Growth Cone Localization of the mRNA Encoding the Chromatin Regulator HMGN5 Modulates Neurite Outgrowth

Neurons exploit local mRNA translation and retrograde transport of transcription factors to regulate gene expression in response to signaling events at distal neuronal ends. Whether epigenetic factors could also be involved in such regulation is not known. We report that the mRNA encoding the high-mobility group N5 (HMGN5) chromatin binding protein localizes to growth cones of both neuron-like cells and of hippocampal neurons, where it has the potential to be translated, and that HMGN5 can be retrogradely transported into the nucleus along neurites. Loss of HMGN5 function induces transcriptional changes and impairs neurite outgrowth, while HMGN5 overexpression induces neurite outgrowth and chromatin decompaction; these effects are dependent on growth cone localization of Hmgn5 mRNA. We suggest that the localization and local translation of transcripts coding for epigenetic factors couple the dynamic neuronal outgrowth process with chromatin regulation in the nucleus.

Growth cone MKK7 mRNA targeting regulates MAP1b-dependent microtubule bundling to control neurite elongation

Local mRNA translation in neurons has been mostly studied during axon guidance and synapse formation but not during initial neurite outgrowth. We performed a genome-wide screen for neurite-enriched mRNAs and identified an mRNA that encodes mitogen-activated protein kinase kinase 7 (MKK7), a MAP kinase kinase (MAPKK) for Jun kinase (JNK). We show that MKK7 mRNA localizes to the growth cone where it has the potential to be translated. MKK7 is then specifically phosphorylated in the neurite shaft, where it is part of a MAP kinase signaling module consisting of dual leucine zipper kinase (DLK), MKK7, and JNK1. This triggers Map1b phosphorylation to regulate microtubule bundling leading to neurite elongation. We propose a model in which MKK7 mRNA localization and translation in the growth cone allows for a mechanism to position JNK signaling in the neurite shaft and to specifically link it to regulation of microtubule bundling. At the same time, this uncouples activated JNK from its functions relevant to nuclear translocation and transcriptional activation.

Two distinct filopodia populations at the growth cone allow to sense nanotopographical extracellular matrix cues to guide neurite outgrowth

BACKGROUND The process of neurite outgrowth is the initial step in producing the neuronal processes that wire the brain. Current models about neurite outgrowth have been derived from classic two-dimensional (2D) cell culture systems, which do not recapitulate the topographical cues that are present in the extracellular matrix (ECM) in vivo. Here, we explore how ECM nanotopography influences neurite outgrowth. METHODOLOGY/PRINCIPAL FINDINGS We show that, when the ECM protein laminin is presented on a line pattern with nanometric size features, it leads to orientation of neurite outgrowth along the line pattern. This is also coupled with a robust increase in neurite length. The sensing mechanism that allows neurite orientation occurs through a highly stereotypical growth cone behavior involving two filopodia populations. Non-aligned filopodia on the distal part of the growth cone scan the pattern in a lateral back and forth motion and are highly unstable. Filopodia at the growth cone tip align with the line substrate, are stabilized by an F-actin rich cytoskeleton and enable steady neurite extension. This stabilization event most likely occurs by integration of signals emanating from non-aligned and aligned filopodia which sense different extent of adhesion surface on the line pattern. In contrast, on the 2D substrate only unstable filopodia are observed at the growth cone, leading to frequent neurite collapse events and less efficient outgrowth. CONCLUSIONS/SIGNIFICANCE We propose that a constant crosstalk between both filopodia populations allows stochastic sensing of nanotopographical ECM cues, leading to oriented and steady neurite outgrowth. Our work provides insight in how neuronal growth cones can sense geometric ECM cues. This has not been accessible previously using routine 2D culture systems.

Contrasting growth and water use efficiency after thinning in mixed Abies pinsapo–Pinus pinaster–Pinus sylvestris forests

Foresters frequently lack sufficient information about thinning intensity effects to optimize semi-natural forest management and their effects and interaction with climate are still poorly understood. In an Abies pinsapo–Pinus pinaster–Pinus sylvestris forest with three thinning intensities, a dendrochronologial approach was used to evaluate the short-term responses of basal area increment (BAI), carbon isotope (δ13C) and water use efficiency (iWUE) to thinning intensity and climate. Thinning generally increased BAI in all species, except for the heavy thinning in P. sylvestris. Across all the plots, thinning increased 13C-derived water-use efficiency on average by 14.49% for A. pinsapo, 9.78% for P. sylvestris and 6.68% for P. pinaster, but through different ecophysiological mechanisms. Our findings provide a robust mean of predicting water use efficiency responses from three coniferous species exposed to different thinning strategies which have been modulated by climatic conditions over time.

NO2-induced nitrate reductase activity in needles of Norway spruce (Picea abies) under laboratory and field conditions

The induction of activity of the enzyme nitrate reductase (NR, EC 1.6.6.1, 1.6.6.2) in needles of Norway spruce (Picea abies[L.] Karst.) by nitrogen dioxide (NO2) was studied under laboratory and field conditions. In fumigation chambers an increase in nitrate reductase activity (NRA) was detected 4 h after the start of the NO2 treatment. During the first 2 days with 100 µg NO2 m−3, NRA reached a constant level and did not change during the following 4 days. At the same level of NO2, NRA was lower in needles from trees grown on NPK-fertilized soil than on non-fertilized soil. After the transfer of spruce trees from fertilized soil to NPK-rich nutrient solution, NRA was transiently increased. This effect was assigned to root injuries causing nitrate transport to the shoot and subsequent induction of NRA. Neither trees on fertilized soil nor trees transferred to NPK-poor nutrient solution had increased NRA unless NO2 was provided. The NO2 gradient in the vicinity of a highway was used to test the long-term effect of elevated levels of NO2 on needle NRA of potted and field-grown spruce trees. Compared with less polluted sites, permanently increased NRAs were detected when NO2 concentrations were above 20 µg m−3. Controls of field measurements some 10 years after the introduction of catalytic converters in cars showed no significant change neither in NO2 levels nor in the decreasing NRA of spruce needles with the distance from the highway.

Phytoplankton Scales of Variability in the California Current System: 2. Latitudinal Variability

The California Current System encompasses a southward flowing current which is perturbed by ubiquitous mesoscale variability. The extent to which latitudinal patterns of physical variability are reflected in the distribution of biological parameters is poorly known. To investigate the latitudinal distribution of chlorophyll variance, a wavelet analysis is applied to nearly 9 years (October 1997 to July 2006) of 1-km-resolution Sea-viewing Wide Field-of-view Sensor (SeaWiFS) chlorophyll concentration data at 5-day resolution. Peaks in the latitudinal distribution of chlorophyll variance coincide with features of the coastal topography. Maxima in variance are located offshore of Vancouver Island and downstream of Heceta Bank, Cape Blanco, Point Arena, and possibly Point Conception. An analysis of dominant wavelengths in the chlorophyll data reveals a transfer of energy into smaller scales is generated in the vicinity of the coastal capes. The latitudinal distribution of variance in sea level anomaly corresponds closely to the chlorophyll variance in the nearshore region (<100 km offshore), suggesting that the same processes determine the distribution of both. Farther offshore, there is no correspondence between latitudinal patterns of sea level anomaly and chlorophyll variance. This likely represents a transition from physical to biological control of the phytoplankton distribution.

Assessment of cone beam computed tomography techniques for imaging lung damage in mice in vivo

Lung damage is a common side effect of chemotherapeutic drugs such as bleomycin. This study used a bleomycin mouse model which simulates the lung damage observed in humans. Noninvasive, in vivo cone-beam computed tomography (CBCT) was used to visualize and quantify fibrotic and inflammatory damage over the entire lung volume of mice. Bleomycin was used to induce pulmonary damage in vivo and the results from two CBCT systems, a micro-CT and flat panel CT (fpCT), were compared to histologic measurements, the standard method of murine lung damage quantification. Twenty C57BL/6 mice were given either 3 U/kg of bleomycin or saline intratracheally. The mice were scanned at baseline, before the administration of bleomycin, and then 10, 14, and 21 days afterward. At each time point, a subset of mice was sacrificed for histologic analysis. The resulting CT images were used to assess lung volume. Percent lung damage (PLD) was calculated for each mouse on both the fpCT (PLDfpcT) and the micro-CT (PLDμCT). Histologic PLD (PLDH) was calculated for each histologic section at each time point (day 10, n = 4; day 14, n = 4; day 21, n = 5; control group, n = 5). A linear regression was applied to the PLDfpCT vs. PLDH, PLDμCT vs. PLDH and PLDfpCT vs. PLDμCT distributions. This study did not demonstrate strong correlations between PLDCT and PLDH. The coefficient of determination, R, was 0.68 for PLDμCT vs. PLDH and 0.75 for the PLD fpCT vs. PLDH. The experimental issues identified from this study were: (1) inconsistent inflation of the lungs from scan to scan, (2) variable distribution of damage (one histologic section not representative of overall lung damage), (3) control mice not scanned with each group of bleomycin mice, (4) two CT systems caused long anesthesia time for the mice, and (5) respiratory gating did not hold the volume of lung constant throughout the scan. Addressing these issues might allow for further improvement of the correlation between PLDCT and PLDH. ^

Body image rating scales to assess obesity

Obesity prevalence in the U.S. has increased during the last three decades with major impact on public health. Screening for obesity in a population with unknown weight status can be time- and resource-consuming, but the information is valuable for prioritizing and allocating scarce resources. The challenge remains to properly assess obesity with the available methods. Body Image Rating Scales (BIRS) have initially been developed to assess body image disturbances, but also seem useful as an alternative method in assessing obesity prevalence. Several different BIRS exists. In this project I reviewed the literature that exists regarding the use of BIRS, and its advantages and limitations for the assessment of obesity status with regards to BMI. The result yielded nine publications that examined eight different scales and their correlation with BMI, ranging from r=.59 for self-reported BMI to r=.94 for measured BMI. One concern is the lack of standardization of this method to assess obesity, given the range of different scales. While many methods for obesity assessment are available, the simplicity, ease of use and cost-effectiveness of BIRS make it very appealing. BIRS remain a potentially attractive option to assess the weight status of a large population with minimal requirements in assets and time, especially in situations where measuring instruments are not available, or when height or weight could not be recalled.^

Design and Optimization of Four-dimensional Cone-beam Computed Tomography in Image-guided Radiation Therapy

The influence of respiratory motion on patient anatomy poses a challenge to accurate radiation therapy, especially in lung cancer treatment. Modern radiation therapy planning uses models of tumor respiratory motion to account for target motion in targeting. The tumor motion model can be verified on a per-treatment session basis with four-dimensional cone-beam computed tomography (4D-CBCT), which acquires an image set of the dynamic target throughout the respiratory cycle during the therapy session. 4D-CBCT is undersampled if the scan time is too short. However, short scan time is desirable in clinical practice to reduce patient setup time. This dissertation presents the design and optimization of 4D-CBCT to reduce the impact of undersampling artifacts with short scan times. This work measures the impact of undersampling artifacts on the accuracy of target motion measurement under different sampling conditions and for various object sizes and motions. The results provide a minimum scan time such that the target tracking error is less than a specified tolerance. This work also presents new image reconstruction algorithms for reducing undersampling artifacts in undersampled datasets by taking advantage of the assumption that the relevant motion of interest is contained within a volume-of-interest (VOI). It is shown that the VOI-based reconstruction provides more accurate image intensity than standard reconstruction. The VOI-based reconstruction produced 43% fewer least-squares error inside the VOI and 84% fewer error throughout the image in a study designed to simulate target motion. The VOI-based reconstruction approach can reduce acquisition time and improve image quality in 4D-CBCT.