Invasive Insects Differ from Non-Invasive in Their Thermal Requirements

We tested whether two basic thermal requirements for insect development, lower developmental thresholds, i.e. temperatures at which development ceases, and sums of effective temperatures, i.e. numbers of day degrees above the lower developmental thresholds necessary to complete development, differ among insect species that proved to be successful invaders in regions outside their native range and those that did not. Focusing on species traits underlying invasiveness that are related to temperature provides insights into the mechanisms of insect invasions. The screening of thermal requirements thus could improve risk-assessment schemes by incorporating these traits in predictions of potentially invasive insect species. We compared 100 pairs of taxonomically-related species originating from the same continent, one invasive and the other not reported as invasive. Invasive species have higher lower developmental thresholds than those never recorded outside their native ranges. Invasive species also have a lower sum of effective temperatures, though not significantly. However, the differences between invasive and non-invasive species in the two physiological measures were significantly inversely correlated. This result suggests that many species are currently prevented from invading by low temperatures in some parts of the world. Those species that will overcome current climatic constraints in regions outside their native distribution due to climate change could become even more serious future invaders than present-day species, due to their potentially faster development.     

Decline and Local Extinction of Caribbean Eusocial Shrimp

The tropical shrimp genus Synalpheus includes the only eusocial marine animals. In much of the Caribbean, eusocial species have dominated the diverse fauna of sponge-dwelling shrimp in coral rubble for at least the past two decades. Here we document a recent, dramatic decline and apparent local extinction of eusocial shrimp species on the Belize Barrier Reef. Our collections from shallow reefs in central Belize in 2012 failed to locate three of the four eusocial species formerly abundant in the area, and showed steep declines in colony size and increases in frequency of queenless colonies prior to their disappearance. Concordant with these declines, several nonsocial, pair-forming Synalpheus species increased in frequency. The decline in eusocial shrimp is explained in part by disappearance of two sponge species on which they specialize. Eusocial shrimp collections from Jamaica in 2012 showed similar patterns of decline in colony size and increased queenlessness compared with prior Jamaican collections. The decline and local extinction of eusocial shrimp happened against a backdrop of changes in coral assemblages during recent decades, and may reflect changes in abundance and quality of dead coral substratum and succession of the diverse cryptic organisms living within it. These changes document potentially worrisome declines in a unique taxon of eusocial marine animals.     

P‐cresyl sulfate causes mitochondrial hyperfusion in H9C2 cardiomyoblasts

Increased circulating level of uraemic solute p‐cresyl sulphate (PCS) in patients with chronic kidney disease (CKD) is known to increase myocardial burden relevant to mitochondrial abnormalities. This study aimed at investigating mitochondrial response to PCS in H9C2 cardiomyoblasts. H9C2 cardiomyoblasts were treated with four different concentrations of PCS (3.125, 6.25, 12.5 and 25.0 µg/mL) to study the changes in cell proliferation, cell size and mitochondrial parameters including morphology, respiration, biogenesis and membrane potential. The lowest effective dose of PCS (6.25 µg/mL) induced mitochondrial hyperfusion with enhanced mitochondrial connectivity, mitochondrial oxygen consumption rates, mitochondrial mass, mitochondrial DNA copy number and increased volume of cardiomyoblasts. After PCS treatment, phosphorylation of energy‐sensing adenosine monophosphate‐activated protein kinase (AMPK) was increased without induction of apoptosis. In contrast, mitochondrial mass was recovered after AMPK silencing. Additionally, mitochondrial hyperfusion and cell volume were reversed after cessation of PCS treatment. The results of the present study showed that low‐level PCS not only caused AMPK‐dependent mitochondrial hyperfusion but also induced cell enlargement in H9C2 cardiomyoblasts in vitro.     

Hypoxic Tumor Cell Modulates Its Microenvironment to Enhance Angiogenic and Metastatic Potential by Secretion of Proteins and Exosomes

Under hypoxia, tumor cells produce a secretion that modulates their microenvironment to facilitate tumor angiogenesis and metastasis. Here, we observed that hypoxic or reoxygenated A431 carcinoma cells exhibited enhanced angiogenic and metastatic potential such as reduced cell-cell and cell-extracellular matrix adhesion, increased invasiveness, and production of a secretion with increased chorioallantoic membrane angiogenic activity. Consistent with these observations, quantitative proteomics revealed that under hypoxia the tumor cells secreted proteins involved in angiogenesis, focal adhesion, extracellular matrix-receptor interaction, and immune cell recruitment. Unexpectedly, the secreted proteins were predominantly cytoplasmic and membrane proteins. Ultracentrifugation at 100,000 × g precipitated 54% of the secreted proteins and enriched for many exosome-associated proteins such as the tetraspanins and Alix and also proteins with the potential to facilitate angiogenesis and metastasis. Two tetraspanins, CD9 and CD81, co-immunoprecipitated. Together, these data suggested that tumor cells secrete proteins and exosomes with the potential to modulate their microenvironment and facilitate angiogenesis and metastasis.Cancer is the leading cause of mortality, causing one in eight deaths worldwide with 90% of these deaths attributable to metastases (1). Generally, a primary non-metastatic cancer begins at a localized focus and is resectable with good prognosis, but once metastasized, it is usually unresectable, and controlling its spread with radio- and chemotherapy remains ineffective (2). In fact, prognoses of highly metastatic cancers have not improved in the last century (3).To sustain growth and survival in their hostile microenvironment, rapidly growing tumors have to overcome hypoxia (Hx)¹ and a lack of nutrients through either angiogenesis to ensure an adequate supply of oxygen and nutrients or metastasis to a more conducive microenvironment. Therefore, therapeutic intervention targeting tumor angiogenesis or metastasis represents a viable strategy for regulating tumor growth. Indeed, antitumor angiogenesis drugs such as anti-VEGF therapy have proven to be clinically efficacious (4). However, the therapeutic efficacy of such treatments is generally short lived as tumors are proficient at adopting alternative pathways to circumvent the therapeutic block. For example, prolonged anti-VEGF treatment on tumors is known to select for the tumor cells that recruit alternative angiogenesis signaling pathways involving fibroblast growth factor, platelet-derived growth factor (PDGF), and angiopoietins (5). Therefore, to develop effective therapeutics, a comprehensive understanding of the complex processes that are central to metastasis and angiogenesis would likely reveal more robust and less redundant therapeutic targets. Because emerging evidence implicates Hx as a key inducer of angiogenesis and metastasis in tumors (6) and because extracellular signals emanating from the tumor cells will be necessary in modulating the extracellular matrix (ECM) to facilitate the cell migration during tumor development (7), we focused on elucidating the secretome (8) of tumor cells in their adaptation to Hx.A431 squamous carcinoma cells have been used as a model to study the oxidative stress- or EGFR-mediated angiogenesis and tumor growth (9, 10) and in a xenograft model for metastasis (11). Here, A431 cells were used to investigate the effects of Hx and hypoxia/reoxygenation (Reox) stresses on metastasis and angiogenic potential. We observed that under Hx the tumor cells exhibited reduced adhesion to their neighboring cells or ECM accompanied by enhanced invasiveness into Matrigel. We also noted that secretion from the hypoxic A431 cells was more efficient at inducing angiogenesis in the chorioallantoic membrane (CAM) assay. These observations suggest that Hx and/or Reox potentiated the angiogenic and metastatic phenotype in A431 cells, possibly through the secretion of proangiogenic and prometastatic factors. To test this hypothesis, we used mass spectrometry-based and cytokine array proteomics approaches to perform high throughput elucidation of the secretome of A431 tumor cells. High throughput proteomics analysis by mass spectrometry has been applied successfully to uncover potential cancer biomarkers as well as elucidate the tumorigenic mechanism (12, 13). Here, we utilized quantitative proteomics in analyzing the tumor secretome and delineating the dynamic changes in the secretome during Hx and Reox with a specific focus on signals that are potentially useful to the survival of a tumor in a hostile tumor microenvironment.     

Digital Microfluidics for Automated Proteomic Processing

Clinical proteomics has emerged as an important new discipline, promising the discovery of biomarkers that will be useful for early diagnosis and prognosis of disease. While clinical proteomic methods vary widely, a common characteristic is the need for (i) extraction of proteins from extremely heterogeneous fluids (i.e. serum, whole blood, etc.) and (ii) extensive biochemical processing prior to analysis. Here, we report a new digital microfluidics (DMF) based method integrating several processing steps used in clinical proteomics. This includes protein extraction, resolubilization, reduction, alkylation and enzymatic digestion. Digital microfluidics is a microscale fluid-handling technique in which nanoliter-microliter sized droplets are manipulated on an open surface. Droplets are positioned on top of an array of electrodes that are coated by a dielectric layer - when an electrical potential is applied to the droplet, charges accumulate on either side of the dielectric. The charges serve as electrostatic handles that can be used to control droplet position, and by biasing a sequence of electrodes in series, droplets can be made to dispense, move, merge, mix, and split on the surface. Therefore, DMF is a natural fit for carrying rapid, sequential, multistep, miniaturized automated biochemical assays. This represents a significant advance over conventional methods (relying on manual pipetting or robots), and has the potential to be a useful new tool in clinical proteomics.Mais J. Jebrail, Vivienne N. Luk, and Steve C. C. Shih contributed equally to this work.Sergio L. S. Freire''s current address is at the University of the Sciences in Philadelphia located at 600 South 43rd Street, Philadelphia, PA 19104.Download video file.^{(130M, mp4)}     

An E. coli lon mutant conferring partial resistance to colicin may reveal a novel role in regulating proteins involved in the translocation of colicin

Initially, we found that a lon mutant confers partial resistance against colicin. The results of Western blotting detected a decrease in the protein expression levels of BtuB and OmpF involved in colicin translocation in the lon mutant. Moreover, 2-D gel analysis revealed that the expression level of some scavenger proteins marks the lon mutant as being in a situation similar to oxidative stress. OxyRS and SoxRS are the two major response regulators for oxidative stress. Our RT-PCR analysis revealed an elevation of expression of the oxyS gene in the lon mutant. An immunoblot assay further confirmed that overexpression of oxyS RNA can negatively control on the expression of BtuB protein. Probably the BtuB is negatively regulated by a global regulator, oxyS, induced during oxidative stress.