Altered Growth Rates Research Articles

PEP3 overexpression shortens lag phase but does not alter growth rate in Saccharomyces cerevisiae exposed to acetic acid stress.

In fungi, two recognized mechanisms contribute to pH homeostasis: the plasma membrane proton-pumping ATPase that exports excess protons and the vacuolar proton-pumping ATPase (V-ATPase) that mediates vacuolar proton uptake. Here, we report that overexpression of PEP3 which encodes a component of the HOPS and CORVET complexes involved in vacuolar biogenesis, shortened lag phase in Saccharomyces cerevisiae exposed to acetic acid stress. By confocal microscopy, PEP3-overexpressing cells stained with the vacuolar membrane-specific dye, FM4-64 had more fragmented vacuoles than the wild-type control. The stained overexpression mutant was also found to exhibit about 3.6-fold more FM4-64 fluorescence than the wild-type control as determined by flow cytometry. While the vacuolar pH of the wild-type strain grown in the presence of 80 mM acetic acid was significantly higher than in the absence of added acid, no significant difference was observed in vacuolar pH of the overexpression strain grown either in the presence or absence of 80 mM acetic acid. Based on an indirect growth assay, the PEP3-overexpression strain exhibited higher V-ATPase activity. We hypothesize that PEP3 overexpression provides protection from acid stress by increasing vacuolar surface area and V-ATPase activity and, hence, proton-sequestering capacity.

Indirect and direct effects of salinity on the quantity and quality of total amino acids in Ulva ohnoi (Chlorophyta).

Salinity can affect the quantity and quality of total amino acids (TAAs) in seaweeds indirectly by altering growth rates and thereby diluting or concentrating the amino acid content of the biomass, or directly by altering the synthesis of specific amino acids and osmolytes. This study attempted to partition the indirect and direct effects of salinity on the quantity and quality of TAAs in the green seaweed Ulva ohnoi by culturing it under a range of salinities without nutrient limitation. Both the quantity and quality of TAAs varied across the salinity treatments. Quantity was most strongly related to the growth rate of the seaweed and was highest in the slowest growing seaweed. In contrast, the quality of TAAs (individual amino acids as a proportion of total content) was most strongly related to salinity for all amino acids, although this varied substantially among individual amino acids. Increases in salinity were positively correlated with the proportion of proline (46% increase), tyrosine (36% increase), and histidine (26% increase), whereas there was a negative correlation with alanine (29% decrease). The proportion of methionine, with strong links to the synthesis of theosmolyte dimethylsulfoniopropionate, did not correlate linearly with salinity and instead was moderately higher at the optimal salinities for growth. These results show that salinity simultaneously affects the quantity and quality of TAAs in seaweed through both indirect and direct mechanisms, with growth rates playing the overarching role in determining the quantity of TAAs.

Anoikis-resistant subpopulations of human osteosarcoma display significant chemoresistance and are sensitive to targeted epigenetic therapies predicted by expression profiling.

BackgroundOsteosarcoma (OS) is the most common type of solid bone cancer, with latent metastasis being a typical mode of disease progression and a major contributor to poor prognosis. For this to occur, cells must resist anoikis and be able to recapitulate tumorigenesis in a foreign microenvironment. Finding novel approaches to treat osteosarcoma and target those cell subpopulations that possess the ability to resist anoikis and contribute to metastatic disease is imperative. Here we investigate anchorage-independent (AI) cell growth as a model to better characterize anoikis resistance in human osteosarcoma while using an expression profiling approach to identify and test targetable signaling pathways.MethodsEstablished human OS cell lines and patient-derived human OS cell isolates were subjected to growth in either adherent or AI conditions using Ultra-Low Attachment plates in identical media conditions. Growth rate was assessed using cell doubling times and chemoresistance was assessed by determining cell viability in response to a serial dilution of either doxorubicin or cisplatin. Gene expression differences were examined using quantitative reverse-transcription PCR and microarray with principal component and pathway analysis. In-vivo OS xenografts were generated by either subcutaneous or intratibial injection of adherent or AI human OS cells into athymic nude mice. Statistical significance was determined using student’s t-tests with significance set at α = 0.05.ResultsWe show that AI growth results in a global gene expression profile change accompanied by significant chemoresistance (up to 75 fold, p < 0.05). AI cells demonstrate alteration of key mediators of mesenchymal differentiation (β-catenin, Runx2), stemness (Sox2), proliferation (c-myc, Akt), and epigenetic regulation (HDAC class 1). AI cells were equally tumorigenic as their adherent counterparts, but showed a significantly decreased rate of growth in-vitro and in-vivo (p < 0.05). Treatment with the pan-histone deacetylase inhibitor vorinostat and the DNA methyltransferase inhibitor 5-azacytidine mitigated AI growth, while 5-azacytidine sensitized anoikis-resistant cells to doxorubicin (p < 0.05).ConclusionsThese data demonstrate remarkable plasticity in anoikis-resistant human osteosarcoma subpopulations accompanied by a rapid development of chemoresistance and altered growth rates mirroring the early stages of latent metastasis. Targeting epigenetic regulation of this process may be a viable therapeutic strategy.Electronic supplementary materialThe online version of this article (doi:10.1186/s12967-015-0466-4) contains supplementary material, which is available to authorized users.

The CpxRA two-component system is essential for Citrobacter rodentium virulence.

Citrobacter rodentium is a murine intestinal pathogen used as a model for the foodborne human pathogens enterohemorrhagic Escherichia coli and enteropathogenic E. coli. During infection, these pathogens use two-component signal transduction systems to detect and adapt to changing environmental conditions. In E. coli, the CpxRA two-component signal transduction system responds to envelope stress by modulating the expression of a myriad of genes. Quantitative real-time PCR showed that cpxRA was expressed in the colon of C57BL/6J mice infected with C. rodentium. To determine whether CpxRA plays a role during C. rodentium infection, a cpxRA deletion strain was generated and found to have a colonization defect during infection. This defect was independent of an altered growth rate or a defective type III secretion system, and single-copy chromosomal complementation of cpxRA restored virulence. The C. rodentium strains were then tested in C3H/HeJ mice, a lethal intestinal infection model. Mice infected with the ΔcpxRA strain survived infection, whereas mice infected with the wild-type or complemented strains succumbed to infection. Furthermore, we found that the cpxRA expression level was higher during early infection than at a later time point. Taken together, these data demonstrate that the CpxRA two-component signal transduction system is essential for the in vivo virulence of C. rodentium. In addition, these data suggest that fine-tuned cpxRA expression is important for infection. This is the first study that identifies a C. rodentium two-component transduction system required for pathogenesis. This study further indicates that CpxRA is an interesting target for therapeutics against enteric pathogens.

Effects of soil nutrients on the sequestration of plant defence chemicals by the specialist insect herbivore, Danaus plexippus

1. Although anthropogenic nitrogen (N) enrichment has significantly changed the growth, survival and reproduction of herbivorous insects, its effects on the defensive sequestration of secondary chemicals by insect herbivores are less well understood. Previous studies have shown that soil nutrient availability can affect sequestration directly through changing concentrations of plant defence chemicals, or indirectly through altering growth rates of herbivores. There has been less exploration of how nutrient deposition affects the consumption of secondary chemicals and subsequent sequestration efficiency. In the current study, the overall effect of soil N availability on cardenolide sequestration by the monarch caterpillar Danaus plexippus was examined. Specifically, the effects of soil nutrient availability on growth, consumption, excretion and sequestration efficiency of cardenolides by D. plexippus larvae fed on the tropical milkweed Asclepias curassavica were measured.2. The results showed that soil N and phosphorus (P) fertilisation significantly reduced caterpillar growth rate and the sequestration efficiency of cardenolides by monarch caterpillars feeding on A. curassavica. The lowered sequestration efficiency was accompanied by higher concentrations of cardenolides in frass. Although the total cardenolide contents of caterpillars were lower under high N or P fertilisation levels, caterpillar cardenolide concentrations were constant across fertilisation treatments because of lower growth rates (and therefore lower body mass) under high fertilisation. It is concluded that anthropogenic N deposition may have multiple effects on insect herbivores, including their ability to defend themselves from predators with sequestered plant defences.

Phosphorylation of pyruvate kinase A by protein kinase J leads to the altered growth and differential rate of intracellular survival of mycobacteria.

PknJ (Rv2088) is a serine/threonine protein kinase of mycobacteria which is present in Mycobacterium tuberculosis (MTB), but its gene is absent in Mycobacterium smegmatis (MS); a fast grower and nonpathogenic species of mycobacteria. The heterologous expression of MTB-specific PknJ in MS altered the growth of recombinant mycobacteria highlighting one of the characteristics of this protein. This nature of the protein was further confirmed when Mycobacterium bovis BCG (BCG) containing antisense copy of pknJ resulted in the increased growth of BCG. The real-time RNA quantification analysis pointed out toward increased expression of this protein during infection of THP-1 macrophage cells which further emphasized that the protein is essential for the intracellular survival of mycobacteria. The differential in gel electrophoresis (DIGE) data followed by mass spectroscopy suggested that PknJ is involved in regulation of pyruvate kinase A (Rv1617). Since pyruvate kinase (PK) A is one of the key enzymes which controls glycolytic cycle in mycobacteria, we looked for its interaction with PknJ during extracellular and intracellular growth of mycobacteria. In order to identify the specific residue(s) involved in post-translational modification, the phospho-null mutants of PK were generated, and their substrate specificities in response to PknJ were assessed through kinase assay. The findings thus underlined that the PK activity is predominantly dependent on the threonine residue at the 94(th) position and further suggested that this site may be plausible in intracellular survival of mycobacteria upon phosphorylation with PknJ.

Yeast Growth Plasticity Is Regulated by Environment-Specific Multi-QTL Interactions

For a unicellular, non-motile organism like Saccharomyces cerevisiae, carbon sources act both as nutrients and as signaling molecules and consequently affect various fitness parameters including growth. It is therefore advantageous for yeast strains to adapt their growth to carbon source variation. The ability of a given genotype to manifest different phenotypes in varying environments is known as phenotypic plasticity. To identify quantitative trait loci (QTL) that drive plasticity in growth, two growth parameters (growth rate and biomass) were measured in a published dataset from meiotic recombinants of two genetically divergent yeast strains grown in different carbon sources. To identify QTL contributing to plasticity across pairs of environments, gene–environment interaction mapping was performed, which identified several QTL that have a differential effect across environments, some of which act antagonistically across pairs of environments. Multi-QTL analysis identified loci interacting with previously known growth affecting QTL as well as novel two-QTL interactions that affect growth. A QTL that had no significant independent effect was found to alter growth rate and biomass for several carbon sources through two-QTL interactions. Our study demonstrates that environment-specific epistatic interactions contribute to the growth plasticity in yeast. We propose that a targeted scan for epistatic interactions, such as the one described here, can help unravel mechanisms regulating phenotypic plasticity.

Vulnerability of Coral Reef Fisheries to a Loss of Structural Complexity

Coral reefs face a diverse array of threats, from eutrophication and overfishing to climate change. As live corals are lost and their skeletons eroded, the structural complexity of reefs declines. This may have important consequences for the survival and growth of reef fish because complex habitats mediate predator-prey interactions [1, 2] and influence competition [3-5] through the provision of prey refugia. A positive correlation exists between structural complexity and reef fish abundance and diversity in both temperate and tropical ecosystems [6-10]. However, it is not clear how the diversity of available refugia interacts with individual predator-prey relationships to explain emergent properties at the community scale. Furthermore, we do not yet have the ability to predict how habitat loss might affect the productivity of whole reef communities and the fisheries they support. Using data from an unfished reserve in The Bahamas, we find that structural complexity is associated not only with increased fish biomass and abundance, but also with nonlinearities in the size spectra of fish, implying disproportionately high abundances of certain size classes. By developing a size spectrum food web model that links the vulnerability of prey to predation with the structural complexity of a reef, we show that these nonlinearities can be explained by size-structured prey refugia that reduce mortality rates and alter growth rates in different parts of the size spectrum. Fitting the model with data from a structurally complex habitat, we predict that a loss of complexity could cause more than a 3-fold reduction in fishery productivity.

Impact of S100A8/A9 Expression on Prostate Cancer Progression In Vitro and In Vivo

The proinflammatory S100A8/A9 proteins, which are expressed in myeloid cells under physiological conditions, are strongly expressed in human prostate cancer epithelial cells. Their role in the tumor cells and in tumor progression is largely unclear. We established a prostate cancer epithelial cell line (PC-3 TO-A8/A9) expressing S100A8 and S100A9 simultaneously under doxycycline control, to study the role of S100A8/A9 on tumor growth and infiltration of immune cells in subcutaneous xenografts in male NMRI nu/nu mice. Colonization of distant organs was studied after intracardial injection of the tumor cells in male NOD/SCID mice. PC-3 TO-A8/A9 cells grown in vitro and subcutaneous xenografts in mice not treated with doxycycline expressed high levels of S100A8/A9 mRNA and protein, whereas doxycycline treatment suppressed S100A8/A9 expression. S100A8/A9 expression did not significantly alter growth rate and invasion of the subcutaneous tumors into surrounding tissues. However, S100A8/A9 expression caused increased infiltration of immune cells, especially neutrophils. In intracardially injected mice sporadic tumor settlement was observed in muscle and lymph nodes. Colonies of tumor cells and micro-metastases were observed in the lung of 64.3% (9 out of 14) of mice not treated with doxycycline and in 33.3% (5 out of 15) of mice treated with doxycycline. Our data demonstrate for the first time that S100A8/A9 expression in epithelial cancer cells causes enhanced infiltration of immune cells, especially neutrophils, and stimulates settlement of the cancer cells in the lung.

Impact of feed restriction on health, digestion and faecal microbiota of growing pigs housed in good or poor hygiene conditions

Feed restriction could be a relevant strategy to preserve gut health, reduce systemic inflammatory response and finally limit antibiotic use. This study assessed the effect of feed restriction on growing pigs submitted to a moderate inflammatory challenge induced by the degradation of the environmental hygiene that is known to alter growth rate. The experiment was run on 80 pigs selected at 7 weeks of age according to a 2×2 factorial design: two feeding levels, ad libitum (AL) and feed restricted (FR) at 60% of AL, and two conditions of environmental hygiene, clean and dirty. Pigs were housed individually throughout the experiment. From 61 to 68 days of age (day 0 to 7), pigs were housed in a post weaning unit and feed restriction was applied to half of the pigs from day 0 to day 29. At 68 days of age (day 7 of the experiment), pigs were transferred in a growing unit where half of FR and half of AL pigs were housed in a dirty environment (poor hygiene) and the other half in a clean environment (good hygiene) until day 42. Growth performance was recorded weekly. Blood and faeces samples were collected to measure indicators of inflammation, nutrient digestibility and microbiota composition. Faecal consistency was monitored daily to detect diarrhoeas. Feed restriction decreased daily weight gain (−35% to −50%, P<0.001), increased the feed conversion ratio (+15%, P<0.001) and CP digestibility (+3%, P<0.05) and reduced the occurrence of diarrhoeas irrespective of hygiene conditions. Poor hygiene conditions decreased growth performance (−20%, P<0.05) and total tract digestibility of all nutrients (P<0.001). Haptoglobin (+50%) concentrations and lymphocyte (+10%) and granulocyte (+40%) numbers were higher in poor hygiene conditions (P<0.05), confirming that the model was effective to induce a systemic inflammatory response. Both feed restriction and hygiene modified the profile of the faecal microbiota. In this study, feed restriction did not reduce the systemic inflammatory response caused by poor hygiene conditions despite the limitation of the occurrence of digestive disorders. However, our study opens discussions regarding the impact of hygiene and feed restriction on gut microbial communities and digestive health.

Subacute oral toxicity of BDE-15, CDE-15, and HODE-15 in ICR male mice: assessing effects on hepatic oxidative stress and metals status and ascertaining the protective role of vitamin E

The present study examined the effects of oral exposure of 4,4'-dibromodiphenyl ether (BDE-15), 4,4'-dichlorodiphenyl ether (CDE-15), and 4,4'-dihydroxydiphenyl ether (HODE-15) on hepatic oxidative stress (OS) and metal status in Institute of Cancer Research (ICR) male mice. Furthermore, the role of vitamin E in ameliorating potential OS caused by BDE-15, CDE-15, and HODE-15 was investigated. Three groups of mice were exposed to 1.20 mg/kg(body weight)/day of each of the three toxicants for 28 days. Results showed that none of the three toxicants altered growth rates of mice, but significantly increased (P<0.05) relative liver weights and decreased relative kidney weights. Pathological changes including cell swelling, inflammation and vacuolization, and hepatocellular hypertrophy in livers were observed. Significant decreases (P<0.05 and P<0.01) in superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) activity, and glutathione (GSH) levels, together with increases in malondialdehyde (MDA) content were recorded in all toxicant-treated groups. Hepatic copper levels increased in all toxicant-treated groups. Hepatic zinc levels decreased in the liver of BDE-15-treated mice, whereas they increased in the livers of CDE-15-treated and HODE-15-treated mice. In conclusion, daily exposure to the three toxicants perturbed metal homeostasis and increased OS in mouse liver. Experimental data indicated the hepatic oxidative toxicity of the three toxicants followed the order BDE-15<HODE-15<CDE-15. Moreover, the study proved that daily supplementation of 50 mg/kg vitamin E is effective to ameliorate the hepatic OS status and metal disturbance in mice.

Will Plants Grow on Moon or Mars?

The cultivation of plants in space and on the Moon and Mars implies exposure to physical factors that are different from Earth: radiation, gravity levels and magnetic fields. As part of the MELiSSA (Micro-Ecological Life Support System Alternative) project where higher plants represent one of the compartments of an enclosed life support system, literature has been reviewed to assemble the relevant knowledge within space plant research. This review has particularly focused on the effect of the physical factors mentioned above on higher plant morphology, anatomy, gene regulation and genetic damage. Radiation studies on ground have shown increased damages to plant cells, and magnetic field studies have indicated some stress responses and altered growth rates, although not conclusive. Both in space and on ground (clinostat experiments) there are inconsistent results regarding gene expression and plant morphology. Anatomical flaws on the cellular level such as a reduced proliferation, displaced statoliths, ovoid chloroplasts, progressive vacuolization, and altered cell walls have been observed in several experiments. A normal plant reproduction process in weightlessness is feasible with a properly designed hardware. The same can be expected with low gravity levels, like those on Moon and Mars. In conclusion, the published results show that plants will grow and reproduce in low Earth orbit, but they might experience genotoxic stress and anatomic changes. Longer space studies, in particular outside the geomagnetic field and its radiation belt, are needed in order to investigate potential adaption responses since many studies show that plants can adapt in one generation to extreme environments.

Nail abnormalities associated with systemic pathologies

Nail abnormalities can arise in conjunction with or as a result of systematic pathologies. These pathologies include single-organ diseases, multisystemic diseases, and drug-induced insults. Clinical signs associated with these conditions include dyschromias, vascular alterations, periungual tissue changes, textural dystrophies, contour alterations, and growth-rate alterations. The associated systemic pathologies may affect any part of the nail apparatus, including the nail matrix, the nail plate, the nail bed, the underlying vasculature, and the periungual tissues. The anatomical location and extent of damage determine the clinically manifested anomaly.

Exposure to ionizing radiations and starvation culture does not modify phenotype, functions and genetic profile of mesenchymal stromal cells isolated from bone marrow of healthy donors

Mesenchymal stromal cells (MSCs) are multipotent cells located in different human tissues, including bone marrow (BM), where they sustain the creation of the hematopoietic stem cell niche by regulating its differentiation and functions. Many studies have demonstrated that BM exposure to ionizing radiations induces the rapid depletion of hematopoietic precursors, however radiation effects on MSCs have been poorly investigated. Moreover, little is known about MSCs behavior in starvation culture conditions. In this study, we examined morphology, proliferative capacity (in terms of population doublings), immunophenotype, differentiation potential, immunomodulatory properties (PHA-induced T cell proliferation assay) and genetic profile (array-CGH and caryotype) of MSCs isolated from BM of healthy donors and exposed to ionizing radiations and starvation culture. MSCs were isolated from 10 healthy donors (median age: 16 years; range: 5-32) and expanded in culture medium supplemented with 5% platelet lysate up to passage 2. Thereafter MSCs were exposed both to escalating doses of ionizing radiations (3000, 10000 and 20000 rad) and to starvation culture conditions (culture medium supplemented with 1% platelet lysate instead of 5%). With escalating doses of ionizing radiations, MSCs lose their typical spindle-shaped morphology, their boundaries are less regular and their growth rate slows down (at 3000 rad) or even stops (at 10000 and 20000 rad). Nonetheless, in the presence of 1% platelet lysate, although showing a slower growth rate as compared with non irradiated MSCs, the effects on morphology are less evident, thus suggesting that the lack of growing factors can slow down the senescent process induced by ionizing radiations. Furthermore, irradiated and starved MSCs maintain the same immunophenotype, differentiation potential, immunomodulatory properties and genetic profile of normal MSCs. Our data indicate that irradiated and starved MSCs, although presenting altered morphology and growth rate, maintain the typical characteristics of normal MSCs.

A phase II study of a yeast-based therapeutic cancer vaccine, GI-6207, targeting CEA in patients with minimally symptomatic, metastatic medullary thyroid cancer.

TPS3127 Background: Saccharomyces cerevisiae has been genetically modified to express CEA protein and developed under a CRADA with GlobeImmune/NCI as a heat-killed immune-stimulating, therapeutic cancer vaccine (GI-6207). A phase I study with GI-6207 demonstrated safety, biomarker stabilization and enhanced immune response in some patients. CEA is over-expressed in multiple malignancies, including medullary thyroid cancer (MTC). Two therapies recently approved by the FDA for metastatic MTC (vandetanib, cabozantinib) come with toxicity and should be reserved for symptomatic/progressive disease. However, a large population of asymptomatic MTC patients has small tumor burden and/or disease that is more indolent. The standard management of these patients is observation. Preliminary data suggest that tumor growth measured by the rate of CEA and calcitonin increase can be quantified in a 3-6 months. Retrospective data from prostate cancer studies suggest vaccines can alter growth rates within 3-4 months. We hypothesize that GI-6207 can alter tumor growth rates in MTC and impact long-term outcome. Methods: A phase II study will evaluate the effect of GI-6207 onthe rates ofincrease in calcitonin in metastatic MTC. 34 patients with minimally symptomatic, radiographically evaluable, metastatic MTC will be randomized 1:1. Arm A will receive vaccine for a year from the time of enrollment. Arm B will receive vaccine after 6 months of surveillance. GI-6207 will be administered subcutaneously at 4 sites (10 yeast units/site), every 2 weeks for 3 months, then monthly up to 1 year. The primary endpoint will compare the effect of GI-6207 on calcitonin kinetics between the vaccine and surveillance arms in the first 6 months. Secondary endpoints include immunologic responses (including antigen-specific T cell responses), objective responses, time to progression, and changes in CEA kinetics. If this trial can prospectively demonstrate that vaccines can alter tumor growth rates, and if such changes are associated with clinical outcomes, then changes in tumor growth rates may become a clinical metric to evaluate vaccine efficacy in MTC and other populations.

Ilizarov treatment for pseudarthrosis of the tibia due to haematogenous osteomyelitis

Haematogenous osteomyelitis is associated with poverty and overcrowding. The infection may extend through the entire length of a long bone and as a part of debridement, the sequestrum may be removed, resulting in a defect. Infection of tibial growth plates would result in altered growth rates of the tibia and fibula and produce deformities and shortening. Fourteen children were treated for pseudarthrosis of the tibia because of haematogenous osteomyelitis. Their ages ranged from 4 to 13 years. There were nine male and five female children. The treatment included the use of Ilizarov techniques in the form of distraction of a stiff pseudarthrosis (n=5) and acute shortening and relengthening for bone defects (n=9). No active infection was present at the time of treatment. In all cases, the bone defects were bridged and consolidated. No residual infection was present. Transient palsy of the common peroneal nerve occurred in two cases during deformity correction and lengthening. Refracture occurred in one case at the site of a previous wire. Residual leg shortening of more than 1 inch was present in two cases. The results were satisfactory in 12 cases (86%) and unsatisfactory in two cases (14%) because of residual shortening and ankle joint fusion. Ilizarov techniques comprehensively address the problems associated with chronic haematogenous osteomyelitis of the tibia in children despite the difficulties in their application because of the nature of the illness.

Short- and Long-Term Effects of LRRK2 on Axon and Dendrite Growth

Mutations in leucine-rich repeat kinase 2 (LRRK2) underlie an autosomal-dominant form of Parkinson's disease (PD) that is clinically indistinguishable from idiopathic PD. The function of LRRK2 is not well understood, but it has become widely accepted that LRRK2 levels or its kinase activity, which is increased by the most commonly observed mutation (G2019S), regulate neurite growth. However, growth has not been measured; it is not known whether mean differences in length correspond to altered rates of growth or retraction, whether axons or dendrites are impacted differentially or whether effects observed are transient or sustained. To address these questions, we compared several developmental milestones in neurons cultured from mice expressing bacterial artificial chromosome transgenes encoding mouse wildtype-LRRK2 or mutant LRRK2-G2019S, Lrrk2 knockout mice and non-transgenic mice. Over the course of three weeks of development on laminin, the data show a sustained, negative effect of LRRK2-G2019S on dendritic growth and arborization, but counter to expectation, dendrites from Lrrk2 knockout mice do not elaborate more rapidly. In contrast, young neurons cultured on a slower growth substrate, poly-L-lysine, show significantly reduced axonal and dendritic motility in Lrrk2 transgenic neurons and significantly increased motility in Lrrk2 knockout neurons with no significant changes in length. Our findings support that LRRK2 can regulate patterns of axonal and dendritic growth, but they also show that effects vary depending on growth substrate and stage of development. Such predictable changes in motility can be exploited in LRRK2 bioassays and guide exploration of LRRK2 function in vivo.

The15N isotope effect inEscherichia coli: A neutron can make the difference

Several techniques based on stable isotope labeling are used for quantitative MS. These include stable isotope metabolic labeling methods for cells in culture as well as live organisms with the assumption that the stable isotope has no effect on the proteome. Here, we investigate the (15) N isotope effect on Escherichia coli cultures that were grown in either unlabeled ((14) N) or (15) N-labeled media by LC-ESI-MS/MS-based relative protein quantification. Consistent protein expression level differences and altered growth rates were observed between (14) N and (15) N-labeled cultures. Furthermore, targeted metabolite analyses revealed altered metabolite levels between (14) N and (15) N-labeled bacteria. Our data demonstrate for the first time that the introduction of the (15) N isotope affects protein and metabolite levels in E. coli and underline the importance of implementing controls for unbiased protein quantification using stable isotope labeling techniques.

The Effects of Disturbance Events on Abundance and Sex Ratios of a Terrestrial Turtle, Terrapene bauri

Abstract Disturbances can affect the structure of ecological communities, and their impacts may have consequences for individual species' population dynamics and long-term persistence. Even without catastrophic mortality, survivorship could be reduced following a disturbance, thus leading to population decline. We used a 16-year mark–recapture dataset to determine the effects of catastrophic storms and the cutting of nonindigenous vegetation on the survivorship and abundance of a population of long-lived terrestrial turtles (Terrapene bauri). Our results indicate that these habitat disturbances had little long-term effect on abundance. Indeed, the population continued to grow despite severe changes to its habitat, although recapture probabilities decreased through time. Tertiary (adult) sex ratios became more highly skewed temporarily but eventually approached their predisturbance levels. In long-lived box turtles, short-term disturbances may alter growth rates and dispersal tendencies, but they appear to...

Underlying causes of habitat-associated differences in size of age-0 walleye pollock (Theragra chalcogramma) in the Gulf of Alaska

Age-0 walleye pollock (Theragra chalcogramma) caught in September in the Gulf of Alaska display habitat-associated differences in standard length (SL). Age-0 fish collected in the region around Sutwik Island and 375 km farther downstream near the Shumagin Islands most likely originate from the Shelikof Strait spawning aggregation. However, age-0 fish resulting from the same spawning aggregation differ in mean size up to 20 mm between areas by September. We examined the otoliths of the larval and age-0 stages of walleye pollock from these two areas in 2000 and 2001 to determine whether growth rate, hatch date, and/or temperature influenced fish size. Circulation models were used to determine whether transport of larvae from an upstream spawning group into the study areas could have occurred. Mean in situ temperature during sampling periods was not defined as a significant factor in altering growth rates. Overlapping hatch date distributions of the larval and age-0 fish in the Shumagin Island area confirmed that the fish were from the Shelikof Strait spawning group. Comparison of hatch date distributions in the upstream Sutwik Island area revealed larger/older larvae from an upstream spawning group mixed with larvae from the Shelikof Strait spawning group. Our results suggest that the offset of 20 mm SL between the groups of age-0 pollock was the result of a combination of enhanced survivorship of early-hatched larvae in the Sutwik area and the introduction and retention of the progeny of another spawning group originating upstream of Shelikof Strait.