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Supplementary Figure S1 from The Glycosphingolipid, Lactosylceramide, Regulates β1-Integrin Clustering and Endocytosis

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Glycosphingolipids are known to play roles in integrin-mediated cell adhesion and migration; however, the mechanisms by which glycosphingolipids affect integrins are unknown. Here, we show that addition of the glycosphingolipid, C8-lactosylceramide (C8-LacCer), or free cholesterol to human fibroblasts at 10°C causes the formation of glycosphingolipid-enriched plasma membrane domains as shown by visualizing a fluorescent glycosphingolipid probe, BODIPY-LacCer, incorporated into the plasma membrane of living cells. Addition of C8-LacCer or cholesterol to cells initiated the clustering of β1-integrins within these glycosphingolipid-enriched domains and the activation of the β1-integrins as assessed using a HUTS antibody that only binds activated integrin. On warming to 37°C, β1-integrins were rapidly internalized via caveolar endocytosis in cells treated with C8-LacCer or cholesterol, whereas little β1-integrin was endocytosed in untreated fibroblasts. Incubation of cells with C8-LacCer or cholesterol followed by warm-up caused src activation, a reorganization of the actin cytoskeleton, translocation of RhoA GTPase away from the plasma membrane as visualized using total internal reflection fluorescence microscopy, and transient cell detachment. These studies show that LacCer can regulate integrin function both by modulating integrin clustering in microdomains and by regulating integrin endocytosis via caveolae. Our findings suggest the possibility that aberrant levels of glycosphingolipids found in cancer cells may influence cell attachment events by direct effects on integrin clustering and internalization.

Nuanced understanding of seasonal movements of partially migratory birds is paramount to species and habitat conservation. Using nascent statistical methods, we identified migratory strategies of birds outfitted with radio-frequency identification (RFID) tags detected at RFID feeders in two sites in California, USA. We quantified proportions of migrants and residents and the seasonal phenology for each movement strategy in Allen’s and Anna’s hummingbirds; we also validated our methodology by fitting our model to obligate migratory black-chinned hummingbirds. Allen’s and Anna’s hummingbirds exhibited characteristics of facultative migratory behaviour. We also quantified apparent annual survival for each migratory strategy and found that residents had significantly higher probabilities of apparent survival. Low survival estimates for migrants suggest that a high proportion of birds in the migrant group permanently emigrated from our study sites. Considered together, our analyses suggest that hummingbirds in both northern and southern California sites partake in diverse and highly plastic migratory behaviours. Our assessment elucidates the dynamics underlying idiosyncratic migratory behaviours of two species of hummingbirds, in addition to describing a framework for similar assessments of migratory behaviours using the multi-state open robust design with state uncertainty model and single-site dynamics.

Genetic screens provide a mechanism to identify genes involved with different cellular and organismal processes. Using a Flp/FRT screen in the Drosophila eye we identified mutations that result in alterations and de-regulation of cell growth and division. From this screen a group of undergraduate researchers part of the Fly-CURE consortium mapped and characterized a new allele of the gene Hippo, HpoN.1.2.

Despite the popular use of hummingbird feeders, there are limited studies evaluating the effects of congregation, sharing food resources and increased contact when hummingbirds visit feeders in urban landscapes. To evaluate behavioral interactions occurring at feeders, we tagged 230 individuals of two species, Anna's and Allen's Hummingbirds, with passive integrated transponder tags and recorded their visits with RFID transceivers at feeders. For detecting the presence of tagged birds, we developed an RFID equipped feeding station using a commercially available antenna and RFID transceiver. Data recorded included the number of feeder visits, time spent at the feeder, simultaneous feeder visitation by different individuals, and identifying which feeders were most commonly visited by tagged birds. For the study period (September 2016 to March 2018), 118,017 detections were recorded at seven feeding stations located at three California sites. The rate of tagged birds returning to RFID equipped feeders at least once was 61.3% (141/230 birds). Females stayed at feeders longer than males per visit. We identified primary, secondary and tertiary feeders at Sites 2 and 3, according to the frequency of visitation to them, with a mean percentage of 86.9% (SD±19.13) visits to a primary feeder for each tagged hummingbird. During spring and summer, hummingbirds visited feeders most often in morning and evening hours. Feeder visits by males overlapped in time with other males more frequently than other females. The analysis of the contact network at the feeders did not distinguish any significant differences between age or sex. Although most hummingbirds visited the feeders during the daytime, our system recorded night feeder visitations (n = 7 hummingbirds) at one site. This efficient use of RFID technology to characterize feeder visitations and contact networks of hummingbirds in urban habitats could be used in the future to elucidate behaviors, population dynamics and community structure of hummingbirds visiting feeders.

Purpose The purpose of this paper is to discuss the implementation of a skills development project aimed at increasing the technology competencies of participating librarians in Columbia University’s Science & Engineering Libraries, in response to changing instructional needs. Design/methodology/approach Adopting a DIY or Maker learning model gave librarians a perfect opportunity to experience learning new technology skills just as their users encounter them. Findings The authors conclude that this collaborative project methodology could potentially fit a large variety of different library environments, providing other institutions with an excellent opportunity to reassess and revamp staff skills, no matter their instructional focus. Originality/value The literature notes that previous library staff training models were largely devised for newly matriculated Library and Information Science graduates preparing to enter the workforce. Burgeoning technology developments require libraries to explore novel methods to expose staff to new technology skills; this case study applies the programmatic lens of the Maker Movement to a collaborative staff learning model.

Glycosphingolipids are known to play roles in integrin-mediated cell adhesion and migration; however, the mechanisms by which glycosphingolipids affect integrins are unknown. Here, we show that addition of the glycosphingolipid, C8-lactosylceramide (C8-LacCer), or free cholesterol to human fibroblasts at 10°C causes the formation of glycosphingolipid-enriched plasma membrane domains as shown by visualizing a fluorescent glycosphingolipid probe, BODIPY-LacCer, incorporated into the plasma membrane of living cells. Addition of C8-LacCer or cholesterol to cells initiated the clustering of β1-integrins within these glycosphingolipid-enriched domains and the activation of the β1-integrins as assessed using a HUTS antibody that only binds activated integrin. On warming to 37°C, β1-integrins were rapidly internalized via caveolar endocytosis in cells treated with C8-LacCer or cholesterol, whereas little β1-integrin was endocytosed in untreated fibroblasts. Incubation of cells with C8-LacCer or cholesterol followed by warm-up caused src activation, a reorganization of the actin cytoskeleton, translocation of RhoA GTPase away from the plasma membrane as visualized using total internal reflection fluorescence microscopy, and transient cell detachment. These studies show that LacCer can regulate integrin function both by modulating integrin clustering in microdomains and by regulating integrin endocytosis via caveolae. Our findings suggest the possibility that aberrant levels of glycosphingolipids found in cancer cells may influence cell attachment events by direct effects on integrin clustering and internalization.

Internalization of some plasma membrane constituents, bacterial toxins, and viruses occurs via caveolae; however, the factors that regulate caveolar internalization are still unclear. Here, we demonstrate that a brief treatment of cultured cells with natural or synthetic glycosphingolipids (GSLs) or elevation of cholesterol (either by acute treatment with mβ-cyclodextrin/cholesterol or by alteration of growth conditions) dramatically stimulates caveolar endocytosis with little or no effect on other endocytic mechanisms. These treatments also stimulated the movement of GFP-labeled vesicles in cells transfected with caveolin-1-GFP and reduced the number of surface-connected caveolae seen by electron microscopy. In contrast, overexpression of caveolin-1 decreased caveolar uptake, but treatment with GSLs reversed this effect and stimulated caveolar endocytosis. Stimulation of caveolar endocytosis did not occur using ceramide or phosphatidylcholine and was not due to GSL degradation because similar results were obtained using a nonhydrolyzable GSL analog. Stimulated caveolar endocytosis required src kinase and PKC-α activity as shown by i) use of pharmacological inhibitors, ii) expression of kinase inactive src or dominant negative PKCα, and iii) stimulation of src kinase activity upon addition of GSLs or cholesterol. These results suggest that caveolar endocytosis is regulated by a balance of caveolin-1, cholesterol, and GSLs at the plasma membrane.

The use of lithium N , O -dimethylhydroxylamide as an alternative in situ protecting agent for aryl aldehydes with low ortho -directing properties has been evaluated and subsequently applied to two practical multi-step one-pot syntheses of developmental drug candidate intermediates.

Sphingolipids (SLs) are plasma membrane constituents in eukaryotic cells which play important roles in a wide variety of cellular functions. However, little is known about the mechanisms of their internalization from the plasma membrane or subsequent intracellular targeting. We have begun to study these issues in human skin fibroblasts using fluorescent SL analogues. Using selective endocytic inhibitors and dominant negative constructs of dynamin and epidermal growth factor receptor pathway substrate clone 15, we found that analogues of lactosylceramide and globoside were internalized almost exclusively by a clathrin-independent (“caveolar-like”) mechanism, whereas an analogue of sphingomyelin was taken up approximately equally by clathrin-dependent and -independent pathways. We also showed that the Golgi targeting of SL analogues internalized via the caveolar-like pathway was selectively perturbed by elevated intracellular cholesterol, demonstrating the existence of two discrete Golgi targeting pathways. Studies using SL-binding toxins internalized via clathrin-dependent or -independent mechanisms confirmed that endogenous SLs follow the same two pathways. These findings (a) provide a direct demonstration of differential SLs sorting into early endosomes in living cells, (b) provide a “vital marker” for endosomes derived from caveolar-like endocytosis, and (c) identify two independent pathways for lipid transport from the plasma membrane to the Golgi apparatus in human skin fibroblasts.

L'etude de graffiti sur les murs de Szeged en Hongrie met en evidence un mode d'expression politique et culturelle. Certains symboles ou slogans vont a l'encontre de l'influence capitaliste anglaise et revelent des ideologies extremistes. Ces expressions variees revelent les contradictions et les ambiguites a l'egard non seulement du pluralisme politique, de l'emergence de plusieurs groupes economiques mais aussi des valeurs de l'expression culturelle : alors que pour les uns la presence occidentale est une menace, pour les autres il s'agit de la liberte de s'approprier des symboles culturels