Structure and receptor complexes of the hemagglutinin from a highly pathogenic H7N7 influenza virus

Recurrence of highly pathogenic avian influenza (HPAI) virus subtype H7 in poultry continues to be a public health concern. In 2003, an HPAI H7N7 outbreak in The Netherlands infected 89 people in close contact with affected poultry and resulted in one fatal case. In previous studies, the virus isolated from this fatal case, A/Netherlands/219/2003 (NL219) caused a lethal infection in mouse models and had increased replication efficiency and a broader tissue distribution than nonlethal isolates from the same outbreak. A mutation which introduces a potential glycosylation site at Asn123 in the NL219 hemagglutinin was postulated to contribute to the pathogenic properties of this virus. To study this further, we have expressed the NL219 hemagglutinin in a baculovirus expression system and performed a structural analysis of the hemagglutinin in complex with avian and human receptor analogs. Glycan microarray and kinetic analysis were performed to compare the receptor binding profile of the wild-type recombinant NL219 HA to a variant with a threonine-to-alanine mutation at position 125, resulting in loss of the glycosylation site at Asn123. The results suggest that the additional glycosylation sequon increases binding affinity to avian-type α2-3-linked sialosides rather than switching to a human-like receptor specificity and highlight the mechanistic diversity of these pathogens, which calls attention to the need for further studies to fully understand the unique properties of these viruses.     

The effects of growth hormone treatment on health-related quality of life in children

BACKGROUND/AIMS: The effects of growth hormone deficiency (GHD) on linear growth in children are well documented, but there is less convincing evidence regarding the impact on health-related quality of life (QOL). We examined QOL in children aged 8-16 years with acquired GHD following treatment for malignancy (AGHD) or idiopathic GHD (IGHD) on commencing growth hormone treatment (GHT) over 6 months. We adopted a longitudinal design involving consecutive patients and their families attending clinic over an 18-month period. Mothers and children were invited to complete questionnaires before GHT (T1) and 6 months later (T2). METHODS: Mothers of 22 children (AGHD n = 14; IGHD n = 8) completed standardized measures of child QOL and behaviour. Children completed parallel measures of QOL, short-term memory tasks and fitness either in clinic or at the family home. RESULTS: For children with AGHD, QOL was significantly below population norms at T1 and improved over time. For children diagnosed with IGHD, QOL at T1 was below, but comparable with population norms. QOL improved over time, though not significantly. CONCLUSION: GHT is potentially valuable for improving QOL in children, especially in cases of AGHD. We conclude that benefits of GHT for QOL need to be evaluated independent of different diagnostic groups.     

Daily activities and sleep quality in college students

There is growing evidence that social rhythms (e.g., daily activities such as getting into or out of bed, eating, and adhering to a work schedule) have important implications for sleep. The present study used a prospective measure of daily activities to assess the relation between sleep and social rhythms. College students (n=243) 18 to 39 yrs of age, completed the Social Rhythm Metric (SRM) each day for 14 d and then completed the Pittsburgh Sleep Quality Index (PSQI). The sample was divided into groups of good or poor sleepers, according to a PSQI cut-off score of 5 points and was compared on the regularity, frequency, timing, and extent of social engagement during activities. There was a lower frequency and less regularity of social rhythms in poor sleepers relative to good sleepers. Good sleepers engaged more regularly in activities with active social engagement. Earlier rise time, first consumption of a beverage, going outdoors for the first time, and bedtime were associated with better sleep. Greater variability in rise time, consuming a morning beverage, returning home for the last time, and bedtime were associated with more disturbed sleep. The results are consistent with previous findings of reduced regularity in bedtime and rise time schedules in undergraduates, other age groups, and in clinical populations. Results augment the current thought that regulating behavioral zeitgebers may be important in influencing bed and rise times, and suggest that engaging in activities with other people may increase regularity.     

Vps9 domain-containing proteins: activators of Rab5 GTPases from yeast to neurons

Endocytosis of cell surface receptors plays an important role in regulating cell signaling cascades. In some cases, internalization of an activated receptor attenuates the signaling process, while in other cases the clustering of activated receptors on early endosomal structures has been proposed to be essential for fully activating signaling cascades. Regulating the movement of receptors and other signaling proteins through the endocytic pathway, therefore, has a direct impact on cellular homeostasis. The small GTPase Rab5 is a crucial regulatory component of the endocytic pathway. Activation of Rab5 is mediated by GDP-GTP exchange factors (GEFs) that generate the Rab5-GTP complex. A large number of proteins have been identified that contain a specific, highly conserved domain (Vps9) that catalyzes nucleotide exchange on Rab5, linking the regulation of cell signaling cascades with intracellular receptor trafficking through the endocytic pathway.     

Biochemical characterization,localization, and tissue distribution of the longer form of mouse SIRT3

SIRT3 is a key mitochondrial protein deacetylase proposed to play key roles in regulating mitochondrial metabolism but there has been considerable debate about its actual size, the sequences required for activity, and its subcellular localization. A previously cloned mouse SIRT3 has high sequence similarity with the C‐terminus of human SIRT3 but lacks an N‐terminal mitochondrial targeting sequence and has no detectable deacetylation activity in vitro. Using 5′ rapid amplification of cDNA ends, we cloned the entire sequence of mouse SIRT3, as well as rat and rabbit SIRT3. Importantly, we find that full‐length SIRT3 protein localizes exclusively to the mitochondria, in contrast to reports of SIRT3 localization to the nucleus. We demonstrate that SIRT3 has no deacetylation activity in vitro unless the protein is truncated, consistent with human SIRT3. In addition, we determined the inhibition constants and mechanism of action for nicotinamide and a small molecule SIRT3 inhibitor against active mouse SIRT3 and show that the mechanisms are different for the two compounds with respect to peptide substrate and NAD⁺. Thus, identification and characterization of the actual SIRT3 sequence should help resolve the debate about the nature of mouse SIRT3 and identify new mechanisms to modulate enzymatic activity.     

Discovery of oxazolo[4,5-b]pyridines and related heterocyclic analogs as novel SIRT1 activators

SIRT1 is an NAD⁺-dependent protein deacetylase that appears to produce beneficial effects on metabolic parameters such as glucose and insulin homeostasis. Activation of SIRT1 by resveratrol (1) has been shown to modulate insulin resistance, increase mitochondrial content and prolong survival in lower organisms and in mice on a high fat diet. Herein, we describe the identification and SAR of a series of oxazolo[4,5-b]pyridines as novel small molecule activators of SIRT1 which are structurally unrelated to and more potent than resveratrol.     

Ancient Leishmaniasis in a Highland Desert of Northern Chile

Background

Leishmaniasis is an infectious disease endemic today in many areas of South America.

Methodology

We discovered morphologic and molecular evidence of ancient infections in 4 female skulls in the archaeological cemetery of Coyo Oriente, in the desert of San Pedro de Atacama, Northern Chile. The boney facial lesions visible in the skulls could have been caused by a number of chronic infections including chronic Leishmaniasis. This diagnosis was confirmed using PCR-sequenced analyses of bone fragments from the skulls of the affected individuals.Leishmaniasis is not normally found in the high-altitude desert of Northern Chile; where the harsh climate does not allow the parasite to complete its life cycle. The presence of Leishmaniasis in ancient skulls from the region implies infection by the protozoan in an endemic area–likely, in our subjects, to have been the lowlands of North-Eastern Argentina or in Southern Bolivia.

Conclusions

We propose that the presence of the disease in ancient times in the high altitude desert of San Pedro de Atacama is the result of an exogamic system of patrilocal marriages, where women from different cultures followed their husbands to their ancestral homes, allowing immigrant women, infected early in life, to be incorporated in the Atacama desert society before they became disfigured by the disease. The present globalization of goods and services and the extraordinary facile movement of people across borders and continents have lead to a resurgence of infectious diseases and re-emergence of infections such as Leishmaniasis. We show here that such factors were already present millennia ago, shaping demographic trends and the epidemiology of infections just as they do today.     

The Epithelial Cell Adhesion Molecule EpCAM Is Required for Epithelial Morphogenesis and Integrity during Zebrafish Epiboly and Skin Development

The aberrant expression of the transmembrane protein EpCAM is associated with tumor progression, affecting different cellular processes such as cell–cell adhesion, migration, proliferation, differentiation, signaling, and invasion. However, the in vivo function of EpCAM still remains elusive due to the lack of genetic loss-of-function studies. Here, we describe epcam (tacstd) null mutants in zebrafish. Maternal-zygotic mutants display compromised basal protrusive activity and epithelial morphogenesis in cells of the enveloping layer (EVL) during epiboly. In partial redundancy with E-cadherin (Ecad), EpCAM made by EVL cells is further required for cell–cell adhesion within the EVL and, possibly, for proper attachment of underlying deep cells to the inner surface of the EVL, thereby also affecting deep cell epiboly movements. During later development, EpCAM per se becomes indispensable for epithelial integrity within the periderm of the skin, secondarily leading to disrupted morphology of the underlying basal epidermis and moderate hyper-proliferation of skin cells. On the molecular level, EVL cells of epcam mutant embryos display reduced levels of membranous Ecad, accompanied by an enrichment of tight junction proteins and a basal extension of apical junction complexes (AJCs). Our data suggest that EpCAM acts as a partner of E-cadherin to control adhesiveness and integrity as well as plasticity and morphogenesis within simple epithelia. In addition, EpCAM is required for the interaction of the epithelia with underlying cell layers.     

Blood Levels of Carbonic Anhydrase 9 Correlate with Clear Cell Renal Cell Carcinoma Activity

Introduction  Biomarkers for early detection of renal cell carcinoma (RCC) may help diagnose minimal residual disease in patients at risk for RCC, can guide anti-angiogenic therapy, or may help identify candidates for adjuvant treatment. In this study, we investigated whether blood levels of carbonic anhydrase 9 (CA9) correlate with RCC tumor burden and therefore disease activity. Methods  CA9 is a von Hippel–Lindau–hypoxia inducible factor target upregulated in clear cell RCC. We used an anti-CA9 antibody (M75)-based enzyme-linked immunosorbent assay test to measure CA9 levels in blood obtained before and after nephrectomy for clinically localized disease in patients with: (1) clear cell RCC, (2) papillary and chromophobe RCC or oncocytoma, or (3) benign kidney lesions, and we compared these samples to blood drawn from normal control individuals. Results  We observed a significant (p < 0.006) decrease in the blood levels of CA9, after nephrectomy for localized disease, in the majority of patients with clear cell RCC (57%). In contrast, patients with nonclear cell RCC, benign disease, or those having undergone debulking nephrectomy for metastatic disease did not have a decrease in CA9 blood levels after nephrectomy. Preliminary longitudinal follow up measurements of CA9 levels in a small group of patients indicated that rising CA9 levels may correlate with disease progression. Conclusions  Plasma CA9 levels correlate with disease activity in a subset of clear cell RCC patients and should be considered in future multiplex RCC biomarker development algorithms.     

ROLE OF NUTRIENT FLUCTUATIONS AND DELAYED DEVELOPMENT IN GAMETOPHYTE REPRODUCTION BY MACROCYSTIS PYRIFERA (PHAEOPHYCEAE) IN SOUTHERN CALIFORNIA1

Organisms occurring in environments subject to severe disturbance and/or periods of poor environmental quality that result in severe adult mortality can survive these periods by relying on alternate life stages that delay their development in a resistant state until conditions improve. In the northeast Pacific, the forest‐forming giant kelp Macrocystis pyrifera (L.) C. Agardh periodically experiences widespread adult mortality during extended periods of extremely low nutrients and high temperatures, such as those associated with El Niño. Recovery following these periods is hypothesized to occur from microscopic life stages that delay their development until the return of favorable conditions. In the laboratory, we experimentally examined the environmental conditions responsible for regulating delayed development of the microscopic stages of M. pyrifera from Southern California, USA. Nutrients controlled the delay and resumption of gametophyte growth and reproduction, perhaps linked to the large fluctuations in nutrients occurring seasonally and interannually in this region. Although growth of gametophytes proceeded in the virtual absence of nitrate, both nitrate and other trace nutrients were necessary for gametogenesis. Upon exposure to elevated nutrients, delayed gametophytes produced sporophytes more quickly (5–20 d) and at smaller sizes (10–200 μm) than gametophytes that had never been delayed (18–80 d, 80–400 μm, respectively), reducing negative density‐dependent effects. This finding demonstrates that delayed gametophytes of M. pyrifera rapidly utilize increased resources to consistently produce sporophytes. Further work is needed to assess their potential role in population recovery following periods of poor environmental quality.