Process Analysis and Synthesis

Contains reports on two research projects

Nonlinear Circuits

Contains reports on four research projects

Immune cell profile of sentinel lymph nodes in patients with malignant melanoma - FOXP3+ cell density in cases with positive sentinel node status is associated with unfavorable clinical outcome

BACKGROUND: Besides being a preferential site of early metastasis, the sentinel lymph node (SLN) is also a privileged site of T-cell priming, and may thus be an appropriate target for investigating cell types involved in antitumor immune reactions. METHODS: In this retrospective study we determined the prevalence of OX40+ activated T lymphocytes, FOXP3+ (forkhead box P3) regulatory T cells, DC-LAMP+ (dendritic cell-lysosomal associated membrane protein) mature dendritic cells (DCs) and CD123+ plasmacytoid DCs by immunohistochemistry in 100 SLNs from 60 melanoma patients. Density values of each cell type in SLNs were compared to those in non-sentinel nodes obtained from block dissections (n = 37), and analyzed with regard to associations with clinicopathological parameters and disease outcome. RESULTS: Sentinel nodes showed elevated amount of all cell types studied in comparison to non-sentinel nodes. Metastatic SLNs had higher density of OX40+ lymphocytes compared to tumor-negative nodes, while no significant difference was observed in the case of the other cell types studied. In patients with positive sentinel node status, high amount of FOXP3+ cells in SLNs was associated with shorter progression-free (P = 0.0011) and overall survival (P = 0.0014), while no significant correlation was found in the case of sentinel-negative patients. The density of OX40+, CD123+ or DC-LAMP+ cells did not show significant association with the outcome of the disease. CONCLUSIONS: Taken together, our results are compatible with the hypothesis of functional competence of sentinel lymph nodes based on the prevalence of the studied immune cells. The density of FOXP3+ lymphocytes showed association with progression and survival in patients with positive SLN status, while the other immune markers studied did not prove of prognostic importance. These results, together with our previous findings on the prognostic value of activated T cells and mature DCs infiltrating primary melanomas, suggest that immune activation-associated markers in the primary tumor may have a higher impact than those in SLNs on the prognosis of the patients. On the other hand, FOXP3+ cell density in SLNs, but not in the primary tumor, was found predictive of disease outcome in melanoma patients

Trends in stream nitrogen concentrations for forested reference catchments across the USA

To examine whether stream nitrogen concentrations in forested reference catchments have changed over time and if patterns were consistent across the USA, we synthesized up to 44 yr of data collected from 22 catchments at seven USDA Forest Service Experimental Forests. Trends in stream nitrogen presented high spatial variability both among catchments at a site and among sites across the USA. We found both increasing and decreasing trends in monthly flow-weighted stream nitrate and ammonium concentrations. At a subset of the catchments, we found that the length and period of analysis influenced whether trends were positive, negative or non-significant. Trends also differed among neighboring catchments within several Experimental Forests, suggesting the importance of catchment-specific factors in determining nutrient exports. Over the longest time periods, trends were more consistent among catchments within sites, although there are fewer long-term records for analysis. These findings highlight the critical value of long-term, uninterrupted stream chemistry monitoring at a network of sites across the USA to elucidate patterns of change in nutrient concentrations at minimally disturbed forested sites

Nonlinear Circuits

Contains reports on four research projects

Changing forest water yields in response to climate warming: results from long-term experimental watershed sites across North America

Climate warming is projected to affect forest water yields but the effects are expected to vary. We investigated how forest type and age affect water yield resilience to climate warming. To answer this question, we examined the variability in historical water yields at long-term experimental catchments across Canada and the United States over 5-year cool and warm periods. Using the theoretical framework of the Budyko curve, we calculated the effects of climate warming on the annual partitioning of precipitation (P) into evapotranspiration (ET) and water yield. Deviation (d) was defined as a catchment’s change in actual ET divided by P [AET/P; evaporative index (EI)] coincident with a shift from a cool to a warm period – a positive d indicates an upward shift in EI and smaller than expected water yields, and a negative d indicates a downward shift in EI and larger than expected water yields. Elasticity was defined as the ratio of inter annual variation in potential ET divided by P (PET/P; dryness index) to inter annual variation in the EI – high elasticity indicates low d despite large range in drying index (i.e., resilient water yields), low elasticity indicates high d despite small range in drying index (i.e., non-resilient water yields). Although the data needed to fully evaluate ecosystems based on these metrics are limited, we were able to identify some characteristics of response among forest types. Alpine sites showed the greatest sensitivity to climate warming with any warming leading to increased water yields. Conifer forests included catchments with lowest elasticity and stable to larger water yields. Deciduous forests included catchments with intermediate elasticity and stable to smaller water yields. Mixed coniferous/deciduous forests included catchments with highest elasticity and stable water yields. Forest type appeared to influence the resilience of catchment water yields to climate warming, with conifer and deciduous catchments more susceptible to climate warming than the more diverse mixed forest catchments

Trends in stream nitrogen concentrations for forested reference catchments across the USA

To examine whether stream nitrogen concentrations in forested reference catchments have changed over time and if patterns were consistent across the USA, we synthesized up to 44 yr of data collected from 22 catchments at seven USDA Forest Service Experimental Forests. Trends in stream nitrogen presented high spatial variability both among catchments at a site and among sites across the USA. We found both increasing and decreasing trends in monthly flow-weighted stream nitrate and ammonium concentrations. At a subset of the catchments, we found that the length and period of analysis influenced whether trends were positive, negative or non-significant. Trends also differed among neighboring catchments within several Experimental Forests, suggesting the importance of catchment-specific factors in determining nutrient exports. Over the longest time periods, trends were more consistent among catchments within sites, although there are fewer long-term records for analysis. These findings highlight the critical value of long-term, uninterrupted stream chemistry monitoring at a network of sites across the USA to elucidate patterns of change in nutrient concentrations at minimally disturbed forested sites.Keywords: forested catchment, nitrate, trends, stream, reference catchments, ammoniumKeywords: forested catchment, nitrate, trends, stream, reference catchments, ammoniu

Decoding a cancer-relevant splicing decision in the RON proto-oncogene using high-throughput mutagenesis

Mutations causing aberrant splicing are frequently implicated in human diseases including cancer. Here, we establish a high-throughput screen of randomly mutated minigenes to decode the cis-regulatory landscape that determines alternative splicing of exon 11 in the proto-oncogene MST1R (RON). Mathematical modelling of splicing kinetics enables us to identify more than 1000 mutations affecting RON exon 11 skipping, which corresponds to the pathological isoform RON Delta 165. Importantly, the effects correlate with RON alternative splicing in cancer patients bearing the same mutations. Moreover, we highlight heterogeneous nuclear ribonucleoprotein H (HNRNPH) as a key regulator of RON splicing in healthy tissues and cancer. Using iCLIP and synergy analysis, we pinpoint the functionally most relevant HNRNPH binding sites and demonstrate how cooperative HNRNPH binding facilitates a splicing switch of RON exon 11. Our results thereby offer insights into splicing regulation and the impact of mutations on alternative splicing in cancer.Institute of Molecular Biology Core Facilities; DFG [ZA 881/2-1, KO 4566/4-1, LE 3473/2-1]; LOEWE program Ubiquitin Networks (Ub-Net) of the State of Hesse (Germany); Deutsche Forschungsgemeinschaft [SFB902 B13]; EMBO [3057]; Fundacao para a Ciencia e a Tecnologia, Portugal (FCT Investigator Starting Grant) [IF/00595/2014]; German Federal Ministry of Research (BMBF; e:bio junior group program) [FKZ: 0316196]; Boehringer Ingelheim Foundation; [INST 47/870-1 FUGG