Oxidative stresses induced by glycoxidized human or bovine serum albumin on human monocytes

Oxidative stress and protein modifications are frequently observed in numerous disease states. Albumin, the major circulating protein in blood, can undergo increased glycoxidation in diabetes. Protein glycoxidation can lead to the formation of advanced glycoxidation end products, which induce various deleterious effects on cells. Herein, we report the effect of glucose or methylglyoxal-induced oxidative modifications on BSA or HSA protein structures and on THP1 monocyte physiology. The occurrence of oxidative modifications was found to be enhanced in glycoxidized BSA and HSA, after determination of their free thiol group content, relative electrophoretic migration, carbonyl content, and antioxidant activities. Cells treated with glycoxidized albumin exhibited an overgeneration of intracellular reactive oxygen species, impairments in proteasomal activities, enhancements in RAGE expression, and an accumulation of carbonylated proteins. These novel observations made in the presence of a range of modified BSA and HSA facilitate the comparison of the glycoxidation extent of albumin with the oxidative stress induced in cultured monocytes. Finally, this study reconfirms the influence of experimental conditions in which AGEs are generated and the concentration levels in experiments designed to mimic pathological conditions.     

Suppression of a Natural Killer Cell Response by Simian Immunodeficiency Virus Peptides

Natural killer (NK) cell responses in primates are regulated in part through interactions between two highly polymorphic molecules, the killer-cell immunoglobulin-like receptors (KIRs) on NK cells and their major histocompatibility complex (MHC) class I ligands on target cells. We previously reported that the binding of a common MHC class I molecule in the rhesus macaque, Mamu-A1*002, to the inhibitory receptor Mamu-KIR3DL05 is stabilized by certain simian immunodeficiency virus (SIV) peptides, but not by others. Here we investigated the functional implications of these interactions by testing SIV peptides bound by Mamu-A1*002 for the ability to modulate Mamu-KIR3DL05⁺ NK cell responses. Twenty-eight of 75 SIV peptides bound by Mamu-A1*002 suppressed the cytolytic activity of primary Mamu-KIR3DL05⁺ NK cells, including three immunodominant CD8⁺ T cell epitopes previously shown to stabilize Mamu-A1*002 tetramer binding to Mamu-KIR3DL05. Substitutions at C-terminal positions changed inhibitory peptides into disinhibitory peptides, and vice versa, without altering binding to Mamu-A1*002. The functional effects of these peptide variants on NK cell responses also corresponded to their effects on Mamu-A1*002 tetramer binding to Mamu-KIR3DL05. In assays with mixtures of inhibitory and disinhibitory peptides, low concentrations of inhibitory peptides dominated to suppress NK cell responses. Consistent with the inhibition of Mamu-KIR3DL05⁺ NK cells by viral epitopes presented by Mamu-A1*002, SIV replication was significantly higher in Mamu-A1*002⁺ CD4⁺ lymphocytes co-cultured with Mamu-KIR3DL05⁺ NK cells than with Mamu-KIR3DL05^- NK cells. These results demonstrate that viral peptides can differentially affect NK cell responses by modulating MHC class I interactions with inhibitory KIRs, and provide a mechanism by which immunodeficiency viruses may evade NK cell responses.     

Internalization of staphylococcal leukotoxins that bind and divert the C5a receptor is required for intracellular Ca2+ mobilization by human neutrophils

A growing number of receptors, often associated with the innate immune response, are being identified as targets for bacterial toxins of the beta‐stranded pore‐forming family. These findings raise the new question of whether the receptors are activated or merely used as docking points facilitating the formation of a pore. To elucidate whether the Staphylococcus aureus Panton‐Valentine leukocidin and the leukotoxin HlgC/HlgB act through the C5a receptor (C5aR) as agonists, antagonists or differ from the C5a complement‐derived peptide, their activity is explored on C5aR‐expressing cells. Both leukotoxins equally bound C5aR in neutrophils and in stable transfected U937 cells and initiated mobilization of intracellular Ca²⁺. HlgC/HlgB requires the presence of robust intracellular acidic Ca²⁺ stores in order to evoke a rise in free [Ca²⁺]_i, while the LukS‐PV/LukF‐PV directly altered reticular Ca²⁺ stores. Intracellular target specificity is conferred by the F‐subunit associated to the S‐subunit binding the receptor. Furthermore, internalization of the two leukotoxin components (S‐ and F‐subunits) associated to C5aR is required for the initiation of [Ca²⁺]_i mobilization. Electrophysiological recordings on living cells demonstrated that LukS‐PV/LukF‐PV does not alter the membrane resistance of C5aR‐expressing cells. The present observations suggest that part of the pore‐forming process occurs in distinct intracellular compartments rather than at the plasma membrane.     

Traditional and Emerging Lifestyle Risk Behaviors and All-Cause Mortality in Middle-Aged and Older Adults: Evidence from a Large Population-Based Australian Cohort

Background

Lifestyle risk behaviors are responsible for a large proportion of disease burden worldwide. Behavioral risk factors, such as smoking, poor diet, and physical inactivity, tend to cluster within populations and may have synergistic effects on health. As evidence continues to accumulate on emerging lifestyle risk factors, such as prolonged sitting and unhealthy sleep patterns, incorporating these new risk factors will provide clinically relevant information on combinations of lifestyle risk factors.

Methods and Findings

Using data from a large Australian cohort of middle-aged and older adults, this is the first study to our knowledge to examine a lifestyle risk index incorporating sedentary behavior and sleep in relation to all-cause mortality. Baseline data (February 2006– April 2009) were linked to mortality registration data until June 15, 2014. Smoking, high alcohol intake, poor diet, physical inactivity, prolonged sitting, and unhealthy (short/long) sleep duration were measured by questionnaires and summed into an index score. Cox proportional hazards analysis was used with the index score and each unique risk combination as exposure variables, adjusted for socio-demographic characteristics.During 6 y of follow-up of 231,048 participants for 1,409,591 person-years, 15,635 deaths were registered. Of all participants, 31.2%, 36.9%, 21.4%, and 10.6% reported 0, 1, 2, and 3+ risk factors, respectively. There was a strong relationship between the lifestyle risk index score and all-cause mortality. The index score had good predictive validity (c index = 0.763), and the partial population attributable risk was 31.3%. Out of all 96 possible risk combinations, the 30 most commonly occurring combinations accounted for more than 90% of the participants. Among those, combinations involving physical inactivity, prolonged sitting, and/or long sleep duration and combinations involving smoking and high alcohol intake had the strongest associations with all-cause mortality. Limitations of the study include self-reported and under-specified measures, dichotomized risk scores, lack of long-term patterns of lifestyle behaviors, and lack of cause-specific mortality data.

Conclusions

Adherence to healthy lifestyle behaviors could reduce the risk for death from all causes. Specific combinations of lifestyle risk behaviors may be more harmful than others, suggesting synergistic relationships among risk factors.     

Technical advance: a high throughput system for transposon tagging and promoter trapping in tomato

We describe new tools for functional analysis of the tomato genome based on insertional mutagenesis with the maize Ac/Ds transposable elements in the background of the miniature cultivar Micro-Tom. 2932 F3 families, in which Ds elements transposed and were stabilized, were screened for phenotypic mutations. Out of 10 families that had a clear mutant phenotype, only one mutant was Ds-tagged. In addition, we developed promoter trapping using the firefly luciferase reporter gene and enhancer trapping, using beta-glucuronidase (GUS). We show that luciferase can be used as a non-invasive reporter to identify, isolate and regenerate somatic sectors, to study the time course of mutant expression, and to identify inducible genes. Out of 108 families screened for luciferase activity 55% showed expression in the flower, 11% in the fruit and 4% in seedlings, suggesting a high rate of Ds insertion into genes. Preferential insertion into genes was supported by the analysis of Ds flanking sequences: 28 out of 50 sequenced Ds insertion sites were similar to known genes or to ESTs. In summary, the 2932 lines described here contain 2-3 Ds inserts per line, representing a collection of approximately 7500 Ds insertions. This collection has potential for use in high-throughput functional analysis of genes and promoter isolation in tomato.     

Mechanisms of sexual selection: sexual swellings and estrogen concentrations as fertility indicators and cues for male consort decisions in wild baboons

Male mate-guarding episodes ('consortships'), are taxonomically widespread, yet costly to individual males. Consequently, males should bias consortships toward females with whom the probability of conception is high. We combined data on consortships with visual scoring of sexual swellings and assays of fecal estrogen concentrations (fE) in a wild population of baboons (Papio cynocephalus) to test the hypotheses that sexual swellings are reliable indicators of (1) within-cycle timing of ovulation, (2) differences in conception probability among females that differ in maturational stage, and (3) conceptive versus non-conceptive cycles of parous females. We also evaluated whether adult males might rely on swellings or other estrogen-dependent signals (e.g., fE) for mate-guarding decisions. We found that sexual swellings reflected conception probability within and among cycles. Adult males limited their consortships to the turgescent phase of cycles, and consorted more with adult females than with newly cycling adolescents. The highest ranking (alpha) males discriminated more than did males of other ranks; they (1) limited their consortships to the 5-day peri-ovulatory period, (2) consorted more with adult than with adolescent females, and (3) consorted more with adult females on conceptive cycles than on non-conceptive cycles, all to a greater extent than did males of other ranks. Male mate choice based on sexual swellings and other estrogenic cues of fertility may result in sexual selection on these female traits and enhance dominance-based reproductive skew in males. Alpha males are the least constrained in their mating behavior and can best take advantage of these cues to mate selectively.     

The Membrane Topology of ALMT1, an Aluminum-Activated Malate Transport Protein in Wheat (Triticum aestivum)

The wheat ALMT1 gene encodes an aluminum (Al)-activated malate transport protein which confers Al-resistance. We investigated the membrane topology of this plasma-membrane localized protein with immunocytochemical techniques. Several green fluorescent protein (GFP)-fused and histidine (His)-tagged chimeras of ALMT1 were prepared based on a computer-predicted secondary structure and transiently expressed in cultured mammalian cells. Antibodies raised to polypeptide epitopes of ALMT1 were used in conjunction with the antibody to the His-tags to determine the topology of ALMT1. This study shows that the ALMT1 protein contains six transmembrane domains with the amino and carboxyl termini located on the extracellular side of the plasma membrane.Key Words: ALMT1, aluminum resistance, immunofluorescent staining, malate transporter, topology, Triticum aestivum     

Whole-genome sequences of Borrelia bissettii, Borrelia valaisiana, and Borrelia spielmanii

It has been known for decades that human Lyme disease is caused by the three spirochete species Borrelia burgdorferi, Borrelia afzelii, and Borrelia garinii. Recently, Borrelia valaisiana, Borrelia spielmanii, and Borrelia bissettii have been associated with Lyme disease. We report the complete genome sequences of B. valaisiana VS116, B. spielmanii A14S, and B. bissettii DN127.