen species (ROS) production was assessed with hydroethidine. Hsp70 and its mutants all significantly reduced the increases in ROS accumulation during 5 hours of GD. The results demonstrate that the protective effect of the carboxyl-terminal half of Hsp70 and of the full-length molecule is associated with better maintained mitochondrial membrane potential, better maintained state IV respiration, and reduced ROS generation during GD.