An ATP13A1-assisted topogenesis pathway for folding multi-spanning membrane proteins.

Many multi-spanning membrane proteins contain poorly hydrophobic transmembrane domains (pTMDs) protected from phospholipid in mature structure. Nascent pTMDs are difficult for translocon to recognize and insert. How pTMDs are discerned and packed into mature, muti-spanning configuration remains unclear. Here, we report that pTMD elicits a post-translational topogenesis pathway for its recognition and integration. Using six-spanning protein adenosine triphosphate-binding cassette transporter G2 (ABCG2) and cultured human cells as models, we show that ABCG2's pTMD2 can pass through translocon into the endoplasmic reticulum (ER) lumen, yielding an intermediate with inserted yet mis-oriented downstream TMDs. After translation, the intermediate recruits P5A-ATPase ATP13A1, which facilitates TMD re-orientation, allowing further folding and the integration of the remaining lumen-exposed pTMD2. Depleting ATP13A1 or disrupting pTMD-characteristic residues arrests intermediates with mis-oriented and exposed TMDs. Our results explain how a "difficult" pTMD is co-translationally skipped for insertion and post-translationally buried into the final correct structure at the late folding stage to avoid excessive lipid exposure.

UBE4A catalyzes NRF1 ubiquitination and facilitates DDI2-mediated NRF1 cleavage.

The transcription factor nuclear factor erythroid 2 like 1 (NFE2L1 or NRF1) regulates constitutive and inducible expression of proteasome subunits and assembly chaperones. The precursor of NRF1 is integrated into the endoplasmic reticulum (ER) and can be retrotranslocated from the ER to the cytosol where it is processed by ubiquitin-directed endoprotease DDI2. DDI2 cleaves and activates NRF1 only when NRF1 is highly polyubiquitinated. It remains unclear how retrotranslocated NRF1 is primed with large amount of ubiquitin and/or very long polyubiquitin chain for subsequent processing. Here, we report that E3 ligase UBE4A catalyzes ubiquitination of retrotranslocated NRF1 and promotes its cleavage. Depletion of UBE4A reduces the amount of ubiquitin modified on NRF1, shortens the average length of polyubiquitin chain, decreases NRF1 cleavage efficiency and causes accumulation of non-cleaved, inactivated NRF1. Expression of a UBE4A mutant lacking ligase activity impairs the cleavage, likely due to a dominant negative effect. UBE4A interacts with NRF1 and the recombinant UBE4A can promote ubiquitination of retrotranslocated NRF1 in vitro. In addition, knocking out UBE4A reduces transcription of proteasomal subunits in cells. Our results indicate that UBE4A primes NRF1 for DDI2-mediated activation to facilitate expression of proteasomal genes.

Effect of Transient Wave Forcing on the Behavior of Arsenic in a Nearshore Aquifer.

Groundwater-coastal water interactions influence the fate of inorganic chemicals in nearshore aquifers and their flux to receiving coastal waters. This study evaluates the impact of variable wave conditions on the geochemical changes and distribution of mobile arsenic (As) in a nearshore aquifer. Field measurements in a sandy nearshore aquifer on Lake Erie, Canada, are presented with geochemical changes analyzed over a period of high waves. A numerical model of wave-induced groundwater flow dynamics, validated against field data, is used to provide insight into the physical flow processes underlying the observed geochemical changes. Rapid changes in dissolved As, Fe, Mn, and S demonstrate the importance of reactions as well as dynamic transport in controlling the behavior of reactive species, especially those that are redox sensitive. Field data suggest the presence of sediment traps, which under certain hydrological and geochemical conditions may result in a "hot moment" with episodic release of As. The study provides new insight into factors controlling the fate of reactive species in dynamic coastal environments as required to better predict chemical fluxes to coastal waters. Additionally, it highlights the need to pay particular attention to "hot moments" for reaction and transport caused by storms and waves.

Effect of Low Energy Waves on the Accumulation and Transport of Fecal Indicator Bacteria in Sand and Pore Water at Freshwater Beaches.

Elevated fecal indicator bacteria (FIB) in beach sand and pore water represent an important nonpoint source of contamination to surface waters. This study examines the physical processes governing the accumulation and distribution of FIB in a beach aquifer. Field data indicate E. coli and enterococci can be transported 1 and 2 m, respectively, below the water table. Data were used to calibrate a numerical model whereby FIB are delivered to a beach aquifer by wave-induced infiltration across the beach face. Simulations indicate FIB rapidly accumulate in a beach aquifer with FIB primarily associated with sand rather than freely residing in the pore water. Simulated transport of E. coli in a beach aquifer is complex and does not correlate with conservative tracer transport. Beaches with higher wave-induced infiltration rate and vertical infiltration velocity (i.e., beaches with higher beach slope and wave height, and lower terrestrial groundwater discharge) had greater E. coli accumulation and E. coli was transported deeper below the beach face. For certain beach conditions, the amount of FIB accumulated in sand over 5-6 days was found to be sufficient to trigger a beach advisory if eroded to surface water.

PHT3D-UZF: A Reactive Transport Model for Variably-Saturated Porous Media.

A modified version of the MODFLOW/MT3DMS-based reactive transport model PHT3D was developed to extend current reactive transport capabilities to the variably-saturated component of the subsurface system and incorporate diffusive reactive transport of gaseous species. Referred to as PHT3D-UZF, this code incorporates flux terms calculated by MODFLOW's unsaturated-zone flow (UZF1) package. A volume-averaged approach similar to the method used in UZF-MT3DMS was adopted. The PHREEQC-based computation of chemical processes within PHT3D-UZF in combination with the analytical solution method of UZF1 allows for comprehensive reactive transport investigations (i.e., biogeochemical transformations) that jointly involve saturated and unsaturated zone processes. Intended for regional-scale applications, UZF1 simulates downward-only flux within the unsaturated zone. The model was tested by comparing simulation results with those of existing numerical models. The comparison was performed for several benchmark problems that cover a range of important hydrological and reactive transport processes. A 2D simulation scenario was defined to illustrate the geochemical evolution following dewatering in a sandy acid sulfate soil environment. Other potential applications include the simulation of biogeochemical processes in variably-saturated systems that track the transport and fate of agricultural pollutants, nutrients, natural and xenobiotic organic compounds and micropollutants such as pharmaceuticals, as well as the evolution of isotope patterns.

Electrokinetic in situ oxidation remediation: assessment of parameter sensitivities and the influence of aquifer heterogeneity on remediation efficiency.

A newly developed groundwater and electrokinetic (EK) flow and reactive transport numerical model was applied to simulate electrokinetic in situ chemical oxidation (EK-ISCO) remediation. Scenario simulations that considered the oxidation of a typical organic contaminant (tetrachloroethene) by permanganate were used to gain a better understanding of the key processes and parameters that control remediation efficiency. In a first step a sensitivity analysis was carried out to investigate a range of EK, hydraulic and engineering parameters on the performance of EK-ISCO. While all investigated parameters affected the remediation process to some extent, the duration and energy required for remediation were shown to be most dependent upon the applied voltage gradient, the natural oxidant demand and the concentration of the injected oxidant. Secondly, the efficacy of EK-induced oxidant transport was further examined for a heterogeneous aquifer system with random permeability fields. Oxidant migration under EK was slower in low-permeability media due to the increased oxidant consumption of competing reductants. Instead of injecting oxidant only at the cathode, locating injection wells between the electrodes greatly increased the contaminant degradation by decreasing the distance the amendment had to migrate before reaching the contaminant.