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The U.S. Army Corps of Engineers (USACE) has a major responsibility for the regulation of the Nation’s streams, rivers, and waterways. This often requires developing water quality models to resolve issues and concerns with regard to the environment and ecosystems. USACE Engineer Research and Development Center (ERDC) is currently developing an Eco-Hydrology Engineering Design Tool supported by years of research and development. This tool development integrates Gridded Surface Subsurface Hydrologic Analysis (GSSHA) model and a suite of Water Quality and Ecosystem Models (Clearwater Water Quality Modules - ClearWater). Weather is not the only cause of flooding, stream erosion, and pollution. These problems usually occur from human impacts to watersheds, including urban development, construction activities, hydrologic modifications, forestry, mining, and agricultural practices. To help control the impact of these serious economic and environmental issues, experts in watershed engineering at the U.S. Army Engineer Research and Development Center (ERDC) developed the Gridded Surface Subsurface Hydrologic Analysis (GSSHA) tool to conduct studies in watershed modeling. With GSSHA, engineers can perform complex studies of atmospheric, land-based, wetland, riverine, and coastal systems to help predict and mitigate watershed management problems. ERDC studies in watershed modeling and analysis integrate hydrology, hydraulics, and water quality. This helps city or military installation planners, civil engineers, and others involved in planning and resource management make informed decisions about watershed management. GSSHA is a multidimensional modeling technology that uniformly couples overland, surface, and subsurface flow for accurate watershed simulation. GSSHA is a physics-based, distributed, hydrologic, sediment and constituent fate and transport model featuring the following: 1) Two-dimensional (2D) overland flow and groundwater flow, and one-dimensional (1D) stream flow and soil moisture; 2) Fully dynamic pipe networks for urban and agricultural drainage systems; 3) Wetland peat layer hydrodynamics and several in-stream weir and culvert models; 4) Lakes, detention basins, levees, rating and rule curve releases; 5) Boundary conditions for hurricane storm surge or levee breach inundation modeling; 6) Full coupling among groundwater, vadose zone, streams, and overland flow; 7) Full-Gr-coupled groundwater to surface-water interaction to model Hortonian and non-Hortonian basins; 8) GSSHA can be used as an episodic or continuous model where soil surface moisture, groundwater levels, stream interactions, and constituent fate are continuously simulated; and 9) Full Energy Based Water and Soil Temperature for Overland and Channel systems. This presentation will discuss current GSSHA capabilities and introduce Water Quality and Ecosystem developments to be presented in more detail in “Part 2 - Eco-Hydrology Engineering Design Tool - ClearWater Capabilities - General Constituents, Nutrients, and Contaminants”.

A key pathological hallmark in >97% of all Amyotrophic Lateral Sclerosis (ALS) cases is the cytoplasmic mislocalization and aggregation of a nuclear RNA binding protein, TDP-43. Driving clearance of cytoplasmic TDP-43 reduces toxicity in various ALS models, though how TDP-43 clearance is regulated remains controversial. To address this, we conducted an unbiased yeast genome-wide screen using high-throughput dot blots to identify genes that affect TDP-43 levels. Our screen identified ESCRT complex factors, which induce membrane invagination (particularly at multi-vesicular bodies; MVBs) and K63-linked ubiquitination as key facilitators of TDP-43 endolysosomal clearance. TDP-43 co-localized and bound Rsp5/NEDD4 and ESCRT proteins, and perturbations to either increased TDP-43 aggregation and accumulation. NEDD4 also ubiquitinates TDP-43. Lastly, TDP-43 accumulation caused formation of “giant” MVBs which could reflect a pathological consequence of TDP-43 pertinent to ALS. Our studies shed light on endolysosomal-mediated cytoplasmic protein degradation, which likely impacts multiple substrates, and may be a target for novel ALS therapeutic strategies.

Non-aqueous gravel pack carrier fluids (GPCF) have been introduced into the industry to eliminate the risks associated with the water-based carrier fluids in the presence of reactive shale interbeds in the reservoir. However, non-aqueous GPCF pose a significant barrier to the effective deployment of post-gravel pack filter cake breaker (FCB) application because all FCB systems are water-based. Therefore, a novel approach was developed for FCB application in non-aqueous GPCF environment to improve the efficiency of the FCB and the overall well performance. The non-aqueous GPCF was redesigned from ground up to promote the better diffusion of the FCB. This was accomplished by introducing a reversible emulsifier package into the non-aqueous GPCF design which allows the gravel to change wettability from an oil-wet state to a water-wet state when a low pH solution i.e., breaker is spotted inside the sand screens after the open hoel gravel pack (OHGP). To complement this, the FCB design was deconstructed, and the in-situ breaker component was blended with the gravel. The concept was to incorporate the in-situ breaker component into the gravel pore space which would promote better diffusion of FCB through the reversible non-aqueous GPCP. The in-situ breaker component is inert to the carrier fluid until it is activated by the temperature and water posing no threat to the stability of the carrier fluid while pumping. The innovative approach was tested in the laboratory setting using ceramic disks and return to flow method to prove the concept before conducting an elaborate return permeability testing with the reservoir core plugs for the final validation. Return to flow method indicated that the novel approach could improve the results by at least 10% compared to the baseline test with no breaker application. In the return permeability tests with reservoir core plugs, the novel approach resulted in 76% of the initial permeability whereas the baseline test was only 50%. Both the tests with ceramic disks and full-sequence formation damage tests with actual reservoir cores highlighted the benefits of the novel approach for gravel packing with non-aqueous GPCF and post-gravel pack FCB scenario. Non-aqueous GPCFs are relatively new to the industry and no record of the filter cake breaker application in such environment exists. This novel approach makes the filter cake breaker application possible in non-aqueous environment and pushes the existing boundaries of filter cake breaker chemistries.

Naik et al. show that GATA3, Runx1, and E2A are essential for hierarchical assembly of a transcriptionally active RAG locus chromatin hub in CD4+CD8+ thymocytes. Signal-dependent down-regulation of RAG expression is associated with hub disassembly and depends on Ikaros., Expression of Rag1 and Rag2 is tightly regulated in developing T cells to mediate TCR gene assembly. Here we have investigated the molecular mechanisms governing the assembly and disassembly of a transcriptionally active RAG locus chromatin hub in CD4+CD8+ thymocytes. Rag1 and Rag2 gene expression in CD4+CD8+ thymocytes depends on Rag1 and Rag2 promoter activation by a distant antisilencer element (ASE). We identify GATA3 and E2A as critical regulators of the ASE, and Runx1 and E2A as critical regulators of the Rag1 promoter. We reveal hierarchical assembly of a transcriptionally active chromatin hub containing the ASE and RAG promoters, with Rag2 recruitment and expression dependent on assembly of a functional ASE–Rag1 framework. Finally, we show that signal-dependent down-regulation of RAG gene expression in CD4+CD8+ thymocytes depends on Ikaros and occurs with disassembly of the RAG locus chromatin hub. Our results provide important new insights into the molecular mechanisms that orchestrate RAG gene expression in developing T cells.

Amyotrophic lateral sclerosis (ALS) is a fatal motor neuron degenerative disease. TDP-43 (TAR DNA-binding protein 43) and FUS (fused in sarcoma) are aggregation-prone RNA-binding proteins that in ALS can mis-localize to the cytoplasm of affected motor neuron cells, often forming cytoplasmic aggregates in the process. Such mis-localization and aggregation are implicated in ALS pathology, though the mechanisms of TDP-43 and FUS cytoplasmic toxicity remains unclear. Recently, we determined that the endocytic function aids turnover of TDP-43 and reduces TDP-43 toxicity. Here, we identified that Cdc48 and Ubx3, a Cdc48 co-factor implicated in endocytic function, regulates the turnover and toxicity of TDP-43 and FUS expressed in S. cerevisiae. Cdc48 physically interacts and co-localizes with TDP-43, as does VCP in ALS patient tissue. In yeast, FUS toxicity also depends strongly on endocytic function, but not autophagy under normal conditions. FUS expression also impairs endocytic function, as previously observed with TDP-43. Taken together, our data identifies a role for Cdc48/VCP and endocytosis function in regulating TDP-43 and FUS toxicity and turnover. Furthermore, endocytic dysfunction may be a common defect affecting cytoplasmic clearance of ALS aggregation-prone proteins and may represent a novel therapeutic target of promise.

Summary: Tcrb locus V(D)J recombination is regulated by positioning at the nuclear periphery. Here, we used DamID to profile Tcrb locus interactions with the nuclear lamina at high resolution. We identified a lamina-associated domain (LAD) border composed of several CTCF-binding elements that segregates active non-LAD from inactive LAD regions of the locus. Deletion of the LAD border causes an enhancer-dependent spread of histone H3 lysine 27 acetylation from the active recombination center into recombination center-proximal LAD chromatin. This is associated with a disruption to nuclear lamina association, increased chromatin looping to the recombination center, and increased transcription and recombination of recombination center-proximal gene segments. Our results show that a LAD and LAD border are critical components of Tcrb locus gene regulation and suggest that LAD borders may generally function to constrain the activity of nearby enhancers. : Chen et al. identify a Tcrb locus lamina-associated domain border that constrains the activity of the Tcrb enhancer. Deletion of the border causes enhancer-dependent loss of nuclear lamina association, spreading of H3K27 acetylation, and elevated transcription and VDJ recombination of gene segments in affected chromatin. Keywords: T cell receptor β, Tcrb, V(D)J recombination, nuclear lamina, lamina-associated domain, LAD border, DamID, CTCF

The anticholinesterase paraoxon (Pxn) is related to military nerve agents that increase acetylcholine levels, trigger seizures, and cause excitotoxic damage in the brain. In rat hippocampal slice cultures, high-dose Pxn was applied resulting in a presynaptic vulnerability evidenced by a 64% reduction in synapsin IIb (syn IIb) levels, whereas the postsynaptic protein GluR1 was unchanged. Other signs of Pxn-induced cytotoxicity include the oxidative stress-related production of stable 4-hydroxynonenal (4-HNE)-protein adducts. Next, the Pxn toxicity was tested for protective effects by the fatty acid amide hydrolase (FAAH) inhibitor AM5206, a compound linked to enhanced repair signaling through the endocannabinoid pathway. The Pxn-mediated declines in syn IIb and synaptophysin were prevented by AM5206 in the slice cultures. To test if the protective results in the slice model translate to an in vivo model, AM5206 was injected i.p. into rats, followed immediately by subcutaneous Pxn administration. The toxin caused a pathogenic cascade initiated by seizure events, leading to presynaptic marker decline and oxidative changes in the hippocampus and frontal cortex. AM5206 exhibited protective effects including the reduction of seizure severity by 86%, and improving balance and coordination measured 24 h post-insult. As observed in hippocampal slices, the FAAH inhibitor also prevented the Pxn-induced loss of syn IIb in vivo. In addition, the AM5206 compound reduced the 4-HNE modifications of proteins and the β1 integrin activation events both in vitro and in vivo. These results indicate that Pxn exposure produces oxidative and synaptic toxicity that leads to the behavioral deficits manifested by the neurotoxin. In contrast, the presence of FAAH inhibitor AM5206 offsets the pathogenic cascade elicited by the Pxn anticholinesterase.