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At least 199 records · Page 11

ClusterCAD 2.0: an updated computational platform for chimeric type I polyketide synthase and nonribosomal peptide synthetase design

Abstract Megasynthase enzymes such as type I modular polyketide synthases (PKSs) and nonribosomal peptide synthetases (NRPSs) play a central role in microbial chemical warfare because they can evolve rapidly by shuffling parts (catalytic domains) to produce novel chemicals. If we can understand the design rules to reshuffle these parts, PKSs and NRPSs will provide a systematic and modular way to synthesize millions of molecules including pharmaceuticals, biomaterials, and biofuels. However, PKS and NRPS engineering remains difficult due to a limited understanding of the determinants of PKS and NRPS fold and function. We developed ClusterCAD to streamline and simplify the process of designing and testing engineered PKS variants. Here, we present the highly improved ClusterCAD 2.0 release, available at https://clustercad.jbei.org. ClusterCAD 2.0 boasts support for PKS-NRPS hybrid and NRPS clusters in addition to PKS clusters; a vastly enlarged database of curated PKS, PKS-NRPS hybrid, and NRPS clusters; a diverse set of chemical ‘starters’ and loading modules; the new Domain Architecture Cluster Search Tool; and an offline Jupyter Notebook workspace, among other improvements. Together these features massively expand the chemical space that can be accessed by enzymes engineered with ClusterCAD.

59 BASIC BIOLOGICAL SCIENCES↗

A novel bivalent chromatin associates with rapid induction of camalexin biosynthesis genes in response to a pathogen signal in Arabidopsis

Temporal dynamics of gene expression underpin responses to internal and environmental stimuli. In eukaryotes, regulation of gene induction includes changing chromatin states at target genes and recruiting the transcriptional machinery that includes transcription factors. As one of the most potent defense compounds in Arabidopsis thaliana , camalexin can be rapidly induced by bacterial and fungal infections. Though several transcription factors controlling camalexin biosynthesis genes have been characterized, how the rapid activation of genes in this pathway upon a pathogen signal is enabled remains unknown. By combining publicly available epigenomic data with in vivo chromatin modification mapping, we found that camalexin biosynthesis genes are marked with two epigenetic modifications with opposite effects on gene expression, trimethylation of lysine 27 of histone 3 (H3K27me3) (repression) and acetylation of lysine 18 of histone 3 (H3K18ac) (activation), to form a previously uncharacterized type of bivalent chromatin. Mutants with reduced H3K27me3 or H3K18ac suggested that both modifications were required to determine the timing of gene expression and metabolite accumulation at an early stage of the stress response. Our study indicates that the H3K27me3-H3K18ac bivalent chromatin, which we name as kairostat, plays an important role in controlling the timely induction of gene expression upon stress stimuli in plants.

59 BASIC BIOLOGICAL SCIENCES↗

Dexmedetomidine preconditioning ameliorates lung injury induced by pulmonary ischemia/reperfusion by upregulating promoter histone H3K4me3 modification of KGF-2

Highlights: • DexP alleviates I/R-induced lung injury and endothelial barrier dysfunction in mice. • DexP reduces the inflammatory response and increases KGF-2 expression in I/R. • DexP regulates the H3K4me3 modification of KGF-2 promoter histone. • DexP promotes KGF-2 expression by downregulating the expression of JMJD3. • DexP has the potential to be used as a means of treating I/R-induced lung injury. Keratinocyte growth factor (KGF)-2 has been highlighted to play a significant role in maintaining the endothelial barrier integrity in lung injury induced by ischemia-reperfusion (I/R). However, the underlying mechanism remains largely unknown. The aims of this study were to determine whether dexmedetomidine preconditioning (DexP) modulates pulmonary I/R-induced lung injury through the alteration in KGF-2 expression. In our I/R-modeled mice, DexP significantly inhibited pathological injury, inflammatory response, and inflammatory cell infiltration, while promoted endothelial barrier integrity and KGF-2 promoter activity in lung tissues. Bioinformatics prediction and ChIP-seq revealed that I/R significantly diminished the level of H3K4me3 modification in the KGF-2 promoter, which was significantly reversed by DexP. Moreover, DexP inhibited the expression of histone demethylase JMJD3, which in turn promoted the expression of KGF-2. In addition, overexpression of JMJD3 weakened the protective effect of DexP on lung injury in mice with I/R. Collectively, the present results demonstrated that DexP ameliorates endothelial barrier dysfunction via the JMJD3/KGF-2 axis.

60 APPLIED LIFE SCIENCES↗

Geologic Mapping of the Meridiani Region, Mars

The light toned bedrock that has been observed at the Mars Exploration Rover Opportunity landing site is an upper layer in a sequence >600 m thick in places. These outcrops contain mineral and textural signatures that require interaction of, and possibly formation from, water. Many distinct layers are visible in the remote sensing data (e.g. Figure 1) and no work has ever characterized the full set of these materials that cover an area >3 105 km2 spanning 20 of longitude. Thus, whatever water-related process( es?) altered, and possibly formed, the rocks at the Opportunity landing site extended over a vast region of Mars. Yet many questions remain to be answered, such as: (1) in what capacity did water form and alter the deposits?, (2) what are the temporal and spatial relations with other major events known from ancient Mars?, and (3) would this type of environment have been conducive to the development of life? To address these questions we are completing a detailed geologic, stratigraphic, and thermophysical properties study of this widespread terrain. Specifically, we are drafting a 1:2M-scale geological map covering the full extent of these water-related deposits. In tandem with the mapping, Hynek and Phillips [1] have conducted a preliminary stratigraphic analysis of the stack of materials. After mapping is complete, we will study the thermophysical properties of the varied layers to derive possible compositional information of the materials. These tasks serve several purposes including gaining an understanding of the complex nature of these materials, their potential source region(s), and their timing of emplacement. All of these efforts are necessary to place the observations by the Opportunity Rover in a broader context and prepare for potential future landed missions to the region. Understanding the large-scale paleohydrology of Mars is central to NASA s goals and vital for determining if life ever arose on the planet.

Hynek, B. M.↗

Nutrient Stability in NASA Spaceflight Experiment Rodent Food Bars

Nutrient-upgraded Rodent Food Bars are the standard diet for mice involved in spaceflight experiments aboard the International Space Station as part of NASA’s Rodent Research Project. These Food Bars are a twin-screw extruded AIN-93G based product with nutrient additives to preserve a healthy and complete diet for the animals, and the bar form factor is necessary for use in the microgravity environment. Due to the nature of NASA flight projects and the lengthy production process of these Food Bars, a shelf-life assessment was undertaken to determine nutrient stability over an extended storage time. To assess palatability and stability over an extended duration, a panel of lipid oxidation markers and both fat- and water-soluble vitamins were assessed under various time and temperature conditions via AOAC International standardized methodology. Thiamine was the least stable vitamin at room temperature. Bars that had been continually kept at 4C (t=6, 12, 27 months) showed significantly higher levels of thiamine (via Student’s t-test) compared to all other bars that underwent some degree of ambient temperature storage. This suggests that refrigeration is a key component in maintaining adequate nutrient levels in these relatively high moisture (average 29.3%) food bars. Also of note was the level of Vitamin A in the form of retinol. There was no correlation to storage temperature, but rather the only factor having an effect on retinol levels was time. As low, yet consistent decrease in retinol was observed over the course of the experiment, though at no time did the retinol levels drop below the rodent Nutrient Requirements for Laboratory Animals (NRC). In conclusion, over the 27-month time course, all analyzed vitamins and lipid oxidation factors remained relatively stable and at levels which provide fully sufficient nutrition to rodents during spaceflight. Some groups however, displayed higher-than-expected aerobic and anaerobic bacteria colony forming units, which is believed to stem from a since-revised handling technique.

Food↗

Substrate stiffness modulates endothelial cell function via the YAP-Dll4-Notch1 pathway

Highlights: • Soft stiffness increases Dll4 expression in endothelial cells via YAP suppression. • Soft substrate-induced Dll4-Notch1 signaling is facilitated by VEGF stimulation. • Elevated Dll4-Notch1 signaling contributes to the regulation of VEGFRs expression. • Substrate stiffness modulates gene expression linked to endothelial cell function. Endothelial cells adapt their functions as a consequence of sensing extracellular substrate stiffness; these alterations allow them to maintain their vascular structure and function. Substrate stiffness-mediated yes-associated protein 1 (YAP) activation plays an important role in mechano-transduction and pro-angiogenic phenotype of endothelial cells, and Delta-like ligand 4 (Dll4)-Notch1 signaling is closely related to angiogenesis; however, the impact of substrate stiffness-mediated interrelation of these pathways on endothelial cell functions remains elusive. We confirmed that endothelial cells on softer substrates not only elongate cellular aspects but also attenuate YAP activation compared to cells on stiffer substrates. Endothelial cells on softer substrates also upregulate the vascular endothelial growth factor receptor 1 (VEGFR1) and VEGFR2 mRNA expression that is enhanced by VEGF stimulation. We determined that endothelial cells on softer substrates increased Dll4 expression, but not Notch1 expression, via YAP signaling. Moreover, endothelial cells on soft substrates induced not only VEGFRs upregulation but also suppression of pro-inflammatory interleukin-6 and plasminogen activator inhibitor-1 mRNA expression and the facilitation of anti-coagulant thrombomodulin and pro-coagulant tissue factor mRNA expression. Our results suggest that endothelial cells activate the YAP-Dll4-Notch signaling pathway in response to substrate stiffness and dictate cellular function.

60 APPLIED LIFE SCIENCES↗

The Collection, Usage, and Preliminary Examination of the Apollo Sample Suite: Lessons for Artemis

Apollo Sample Collection and Usage: From 1969 to 1972 there were six Apollo missions to the surface of the Moon during which the astronauts collected 382 kg of rock and regolith (~2200 samples). The samples collected fall into these general categories: rocks (~66% by mass), rake samples (~4%), bulk regolith (~24%), and specialty regolith (deep drill cores, drive tubes, sealed bulk regolith) samples (~6%). In each category there are a variety of different subtypes available for study, e.g., among the bulk regolith samples there are also skim, trench, and (partially) shaded regolith samples each sampling unique types or depths of regolith. This variety of subsamples has enabled a multitude of different studies over the past 55 years (>3400 individual requests). We are still averaging ~50 unique requests and have allocated >500 individual Apollo samples annually for the past 10 years (2020 excepted). Looking at the 4,675 non-ANGSA (Apollo Next Generation Sample Analysis) samples allocated over the past 10 years, the proportions of allocated samples do not precisely align with the abundance (by mass) of those samples withing the collection: Rock (69.4 %); Rake (10.8 %); Bulk Regolith (15.5 %); Drive Tube (3.3 %); Core/Specialty (1.0 %). Apollo Preliminary Examination (PE): The PE process differs significantly for the various sample types enumerated above; we focus on regolith and rock samples here. During the Apollo mission era, the PE process evolved over the course of the missions; below is what was done for the Apollo 17 mission. For regolith samples, the PE process was: (1) documented bags containing regolith are opened, photographed, and described; (2) large rocks are removed and treated separately; (3) 25% to 33% of the bulk soil is scooped out, weighed, and stored in reserve; (4) the remaining sample is sieved to produce the size fractions <1, 1- 2, 2-4, and 4-10 mm, all of which are weighed. For rock samples, the process is: (1) removing rocks from the container(s) it came back from the Moon in; (2) rematching any materials that spalled off the rock to their original location; (3) numbering, weighing, and basic photographic documentation; (4) dusting with a gentle N2 gas jet; (5) Orthogonal photography; (6) detailed description of the textures and features of the rock; (7) rock modelling and measurement; (8) stereophotography; (9) determination of the orientation of the rock on the lunar surface. Drive tubes and deep drill cores were not characterized during PE beyond an initial weight and a sketch of the interior tube materials derived from 2D medical X-ray images. Catalogs: The ongoing utility of the Apollo samples is enabled by the robust cataloguing process for the samples [3-5], which allows the scientific community to accurately request samples uniquely suited to their proposed studies. A common misconception, however, is the amount of detail that goes into the initial catalog (e.g., [6]) for a collection from the preliminary examination (PE) period, versus what goes into the catalogs that come later in the life cycle of the samples from that mission (e.g., [7]). The only required data for a PE catalog is a weight, a basic photograph, and a description of the nature of the sample. Artemis PE: Over the past few years, the ongoing ANGSA project studied previously unopened Apollo 17 double drive tube samples 73001/2 [1], and a PE of the drive tubes was done. The PE took the existing core dissection process (developed during PE of Apollo cores in the 1970s, 1980s), and modernized it [7]. The main lesson from the ANGSA PE relative to future missions was that the physical work done during PE of lunar samples has not changed much over the past 5 decades. The use of “modern” technology during PE (e.g., XCT; multispectral analyses) resulted in an enhanced initial understanding of the 73001 and 73002 drive tubes, but greatly increased the time required. The lessons learned from recent astromaterial PEs (e.g., ANGSA and OREx) are important to consider when planning for Artemis, but the unique nature of the Artemis Campaign means many lessons learned from these mission will not be applicable. Given the time constraints (6 months) and likely number of samples that will be returned by Artemis (>200), the Artemis PE catalog will necessarily look much more like [4] than [6].

J Gross↗

Next-generation poly-L-histidine formulations for miRNA mimic delivery

Many diseases, especially cancer, are caused by the abnormal expression of non-coding microRNAs (miRNAs), which regulate gene expression, leading to the development of miRNA-based therapeutics. Synthetic miRNA inhibitors have shown promising efficacy in blocking the activity of aberrant miRNAs that are upregulated in disease-specific pathologies. On the other hand, miRNAs that aid in preventing certain diseases and are reduced in expression in the disease state need different strategies. To tackle this, miRNA mimics, which mimic the activity of endogenous miRNAs, can be delivered for those miRNAs downregulated in different disease states. However, the delivery of miRNA mimics remains a challenge. Here, we report a cationic polylactic-co-glycolic acid (PLGA)-poly-L-histidine delivery system to deliver miRNA mimics. We chose miR-34a mimics as a proof of concept for miRNA delivery. miR-34a-loaded PLGA-poly-L-histidine nanoparticles (NPs) were formulated and biophysically characterized to analyze the structural properties of miRNA mimic-loaded NPs. In vitro efficacy was determined by investigating miR-34a and downstream target levels and performing cell viability and apoptosis assays. We confirmed in vivo efficacy through prolonged survival of miR-34a NP-treated A549-derived xenograft mice treated intratumorally. The results of these studies establish PLGA-poly-L-histidine NPs as an effective delivery system for miRNA mimics for treating diseases characterized by downregulated miRNAs.

60 APPLIED LIFE SCIENCES↗

A Proposal for an Integrated Geophysical Strategy to "Follow the Water" on Mars

The search for subsurface water has become a primary focus of Mars exploration. Its abundance and distribution (both as ground ice and groundwater) have important implications for understanding the geologic, hydrologic, and climatic evolution of the planet; the potential origin and continued survival of life; and the accessibility of a critical in situ resource for sustaining future human explorers. For these reasons, a principal goal of the Mars science, astrobiology, and the HEDS programs is to determine the 3-D distribution and state of subsurface H2O, at a resolution sufficient to permit reaching any desired volatile target by drilling. The three targets most often discussed are: groundwater, massive deposits of near-surface ground ice (associated with the ponded discharge of the outflow channels or the relic of a former ocean), and ice-saturated frozen ground. Based on the present best estimates of mean annual surface temperature, crustal thermal conductivity, geothermal heat flow, and groundwater freezing temperature, the mean thickness of frozen ground on Mars is expected to vary from approx. = 2.5 - 5 km at the equator to approx. = 6.5 - 13 km at the poles. However, natural variations in both crustal heat flow and thermal conductivity are likely to result in significant local departures from these predicted values. The recent discovery of "young" fluvial-like features, emanating from the slopes of local scarps, raises the possibility that liquid water may also exist episodically at shallow (approx. = 100 - 500 m) depth; however, the true nature and absolute age of these features remains highly uncertain. Although the belief that Mars is water-rich is supported by a wide variety geologic evidence, our ignorance about the heterogeneous nature and thermal evolution of the planet's crust effectively precludes geomorphic or theoretical attempts to quantitatively assess the current geographic and subsurface vertical distribution of ground ice and groundwater . For this reason, any exploration activity (such as drilling) whose success is contingent on the presence of subsurface water, must be preceded by a comprehensive high-resolution geophysical survey capable of assessing whether local reservoirs of water and ice actually exist. Terrestrial experience has demonstrated that the accurate identification of such targets is likely to require the application of multiple geophysical techniques. In this abstract we propose an integrated strategy for the geophysical exploration of Mars that we believe represents the fastest, most cost-effect, and technically capable approach to identifying the state and distribution of subsurface water. Additional information is contained in the original extended abstract.

Clifford, S. M.↗

Structural basis for self-discrimination by neoantigen-specific TCRs

T cell receptors (TCR) are pivotal in mediating tumour cell cytolysis via recognition of mutation-derived tumour neoantigens (neoAgs) presented by major histocompatibility class-I (MHC-I). Understanding the factors governing the emergence of neoAg from somatic mutations is a major focus of current research. However, the structural and cellular determinants controlling TCR recognition of neoAgs remain poorly understood. This study describes the multi-level analysis of a model neoAg from the B16F10 murine melanoma, H2-D b /Hsf2 p.K72N 68-76 , as well as its cognate TCR 47BE7. Through cellular, molecular and structural studies we demonstrate that the p.K72N mutation enhances H2-D b binding, thereby improving cell surface presentation and stabilizing the TCR 47BE7 epitope. Furthermore, TCR 47BE7 exhibited high functional avidity and selectivity, attributable to a broad, stringent, binding interface enabling recognition of native B16F10 despite low antigen density. Our findings provide insight into the generation of anchor-residue modified neoAg, and emphasize the value of molecular and structural investigations of neoAg in diverse MHC-I contexts for advancing the understanding of neoAg immunogenicity.

60 APPLIED LIFE SCIENCES↗

Multisubstrate specificity shaped the complex evolution of the aminotransferase family across the tree of life

Aminotransferases (ATs) are an ancient enzyme family that play central roles in core nitrogen metabolism, essential to all organisms. However, many of the AT enzyme functions remain poorly defined, limiting our fundamental understanding of the nitrogen metabolic networks that exist in different organisms. Here, we traced the deep evolutionary history of the AT family by analyzing AT enzymes from 90 species spanning the tree of life (ToL). We found that each organism has maintained a relatively small and constant number of ATs. Mapping the distribution of ATs across the ToL uncovered that many essential AT reactions are carried out by taxon-specific AT enzymes due to wide-spread nonorthologous gene displacements. This complex evolutionary history explains the difficulty of homology-based AT functional prediction. Biochemical characterization of diverse aromatic ATs further revealed their broad substrate specificity, unlike other core metabolic enzymes that evolved to catalyze specific reactions today. Interestingly, however, we found that these AT enzymes that diverged over billion years share common signatures of multisubstrate specificity by employing different nonconserved active site residues. These findings illustrate that AT family enzymes had leveraged their inherent substrate promiscuity to maintain a small yet distinct set of multifunctional AT enzymes in different taxa. This evolutionary history of versatile ATs likely contributed to the establishment of robust and diverse nitrogen metabolic networks that exist throughout the ToL. The study provides a critical foundation to systematically determine diverse AT functions and underlying nitrogen metabolic networks across the ToL.

59 BASIC BIOLOGICAL SCIENCES↗

Extracellular matrix controls tubulin monomer levels in hepatocytes by regulating protein turnover

Cells have evolved an autoregulatory mechanism to dampen variations in the concentration of tubulin monomer that is available to polymerize into microtubules (MTs), a process that is known as tubulin autoregulation. However, thermodynamic analysis of MT polymerization predicts that the concentration of free tubulin monomer must vary if MTs are to remain stable under different mechanical loads that result from changes in cell adhesion to the extracellular matrix (ECM). To determine how these seemingly contradictory regulatory mechanisms coexist in cells, we measured changes in the masses of tubulin monomer and polymer that resulted from altering cell-ECM contacts. Primary rat hepatocytes were cultured in chemically defined medium on bacteriological petri dishes that were precoated with different densities of laminin (LM). Increasing the LM density from low to high (1-1000 ng/cm2), promoted cell spreading (average projected cell area increased from 1200 to 6000 microns2) and resulted in formation of a greatly extended MT network. Nevertheless, the steady-state mass of tubulin polymer was similar at 48 h, regardless of cell shape or ECM density. In contrast, round hepatocytes on low LM contained a threefold higher mass of tubulin monomer when compared with spread cells on high LM. Furthermore, similar results were obtained whether LM, fibronectin, or type I collagen were used for cell attachment. Tubulin autoregulation appeared to function normally in these cells because tubulin mRNA levels and protein synthetic rates were greatly depressed in round cells that contained the highest level of free tubulin monomer. However, the rate of tubulin protein degradation slowed, causing the tubulin half-life to increase from approximately 24 to 55 h as the LM density was lowered from high to low and cell rounding was promoted. These results indicate that the set-point for the tubulin monomer mass in hepatocytes can be regulated by altering the density of ECM contacts and changing cell shape. This finding is consistent with a mechanism of MT regulation in which the ECM stabilizes MTs by both accepting transfer of mechanical loads and altering tubulin degradation in cells that continue to autoregulate tubulin synthesis.

Non-NASA Center↗

Inflatable Softgoods Testing Capability Development for Habitation

The Habitation Systems Development Office (HP40) at NASA Marshall Space Flight Center supports systems engineering, integration, and project management for next generation space habitats. The office is responsible for formulation activities for Lunar Surface Habitat and Mars Transit Habitat. HP40 also manages commercial partnerships for exploration habitat development. The Lunar Surface Habitat is intended to initially support a minimum of two crew on the lunar surface for an approximately 30-day mission, with the ability to accommodate four crew for short durations. The Mars Transit Habitat will transport four crew to and from Mars orbit on a potentially 1,200 day mission. Both concepts are envisioned to leverage inflatable softgoods as the primary structural material for the habitation portion of the architecture. Inflatable softgoods are a complex, multi-material system consisting of multiple layers which perform specific functions. A bladder layer (polymer) contains the habitat’s atmosphere. The bladder is indexed to a woven restraint layer, usually Kevlar or Vectran, which serves as the structural layer. The remaining outer layers are for micro-meteoroid orbital debris (MMOD) shielding, protection from atomic Oxygen (if the material system is to be deployed in low earth orbit), and thermal insulation. The primary advantage of inflatable softgoods is that they can offer a larger volume per unit mass relative to a rigid metal structure. Inflatable softgoods have a strong development heritage within NASA, beginning with work in the 1960s in partnership with Goodyear Aerospace and continuing with Johnson Space Center’s TransHab project in the 1990s . Some development efforts subsequently transitioned into the private sector to mature the material technology for flight applications, including human habitation. In August 2022, NASA published its guidelines for certification of crewed inflatable softgoods structures (JSC-67721). This document outlines a series of tests which will be needed to certify inflatable softgoods material systems for use in crewed mission applications. It includes burst testing at the subscale and full-scale to determine the ultimate burst pressure of the softgoods architecture. Creep (long term deformation of the material under stress) is a key failure mode which must be characterized. Creep testing at the component and module levels is also recommended to generate time to failure curves and verify that the inflatable system has a predicted operational life which exceeds mission parameters. ET (Test Engineering) at MSFC and HP40 have worked together to establish softgoods testing capabilities for the center. This includes the ability to conduct module level burst tests and creep tests. The poster will highlight these capabilities and provide examples of testing which has been performed to date by ET. NASA MSFC anticipates that these test capabilities will continue to offer critical support for companies pursuing inflatable softgoods for exploration applications (lunar surface habitation, Mars transit) as well as commercial low earth orbit platforms.

habitation↗

Structural mechanisms for VMAT2 inhibition by tetrabenazine

The vesicular monoamine transporter 2 (VMAT2) is a proton-dependent antiporter responsible for loading monoamine neurotransmitters into synaptic vesicles. Dysregulation of VMAT2 can lead to several neuropsychiatric disorders including Parkinson’s disease and schizophrenia. Furthermore, drugs such as amphetamine and MDMA are known to act on VMAT2, exemplifying its role in the mechanisms of actions for drugs of abuse. Despite VMAT2’s importance, there remains a critical lack of mechanistic understanding, largely driven by a lack of structural information. Here, we report a 3.1 Å resolution cryo-electron microscopy (cryo-EM) structure of VMAT2 complexed with tetrabenazine (TBZ), a non-competitive inhibitor used in the treatment of Huntington’s chorea. We find TBZ interacts with residues in a central binding site, locking VMAT2 in an occluded conformation and providing a mechanistic basis for non-competitive inhibition. We further identify residues critical for cytosolic and lumenal gating, including a cluster of hydrophobic residues which are involved in a lumenal gating strategy. Our structure also highlights three distinct polar networks that may determine VMAT2 conformational dynamics and play a role in proton transduction. The structure elucidates mechanisms of VMAT2 inhibition and transport, providing insights into VMAT2 architecture, function, and the design of small-molecule therapeutics.

59 BASIC BIOLOGICAL SCIENCES↗

The Search for Chiral Asymmetry as a Potential Biosignature in Samples from Mars

The search for evidence of extraterrestrial life in our solar system is currently guided by our understanding of terrestrial biology and its associated biosignatures. The observed homochirality in all life on Earth, that is, the predominance of “left-handed” or L-amino acids and “right-handed” or D-sugars, is a unique property of life that is crucial for molecular recognition, enzymatic function, information storage and structure and is thought to be a prerequisite for the origin or early evolution of life. Therefore, the detection of L- or D-enantiomeric excesses of chiral amino acids and sugars could be a powerful indicator of extant or extinct life on Mars or other habitable environments in our solar system. The exploration of habitable environments on Mars, including an assessment of the preservation potential for complex organics of either abiotic or biological origin, remains a key goal of both current and future Mars missions. Now with the unambiguous detection of indigenous organic matter in sedimentary rocks on Mars [1-3], NASA’s Curiosity rover has significantly advanced our understanding of the preservation of potential chemical biosignatures in the martian near surface. Although amino acids have not yet been identified by in situ measurements on Mars, if they were present, it is expected that amino acid hydrolysis and racemization would be very slow and any chiral or isotopic signatures from an extinct martian biota could be preserved for billions of years, given the extremely cold and dry surface conditions [4]. One of ESA’s Rosalind Franklin rover payload instruments called the Mars Organic Molecule Analyzer (MOMA) includes a wet chemistry package capable of measuring the enantiomeric ratios of any chiral amino acids present at part-per-million concentrations or higher [5]. The complexity and limited duration of spaceflight operations and the known analytical challenges associated with in situ extraction and characterization of trace reduced organic compounds in ancient rocks make it challenging to determine the origins of martian organic matter found to date. Coordinated state-of-the-art laboratory measurements of returned samples from Mars that include spatially resolved chemical, mineralogical, and isotopic studies and molecule-specific isotopic and enantiomeric measurements will be required to firmly establish whether the complex organic matter detected on Mars comes from biotic or abiotic sources. Ultimately, sample return may be our best chance of identifying chemical biosignatures from a past or present martian biota, if one ever existed on Mars. NASA’s Perseverance rover will collect dozens of surface sample cores for possible future return to Earth by NASA and ESA. Here we review our current knowledge of the distributions and enantiomeric and isotopic compositions of non-biological amino acids found in meteorites compared to terrestrial biochemistry and propose a set of measurement criteria that should be used to help establish the origin of any chiral asymmetry detected in samples from Mars [6].

Daniel P Glavin↗

The mesoscale forcing of a midlatitude upper-tropospheric jet streak by a simulated convective system. 2: Kinetic energy and resolution analysis

A kinetic energy (KE) analysis of the forcing of a mesoscale upper-tropospheric jet streak by organized diabatic processes within the simulated convective system (SCS) that was discussed in Part 1 is presented in this study. The relative contributions of the ageostrophic components of motion to the generation of KE of the convectively generated jet streak are compared, along with the KE generation by the rotational (nondivergent) and irrotational (divergent) mass transport. The sensitivity of the numerical simulations of SCS development to resolution is also briefly examined. Analysis within isentropic coordinates provides for an explicit determination of the influence of the diabatic processes on the generation of KE. The upper-level production of specific KE is due predominatly to the inertial advective ageostrophic component (IAD), and as such represents the primary process through which the KE of the convectively generated jet streak is realized. A secondary contribution by the inertial diabatic (IDI) term is observed. Partitioning the KE generation into its rotational and irrotational components reveals that the latter, which is directly linked to the diabatic heating within the SCS through isentropic continuity requirements, is the ultimate source of KE generation as the global area integral of generation by the rotational component vanishes. Comparison with an identical dry simulation reveals that the net generation of KE must be attributed to latent heating. Both the IAD and IDI ageostrophic components play important roles in this regard. Examination of results from simulations conducted at several resolutions supports the previous findings in that the effects of diabatic processes and ageostrophic motion on KE generation remain consistent. Resolution does impact the location and timing of SCS development, a result that has important implications in forecasting the onset of convection that develops from evolution of the large-scale flow and moisture transport. Marked differences are observed in the momentum field aloft subsequent to the life cycle of the SCS in the 1 deg, 30-level base case (MP130) simulation discussed in Part 1 versus its 2 deg counterparts in that the MP130 simulation with higher spatial resolution contains from 14% to 30% more total KE.

Wolf, Bart J.↗

Radiation Synthesis of New Molecules on Jupiter's Icy Satellites

Spectra of Jupiter's icy satellites reveal surfaces dominated by water-ice, minor amounts of SO2 and CO2, and (for Europa) H2O2 along with hydrated materials. Jovian magnetospheric ions (protons, sulfur, and oxygen) and electrons significantly modify the chemical composition of these moons' surfaces in times ranging from a few years for Europa to thousands of years for Callisto at micrometer depths. Appropriate laboratory studies examining relevant volatile and non-volatile materials under low-temperature radiation conditions can provide information on likely radiation chemical mechanisms, on the stability and evolution of species, and on new species awaiting detection. Although the molecules detected on the icy moons are relatively simple, predicting their responses to radiation in space remains difficult. One problem is that there is a dearth of fundamental data examining solid-phase reactions. Our laboratory experiments have focused on infrared studies (2.5 to 25 microns) of a few simple irradiated ices. We have measured the spectra of proton-irradiated H2O ice containing SO2, H2S, and/or CO2. Ices with H2O/SO2 or H2O/H2S ratios of 3 and 30 have been irradiated at 86 K, 110 K, and 132 K. In irradiated H2O + SO2 ices new ions have been identified: SO4(-2), HSO4(-) and H3O(+). After warming to 260 K the residual spectrum is similar to that of H2SO4. Ices with H2O + H2S form SO2. After warming to 175 K, the residual sample matches the spectrum of hydrated H2SO4. H2O + CO2 ice forms carbonic acid, H2CO3 which is stable to temperatures near 230 K. In addition, OCS has been detected in irradiated ices containing H2O + SO2 + CO2. The radiation half-life of SO2 and H2S in H2O has been calculated. Our results give compelling evidence for the presence of new species awaiting detection. Future experiments will examine the signatures of these ices and hydrated materials in the 1 to 5 micron region, where possible weaker overtone bands may occur. In addition, absolute strengths for both the fundamental and overtone bands will be determined. Finally, good arguments can be made, based on current information, for remote sensing observations that have spectral coverage to at least 5 microns on the long wavelength end. This range would include some of the characteristic bands of H2O, H2O2, CO2, SO2, H2CO3, H2S, and OCS.

Moore, M. H.↗

A Simulated High CO 2 Spaceflight Environment Increases Plant Preference for Ammonium as a Nitrogen Source

Long-duration exploration missions will require a sustainable supply of food to support human crews. The spaceflight environment contains high concentrations of CO 2 due to release by astronauts that cannot be completely scrubbed. It is therefore crucial to understand plant responses to elevated CO 2 (eCO 2 ) environments. Nitrogen (N) is crucial for plant survival, though the effects of eCO 2 on plant N uptake remain poorly understood. Shoot nitrate reduction may be reduced at eCO 2 , due to reduced reductant availability for nitrate reduction due to reduced photorespiration and increased carbon fixation. Relative growth rate may be reduced at eCO 2 when N is provided only as nitrate and can be unaffected when N is supplied as ammonium, suggesting eCO 2 may drive increased plant ‘preference’ for ammonium. However, changes in N preference in response to eCO 2 have, to our knowledge, not yet been studied. In this study, novel stable isotope approaches were used in conjunction with hydroponics and isotope ratio mass spectrometry to determine the effect of space station-like eCO 2 (3000 ppm) on preference for ammonium or nitrate in several lettuce varieties previously grown in space, when both N forms are provided equally. All varieties displayed significant ammonium preference irrespective of CO 2 concentration, however the extent of this preference varied. Increased ammonium preference was observed for all varieties at eCO 2 compared to ambient CO 2 (410 ppm), driven by increases in nitrogen uptake which plants disproportionately took up as ammonium. These results suggest that future nutrient formulations should favor ammonium as a major N source for space crop production. Moreover, increased ammonium preference may play a role in future plant-based bioregenerative life support systems with higher ammonium concentrations due to waste recycling. Additionally, this research furthers our understanding of plant responses to future high CO 2 climates, allowing the development of future-proof crops to maintain food security.

space crop production↗