Differential regulation of cell death programs in males and females by Poly (ADP-Ribose) Polymerase-1 and 17 β estradiol

Cell death can be divided into the anti-inflammatory process of apoptosis and the pro-inflammatory process of necrosis. Necrosis, as apoptosis, is a regulated form of cell death, and Poly-(ADP-Ribose) Polymerase-1 (PARP-1) and Receptor-Interacting Protein (RIP) 1/3 are major mediators. We previously showed that absence or inhibition of PARP-1 protects mice from nephritis, however only the male mice. We therefore hypothesized that there is an inherent difference in the cell death program between the sexes. We show here that in an immune-mediated nephritis model, female mice show increased apoptosis compared to male mice. Treatment of the male mice with estrogens induced apoptosis to levels similar to that in female mice and inhibited necrosis. Although PARP-1 was activated in both male and female mice, PARP-1 inhibition reduced necrosis only in the male mice. We also show that deletion of RIP-3 did not have a sex bias. We demonstrate here that male and female mice are prone to different types of cell death. Our data also suggest that estrogens and PARP-1 are two of the mediators of the sex-bias in cell death. We therefore propose that targeting cell death based on sex will lead to tailored and better treatments for each gender.     

Acquired Antibiotic Resistance: Are We Born with It?

The rapid emergence of antibiotic resistance (AR) is a major public health concern. Recent findings on the prevalence of food-borne antibiotic-resistant (ART) commensal bacteria in ready-to-consume food products suggested that daily food consumption likely serves as a major avenue for dissemination of ART bacteria from the food chain to human hosts. To properly assess the impact of various factors, including the food chain, on AR development in hosts, it is important to determine the baseline of ART bacteria in the human gastrointestinal (GI) tract. We thus examined the gut microbiota of 16 infant subjects, from the newborn stage to 1 year of age, who fed on breast milk and/or infant formula during the early stages of development and had no prior exposure to antibiotics. Predominant bacterial populations resistant to several antibiotics and multiple resistance genes were found in the infant GI tracts within the first week of age. Several ART population transitions were also observed in the absence of antibiotic exposure and dietary changes. Representative AR gene pools including tet(M), ermB, sul2, and bla(TEM) were detected in infant subjects. Enterococcus spp., Staphylococcus spp., Klebsiella spp., Streptococcus spp., and Escherichia coli/Shigella spp. were among the identified AR gene carriers. ART bacteria were not detected in the infant formula and infant foods examined, but small numbers of skin-associated ART bacteria were found in certain breast milk samples. The data suggest that the early development of AR in the human gut microbiota is independent of infants' exposure to antibiotics but is likely impacted by exposure to maternal and environmental microbes during and after delivery and that the ART population is significantly amplified within the host even in the absence of antibiotic selective pressure.     

Aquaporin-2 in the ��-omics�� Era

Vasopressin controls renal water excretion largely through actions to regulate the water channel aquaporin-2 in collecting duct principal cells. Our knowledge of the mechanisms involved has increased markedly in recent years with the advent of methods for large-scale systems-level profiling such as protein mass spectrometry, yeast two-hybrid analysis, and oligonucleotide microarrays. Here we review this progress.Regulation of water excretion by the kidney is one of the most visible aspects of everyday physiology. An outdoor tennis game on a hot summer day can result in substantial water losses by sweating, and the kidneys respond by reducing water excretion. In contrast, excessive intake of water, a frequent occurrence in everyday life, results in excretion of copious amounts of clear urine. These responses serve to exact tight control on the tonicity of body fluids, maintaining serum osmolality in the range of 290–294 mosmol/kg of H₂O through the regulated return of water from the pro-urine in the renal collecting ducts to the bloodstream.The importance of this process is highlighted when the regulation fails. For example, polyuria (rapid uncontrolled excretion of water) is a sometimes devastating consequence of lithium therapy for bipolar disorder. On the other side of the coin are water balance disorders that result from excessive renal water retention causing systemic hypo-osmolality or hyponatremia. Hyponatremia due to excessive water retention can be seen with severe congestive heart failure, hepatic cirrhosis, and the syndrome of inappropriate antidiuresis.The chief regulator of water excretion is the peptide hormone AVP,² whereas the chief molecular target for regulation is the water channel AQP2. In this minireview, we describe new progress in the understanding of the molecular mechanisms involved in regulation of AQP2 by AVP in collecting duct cells, with emphasis on new information derived from “systems-level” approaches involving large-scale profiling and screening techniques such as oligonucleotide arrays, protein mass spectrometry, and yeast two-hybrid analysis. Most of the progress with these techniques is in the identification of individual molecules involved in AVP signaling and binding interactions with AQP2. Additional related issues are addressed in several recent reviews (1–4).     

Klebsiella Phage vB_KleM-RaK2 — A Giant Singleton Virus of the Family Myoviridae

At 346 kbp in size, the genome of a jumbo bacteriophage vB_KleM-RaK2 (RaK2) is the largest Klebsiella infecting myovirus genome sequenced to date. In total, 272 out of 534 RaK2 ORFs lack detectable database homologues. Based on the similarity to biologically defined proteins and/or MS/MS analysis, 117 of RaK2 ORFs were given a functional annotation, including 28 RaK2 ORFs coding for structural proteins that have no reliable homologues to annotated structural proteins in other organisms. The electron micrographs revealed elaborate spike-like structures on the tail fibers of Rak2, suggesting that this phage is an atypical myovirus. While head and tail proteins of RaK2 are mostly myoviridae-related, the bioinformatics analysis indicate that tail fibers/spikes of this phage are formed from podovirus-like peptides predominantly. Overall, these results provide evidence that bacteriophage RaK2 differs profoundly from previously studied viruses of the Myoviridae family.     

Does Leaf Position within a Canopy Affect Acclimation of Photosynthesis to Elevated CO2? : Analysis of a Wheat Crop under Free-Air CO2 Enrichment

Previous studies of photosynthetic acclimation to elevated CO₂ have focused on the most recently expanded, sunlit leaves in the canopy. We examined acclimation in a vertical profile of leaves through a canopy of wheat (Triticum aestivum L.). The crop was grown at an elevated CO₂ partial pressure of 55 Pa within a replicated field experiment using free-air CO₂ enrichment. Gas exchange was used to estimate in vivo carboxylation capacity and the maximum rate of ribulose-1,5-bisphosphate-limited photosynthesis. Net photosynthetic CO₂ uptake was measured for leaves in situ within the canopy. Leaf contents of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco), light-harvesting-complex (LHC) proteins, and total N were determined. Elevated CO₂ did not affect carboxylation capacity in the most recently expanded leaves but led to a decrease in lower, shaded leaves during grain development. Despite this acclimation, in situ photosynthetic CO₂ uptake remained higher under elevated CO₂. Acclimation at elevated CO₂ was accompanied by decreases in both Rubisco and total leaf N contents and an increase in LHC content. Elevated CO₂ led to a larger increase in LHC/Rubisco in lower canopy leaves than in the uppermost leaf. Acclimation of leaf photosynthesis to elevated CO₂ therefore depended on both vertical position within the canopy and the developmental stage.     

Leishmania major Glycosylation Mutants Require Phosphoglycans (lpg2 ?) but Not Lipophosphoglycan (lpg1 ?) for Survival in Permissive Sand Fly Vectors

Background

Sand fly species able to support the survival of the protozoan parasite Leishmania have been classified as permissive or specific, based upon their ability to support a wide or limited range of strains and/or species. Studies of a limited number of fly/parasite species combinations have implicated parasite surface molecules in this process and here we provide further evidence in support of this proposal. We investigated the role of lipophosphoglycan (LPG) and other phosphoglycans (PGs) in sand fly survival, using Leishmania major mutants deficient in LPG (lpg1 ⁻), and the phosphoglycan (PG)-deficient mutant lpg2 ⁻. The sand fly species used were the permissive species Phlebotomus perniciosus and P. argentipes, and the specific vector P. duboscqi, a species resistant to L. infantum development.

Principal Findings

The lpg2 ⁻ mutants did not survive well in any of the three sand fly species, suggesting that phosphoglycans and/or other LPG2-dependent molecules are required for parasite development. In vitro, all three L. major lines were equally resistant to proteolytic activity of bovine trypsin, suggesting that sand fly-specific hydrolytic proteases or other factors are the reason for the early lpg2 ⁻ parasite killing. The lpg1 ⁻ mutants developed late-stage infections in two permissive species, P. perniciosus and P. argentipes, where their infection rates and intensities of infections were comparable to the wild type (WT) parasites. In contrast, in P. duboscqi the lpg1 ⁻ mutants developed significantly worse than the WT parasites.

Conclusions

In combination with previous studies, the data establish clearly that LPG is not required for Leishmania survival in permissive species P. perniciosus and P. argentipes but plays an important role in the specific vector P. duboscqi. With regard to PGs other than LPG, the data prove the importance of LPG2-related molecules for survival of L. major in the three sand fly species tested.     

Polo-like Kinase 2 (PLK2) Phosphorylates ��-Synuclein at Serine 129 in Central Nervous System

Several neurological diseases, including Parkinson disease and dementia with Lewy bodies, are characterized by the accumulation of α-synuclein phosphorylated at Ser-129 (p-Ser-129). The kinase or kinases responsible for this phosphorylation have been the subject of intense investigation. Here we submit evidence that polo-like kinase 2 (PLK2, also known as serum-inducible kinase or SNK) is a principle contributor to α-synuclein phosphorylation at Ser-129 in neurons. PLK2 directly phosphorylates α-synuclein at Ser-129 in an in vitro biochemical assay. Inhibitors of PLK kinases inhibited α-synuclein phosphorylation both in primary cortical cell cultures and in mouse brain in vivo. Finally, specific knockdown of PLK2 expression by transduction with short hairpin RNA constructs or by knock-out of the plk2 gene reduced p-Ser-129 levels. These results indicate that PLK2 plays a critical role in α-synuclein phosphorylation in central nervous system.The importance of α-synuclein to the pathogenesis of Parkinson disease (PD)⁴ and the related disorder, dementia with Lewy bodies (DLB), is suggested by its association with Lewy bodies and Lewy neurites, the inclusions that characterize these diseases (1–3), and demonstrated by the existence of mutations that cause syndromes mimicking sporadic PD and DLB (4–6). Furthermore, three separate mutations cause early onset forms of PD and DLB. It is particularly telling that duplications or triplications of the gene (7–9), which increase levels of α-synuclein with no alteration in sequence, also cause PD or DLB.α-Synuclein has been reported to be phosphorylated on serine residues, at Ser-87 and Ser-129 (10), although to date only the Ser-129 phosphorylation has been identified in the central nervous system (11, 12). Phosphorylation at tyrosine residues has been observed by some investigators (13, 14) but not by others (10–12). Phosphorylation at Ser-129 (p-Ser-129) is of particular interest because the majority of synuclein in Lewy bodies contains this modification (15). In addition, p-Ser-129 was found to be the most extensive and consistent modification in a survey of synuclein in Lewy bodies (11). Results have been mixed from studies investigating the function of phosphorylation using S129A and S129D mutations to respectively block and mimic the modification. Although the phosphorylation mimic was associated with pathology in studies in Drosophila (16) and in transgenic mouse models (17, 18), studies using adeno-associated virus vectors to overexpress α-synuclein in rat substantia nigra found an exacerbation of pathology with the S129A mutation, whereas the S129D mutation was benign, if not protective (19). Interpretation of these studies is complicated by a recent study showing that the S129D and S129A mutations themselves have effects on the aggregation properties of α-synuclein independent of their effects on phosphorylation, with the S129A mutation stimulating fibril formation (20). Clearly, determination of the role of p-Ser-129 phosphorylation would be helped by identification of the responsible kinase. In addition, identification will provide a pathologically relevant way to increase phosphorylation in a cell or animal model.Several kinases have been proposed to phosphorylate α-synuclein, including casein kinases 1 and 2 (10, 12, 21) and members of the G-protein-coupled receptor kinase family (22). In this report, we offer evidence that a member of the polo-like kinase (PLK) family, PLK2 (or serum-inducible kinase, SNK), functions as an α-synuclein kinase. The ability of PLK2 to directly phosphorylate α-synuclein at Ser-129 is established by overexpression in cell culture and by in vitro reaction with the purified kinase. We show that PLK2 phosphorylates α-synuclein in cells, including primary neuronal cultures, using a series of kinase inhibitors as well as inhibition of expression with RNA interference. In addition, inhibitor and knock-out studies in mouse brain support a role for PLK2 as an α-synuclein kinase in vivo.     

Identifying the African Wintering Grounds of Hybrid Flycatchers Using a Multi–Isotope (δ 2H, δ 13C, δ 15N) Assignment Approach

Migratory routes and wintering grounds can have important fitness consequences, which can lead to divergent selection on populations or taxa differing in their migratory itinerary. Collared (Ficedula albicollis) and pied (F. hypoleuca) flycatchers breeding in Europe and wintering in different sub-Saharan regions have distinct migratory routes on the eastern and western sides of the Sahara desert, respectively. In an earlier paper, we showed that hybrids of the two species did not incur reduced winter survival, which would be expected if their migration strategy had been a mix of the parent species'' strategies potentially resulting in an intermediate route crossing the Sahara desert to different wintering grounds. Previously, we compared isotope ratios and found no significant difference in stable-nitrogen isotope ratios (δ ¹⁵N) in winter-grown feathers between the parental species and hybrids, but stable-carbon isotope ratios (δ ¹³C) in hybrids significantly clustered only with those of pied flycatchers. We followed up on these findings and additionally analyzed the same feathers for stable-hydrogen isotope ratios (δ ²H) and conducted spatially explicit multi-isotope assignment analyses. The assignment results overlapped with presumed wintering ranges of the two species, highlighting the efficacy of the method. In contrast to earlier findings, hybrids clustered with both parental species, though most strongly with pied flycatcher.     

In Silico Modeling of Human α2C-Adrenoreceptor Interaction with Filamin-2

Vascular smooth muscle α_2C-adrenoceptors (α_2C-ARs) mediate vasoconstriction of small blood vessels, especially arterioles. Studies of endogenous receptors in human arteriolar smooth muscle cells (referred to as microVSM) and transiently transfected receptors in heterologous HEK293 cells show that the α_2C-ARs are perinuclear receptors that translocate to the cell surface under cellular stress and elicit a biological response. Recent studies in microVSM unraveled a crucial role of Rap1A-Rho-ROCK-F-actin pathways in receptor translocation, and identified protein-protein interaction of α_2C-ARs with the actin binding protein filamin-2 as an essential step in the process. To better understand the molecular nature and specificity of this interaction, in this study, we constructed comparative models of human α_2C-AR and human filamin-2 proteins. Finally, we performed in silico protein-protein docking to provide a structural platform for the investigation of human α_2C-AR and filamin-2 interactions. We found that electrostatic interactions seem to play a key role in this complex formation which manifests in interactions between the C-terminal arginines of α_2C-ARs (particularly R454 and R456) and negatively charged residues from filamin-2 region between residues 1979 and 2206. Phylogenetic and sequence analysis showed that these interactions have evolved in warm-blooded animals.     

Pen2 and Presenilin-1 Modulate the Dynamic Equilibrium of Presenilin-1 and Presenilin-2 ��-Secretase Complexes

γ-Secretase is known to play a pivotal role in the pathogenesis of Alzheimer disease through production of amyloidogenic Aβ42 peptides. Early onset familial Alzheimer disease mutations in presenilin (PS), the catalytic core of γ-secretase, invariably increase the Aβ42:Aβ40 ratio. However, the mechanism by which these mutations affect γ-secretase complex formation and cleavage specificity is poorly understood. We show that our in vitro assay system recapitulates the effect of PS1 mutations on the Aβ42:Aβ40 ratio observed in cell and animal models. We have developed a series of small molecule affinity probes that allow us to characterize active γ-secretase complexes. Furthermore we reveal that the equilibrium of PS1- and PS2-containing active complexes is dynamic and altered by overexpression of Pen2 or PS1 mutants and that formation of PS2 complexes is positively correlated with increased Aβ42:Aβ40 ratios. These data suggest that perturbations to γ-secretase complex equilibrium can have a profound effect on enzyme activity and that increased PS2 complexes along with mutated PS1 complexes contribute to an increased Aβ42:Aβ40 ratio.β-Amyloid (Aβ)⁵ peptides are believed to play a causative role in Alzheimer disease (AD). Aβ peptides are generated from the processing of the amyloid precursor protein (APP) by two proteases, β-secretase and γ-secretase. Although γ-secretase generates heterogenous Aβ peptides ranging from 37 to 46 amino acids in length, significant work has focused mainly on the Aβ40 and Aβ42 peptides that are the major constituents of amyloid plaques. γ-Secretase is a multisubunit membrane aspartyl protease comprised of at least four known subunits: presenilin (PS), nicastrin (Nct), anterior pharynx-defective (Aph), and presenilin enhancer 2 (Pen2). Presenilin is thought to contain the catalytic core of the complex (1–4), whereas Aph and Nct play critical roles in the assembly, trafficking, and stability of γ-secretase as well as substrate recognition (5, 6). Lastly Pen2 facilitates the endoproteolysis of PS into its N-terminal (NTF) and C-terminal (CTF) fragments thereby yielding a catalytically competent enzyme (5, 7–10). All four proteins (PS, Nct, Aph1, and Pen2) are obligatory for γ-secretase activity in cell and animal models (11, 12). There are two homologs of PS, PS1 and PS2, and three isoforms of Aph1, Aph1aS, Aph1aL, and Aph1b. At least six active γ-secretase complexes have been reported (two presenilins × three Aph1s) (13, 14). The sum of apparent molecular masses of the four proteins (PS1-NTF/CTF ≈ 53 kDa, Nct ≈ 120 kDa, Aph1 ≈ 30 kDa, and Pen2 ≈ 10kDa) is ∼200 kDa. However, active γ-secretase complexes of varying sizes, ranging from 250 to 2000 kDa, have been reported (15–19). Recently a study suggested that the γ-secretase complex contains only one of each subunit (20). Collectively these studies suggest that a four-protein complex around 200–250 kDa may be the minimal functional γ-secretase unit with additional cofactors and/or varying stoichiometry of subunits existing in the high molecular weight γ-secretase complexes. CD147 and TMP21 have been found to be associated with the γ-secretase complex (21, 22); however, their role in the regulation of γ-secretase has been controversial (23, 24).Mutations of PS1 or PS2 are associated with familial early onset AD (FAD), although it is debatable whether these familial PS mutations act as “gain or loss of function” alterations in regard to γ-secretase activity (25–27). Regardless the overall outcome of these mutations is an increased ratio of Aβ42:Aβ40. Clearly these mutations differentially affect γ-secretase activity for the production of Aβ40 and Aβ42. Despite intensive studies of Aβ peptides and γ-secretase, the molecular mechanism controlling the specificity of γ-secretase activity for Aβ40 and Aβ42 production has not been resolved. It has been found that PS1 mutations affect the formation of γ-secretase complexes (28). However, the precise mechanism by which individual subunits alter the dynamics of γ-secretase complex formation and activity is largely unresolved. A better mechanistic understanding of γ-secretase activity associated with FAD mutations has been hindered by the lack of suitable assays and probes that are necessary to recapitulate the effect of these mutations seen in cell models and to characterize the active γ-secretase complex.In our present studies, we have determined the overall effect of Pen2 and PS1 expression on the dynamics of PS1- and PS2-containing complexes and their association with γ-secretase activity. Using newly developed biotinylated small molecular probes and activity assays, we revealed that expression of Pen2 or PS1 FAD mutants markedly shifts the equilibrium of PS1-containing active complexes to that of PS2-containing complexes and results in an overall increase in the Aβ42:Aβ40 ratio in both stable cell lines and animal models. Our studies indicate that perturbations to the equilibrium of active γ-secretase complexes by an individual subunit can greatly affect the activity of the enzyme. Moreover they serve as further evidence that there are multiple and distinct γ-secretase complexes that can exist within the same cells and that their equilibrium is dynamic. Additionally the affinity probes developed here will facilitate further study of the expression and composition of endogenous active γ-secretase from a variety of model systems.     

表观遗传学——理论·方法·研究进展(2)

4 表观遗传学重点内容之二--染色体重塑 基因的活化和转录需要一系列重要的染色质变化,如染色质去凝集、核小体变成疏松的开放式结构等,使转录因子等调节因子更易接近和结合核小体DNA.与基因表达调节所伴随的这类染色质结构改变现象就叫做染色质重塑(chromatin remodeling),需要特异的因子结合在启动子和增强子等顺式作用元件上,并和RNA聚合酶之问建立起有效的联系.染色质重塑的机理涉及特殊的蛋白复合体.     

Trends in 1-year survival of people admitted to hospital in Ontario, 1994–2009

Background:

Changes in the long-term survival of people admitted to hospital is unknown. This study examined trends in 1-year survival of patients admitted to hospital adjusted for improved survival in the general population.

Methods:

One-year survival after admission to hospital was determined for all adults admitted to hospital in Ontario in 1994, 1999, 2004, or 2009 by linking to vital statistics datasets. Annual survival in the general population was determined from life tables for Ontario.

Results:

Between 1994 and 2009, hospital use decreased (from 8.8% to 6.3% of the general adult population per year), whereas crude 1-year mortality among people with hospital admissions increased (from 9.2% to 11.6%). During this time, patients in hospital became significantly older (median age increased from 51 to 58 yr) and sicker (the proportion with a Charlson comorbidity index score of 0 decreased from 68.2% to 60.0%), and were more acutely ill on admission (elective admissions decreased from 47.4% to 42.0%; proportion brought to hospital by ambulance increased from 16.1% to 24.8%). Compared with 1994, the adjusted odds ratio (OR) for death at 1 year in 2009 was 0.78 (95% confidence interval [CI] 0.77–0.79). However, 1-year risk of death in the general population decreased by 24% during the same time. After adjusting for improved survival in the general population, risk of death at 1 year for people admitted to hospital remained significantly lower in 2009 than in 1994 (adjusted relative excess risk 0.81, 95% CI 0.80–0.82).

Interpretation:

After accounting for both the increased burden of patient sickness and improved survival in the general population, 1-year survival for people admitted to hospital increased significantly from 1994 to 2009. The reasons for this improvement cannot be determined from these data. Hospitals have a special place in most health care systems. Hospital staff care for the people with the most serious illnesses and the most vulnerable. They are frequently the location of many life-defining moments — including birth, surgery, acute medical illness and death — of many people and their families. Hospitals serve as a focus in the training of most physicians. In addition, they consume a considerable proportion of health care expenditures worldwide. ¹ Given the prominence of hospitals in health care systems, measuring outcomes related to hospital care is important. In particular, the measurement of trends for outcomes of hospital care can help us to infer whether the care provided to hospital patients is improving. Previous such studies have focused on survival trends for specific diseases or patients who received treatment in specific departments. ^{2 – 12} None of these studies have adjusted for survival trends in the general population, the adjustment for which is important to determine whether changes in survival of patients in hospital merely reflect changes in the overall population. In this study, whether or not patient outcomes have changed over time was determined by examining trends in 1-year survival in all patients admitted to hospital, adjusting for improved survival in the general population.     

Bellisotoma,a new genus of Isotomidae from North?America (Hexapoda,Collembola)

A new genus of Isotomidae, Bellisotoma gen. n., is described. The new genus is a member of the Proisotoma genus complex and is characterized by a combination of having a bidentate mucro with wide dorsal lamellae that join clearly before the end of mucronal axis without forming a tooth and one strong ventral rib with basal notch that articulates with dens; having abundant chaetotaxy on both faces of dens; and abundant tergal sensilla. Bellisotoma gen. n. shows a furcula adapted to a neustonic mode of life, and may be a Isotopenola-like derivative adapted to neustonic habitats. Subisotoma joycei Soto-Adames & Giordano, 2011 and Ballistura ewingi James, 1933 are transferred to the new genus.