Search NASA⌕ Search

SEARCH · Search NASA

Results for “cellular degradation”

Search indexed NASA NTRS and DOE OSTI research on propulsion, heat transfer, battery materials and energy systems. Follow report and document links to the original sources.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 73 records · Page 4
All sourcesNASA NTRSDOE OSTI
DOE OSTI2021

Degradation of aromatic compounds and lignin by marine protist Thraustochytrium striatum

This paper aimed at studying a novel high biomass strain (HBS) of marine protist, Thraustochytrium striatum, for lignin valorization to lipids and carotenoids. In the screening of 15 aromatic compounds, T. striatum HBS can utilize most compounds while achieving the best growth on 3,4-dihydroxybenzoic acid (3,4-DHBA) and 4-hydroxybenzoic acid (4-HBA). The maximum cell mass concentration reached 2-3 g/L under 5 g/L of 3,4-DHBA or 2 g/L of 4-HBA, which was comparable to that under equivalent concentration of glucose. The long-chain fatty acids content was 13 % and 7 % with 3,4-DHBA and 4-HBA, respectively. The adoption of fed-batch fermentation mode improved the degradation of carbon source and prevented the inhibitory effect caused by overloading of aromatic compounds. Further, both final cellular fatty acids and carotenoids contents were doubled in the fermentation of 3,4-DHBA. T. striatum HBS exhibited limited lignin depolymerization (~24 %) ability on technical lignins.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2020

Large fluxes of fatty acids from membranes to triacylglycerol and back during N-deprivation and recovery in Chlamydomonas

Abstract Microalgae accumulate triacylglycerol (TAG) during nutrient deprivation and break it down after nutrient resupply, and these processes involve dramatic shifts in cellular carbon allocation. Due to the importance of algae in the global carbon cycle, and the potential of algal lipids as feedstock for chemical and fuel production, these processes are of both ecophysiological and biotechnological importance. However, the metabolism of TAG is not well understood, particularly the contributions of fatty acids (FAs) from different membrane lipids to TAG accumulation and the fate of TAG FAs during degradation. Here, we used isotopic labeling time course experiments on Chlamydomonas reinhardtii to track FA synthesis and transfer between lipid pools during nitrogen (N)-deprivation and resupply. When cells were labeled before N-deprivation, total levels of label in cellular FAs were unchanged during subsequent N-deprivation and later resupply, despite large fluxes into and out of TAG and membrane lipid pools. Detailed analyses of FA levels and labeling revealed that about one-third of acyl chains accumulating in TAG during N-deprivation derive from preexisting membrane lipids, and in total, at least 45% of TAG FAs passed through membrane lipids at one point. Notably, most acyl chains in membrane lipids during recovery after N-resupply come from TAG. Fluxes of polyunsaturated FAs from plastidic membranes into TAG during N-deprivation were particularly noteworthy. These findings demonstrate a high degree of integration of TAG and membrane lipid metabolism and highlight a role for TAG in storage and supply of membrane lipid components.

Plant Sciences↗
DOE OSTI2023

Expression and characterization of spore coat Cot$\mathrm{H}$ kinases from the cellulosomes of anaerobic fungi ( Neocallimastigomycetes )

Anaerobic fungi (Neocallimastigomycetes) found in the guts of herbivores are biomass deconstruction specialists with a remarkable ability to extract sugars from recalcitrant plant material. Anaerobic fungi, as well as many species of anaerobic bacteria, deploy multi-enzyme complexes called cellulosomes, which modularly tether together hydrolytic enzymes, to accelerate biomass hydrolysis. While the majority of genomically encoded cellulosomal genes in Neocallimastigomycetes are biomass degrading enzymes, the second largest family of cellulosomal genes encode spore coat CotH domains, whose contribution to fungal cellulosome and/or cellular function is unknown. Structural bioinformatics of CotH proteins from the anaerobic fungus Piromyces finnis shows anaerobic fungal CotH domains conserve key ATP and Mg 2+ binding motifs from bacterial Bacillus CotH proteins known to act as protein kinases. Experimental characterization further demonstrates ATP hydrolysis activity in the presence and absence of substrate from two cellulosomal P. finnis CotH proteins when recombinantly produced in E. coli. These results present foundational evidence for CotH activity in anaerobic fungi and provide a path towards elucidating the functional contribution of this protein family to fungal cellulosome assembly and activity.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

Conformational heterogeneity in the dGsw purine riboswitch: role of Mg²⁺ and 2’-dG in aptamer folding

Recent advancements in RNA structural biology have focused on unraveling the complexities of non-coding mRNA elements like riboswitches. These cis-acting regulatory regions undergo structural changes in response to specific cellular metabolites, leading to up or downregulation of downstream genes. The purine riboswitch family regulates many prokaryotic genes involved in purine degradation and biosynthesis. They feature an aptamer domain organized around a 3-way helical junction, where ligand encapsulation occurs at the junctional core. In our study, we chemically probed the aptamer domain of the 2’-dG-sensing purine riboswitch from Mesoplasma florum (dGsw) under various solution conditions to understand how Mg²⁺ and 2’-dG influence riboswitch folding. Here, we find that efficient 2’-dG binding strongly depends on Mg²⁺, indicating that Mg²⁺ is essential for priming dGsw for ligand interactions. We identified a previously undescribed sequence in the 5’ tail of dGsw that is complementary to a conserved helix. The inclusion of this region in a construct led to intramolecular competition between the alternate helix, Palt, and P1. Mutational analysis confirmed that 5’ flanking end of the aptamer domain forms an alternate helix in the absence of ligand. Molecular dynamics simulations revealed that this alternative conformation is stable. This helix may, therefore, facilitate the formation of an anti-terminator helix by opening the 3-way junction surrounding the 2’-dG binding site. Our study further establishes the importance of a closed terminal P1 helix conformation for metabolite binding and suggests that the delicate interplay between P1 and Palt may fine-tune downstream gene regulation. These insights offer a new perspective on riboswitch structure and enhance our understanding of the role that a conformational ensemble plays in riboswitch activity and regulation.

Biochemistry & Molecular Biology↗
NASA NTRS1990

Study on the spectral efficiency of SFH-GMSK in land mobile telephone communication by direct simulation

A spread spectrum system, slow frequency hopping with GMSK (Gaussian minimum shift keying) modulation (SFH-GMSK), is proposed for mobile telephone communications. The system performance is evaluated using computer simulation and is compared with an unspread system. Results show that under multipath fading conditions, when the signal-to-noise ratio (SNR) is greater than 15 dB, slow frequency hopping gives some bit error rate improvement over the unspread system. Theoretical predictions indicate that a system efficiency of 20-65 users per cell can be achieved in the cellular configuration. Joint use of SFH-GMSK and FM is also investigated. It is shown that FM interference can cause serious degradation to the SFH-GMSK performance.

Bao, H.↗
DOE OSTI2021

Enhancement of polyhydroxyalkanoate production by co-feeding lignin derivatives with glycerol in Pseudomonas putida KT2440

Abstract Background Efficient utilization of all available carbons from lignocellulosic biomass is critical for economic efficiency of a bioconversion process to produce renewable bioproducts. However, the metabolic responses that enable Pseudomonas putida to utilize mixed carbon sources to generate reducing power and polyhydroxyalkanoate (PHA) remain unclear. Previous research has mainly focused on different fermentation strategies, including the sequential feeding of xylose as the growth stage substrate and octanoic acid as the PHA-producing substrate, feeding glycerol as the sole carbon substrate, and co-feeding of lignin and glucose. This study developed a new strategy—co-feeding glycerol and lignin derivatives such as benzoate, vanillin, and vanillic acid in Pseudomonas putida KT2440—for the first time, which simultaneously improved both cell biomass and PHA production. Results Co-feeding lignin derivatives (i.e. benzoate, vanillin, and vanillic acid) and glycerol to P. putida KT2440 was shown for the first time to simultaneously increase cell dry weight (CDW) by 9.4–16.1% and PHA content by 29.0–63.2%, respectively, compared with feeding glycerol alone. GC–MS results revealed that the addition of lignin derivatives to glycerol decreased the distribution of long-chain monomers (C10 and C12) by 0.4–4.4% and increased the distribution of short-chain monomers (C6 and C8) by 0.8–3.5%. The 1 H– 13 C HMBC, 1 H– 13 C HSQC, and 1 H– 1 H COSY NMR analysis confirmed that the PHA monomers (C6–C14) were produced when glycerol was fed to the bacteria alone or together with lignin derivatives. Moreover, investigation of the glycerol/benzoate/nitrogen ratios showed that benzoate acted as an independent factor in PHA synthesis. Furthermore, 1 H, 13 C and 31 P NMR metabolite analysis and mass spectrometry-based quantitative proteomics measurements suggested that the addition of benzoate stimulated oxidative-stress responses, enhanced glycerol consumption, and altered the intracellular NAD + /NADH and NADPH/NADP + ratios by up-regulating the proteins involved in energy generation and storage processes, including the Entner–Doudoroff (ED) pathway, the reductive TCA route, trehalose degradation, fatty acid β -oxidation, and PHA biosynthesis. Conclusions This work demonstrated an effective co-carbon feeding strategy to improve PHA content/yield and convert lignin derivatives into value-added products in P. putida KT2440. Co-feeding lignin break-down products with other carbon sources, such as glycerol, has been demonstrated as an efficient way to utilize biomass to increase PHA production in P. putida KT2440. Moreover, the involvement of aromatic degradation favours further lignin utilization, and the combination of proteomics and metabolomics with NMR sheds light on the metabolic and regulatory mechanisms for cellular redox balance and potential genetic targets for a higher biomass carbon conversion efficiency.

09 BIOMASS FUELS↗
DOE OSTI2024

Polyphosphate kinase deletion increases laboratory productivity in cyanobacteria

Identification and manipulation of cellular energy regulation mechanisms may be a strategy to increase productivity in photosynthetic organisms. This work tests the hypothesis that polyphosphate synthesis and degradation play a role in energy management by storing or dissipating energy in the form of ATP. A polyphosphate kinase ( ppk ) knock-out strain unable to synthesize polyphosphate was generated in the cyanobacterium Synechocystis sp. PCC 6803. This mutant strain demonstrated higher ATP levels and faster growth than the wildtype strain in high-carbon conditions and had a growth defect under multiple stress conditions. In a strain that combined ppk deletion with heterologous expression of ethylene-forming enzyme, higher ethylene productivity was observed than in the wildtype background. These results support the role of polyphosphate synthesis and degradation as an energy regulation mechanism and suggest that such mechanisms may be effective targets in biocontainment design.

59 BASIC BIOLOGICAL SCIENCES↗
NASA NTRS2007

Comparative MicroRNA Expression Patterns in Fibroblasts after Low and High Doses of Low-LET Radiation Exposure

Exposure to ionizing radiation causes DNA damage to cells, and provokes a plethora of cellular responses controlled by unique gene-directed signaling pathways. MicroRNAs (miRNAs) are small (~22-nucleotide), non-coding RNAs which functionally silence gene expression by either degrading the messages or inhibiting translation. Here we investigate radiation-dependent changes in these negative regulators by comparing the expression patterns of all 462 known human miRNAs in fibroblasts, after exposure to low (0.1 Gy) or high (2 Gy) doses of X-rays at 30 min, 2, 6 and 24 hrs post-treatment. The expression patterns of microRNAs after low and high doses of radiation show a similar qualitative down-regulation trend at early (0.5 hr) and late (24 hr) time points, with a quantitatively steeper slope following the 2 Gy exposures. Interestingly, an interruption of this downward trend is observed after the 2 Gy exposure, i.e. a significant up-regulation of microRNAs at 2 hrs, then reverting to the downward trend by 6 hrs; this interruption at the intermediate time point was not observed with the 0.1 Gy exposure. At the early time point (0.5 hr), candidate gene targets of selected down-regulated microRNAs, common to both 0.1 and 2 Gy exposures, were those functioning in chromatin remodeling. Candidate target genes of unique up-regulated microRNAs seen at a 2 hr intermediate time point, after the 2 Gy exposure only, are those involved in cell death signaling. Finally, putative target genes of down-regulated microRNAs seen at the late (24 hr) time point after either doses of radiation are those involved in the up-regulation of DNA repair, cell signaling and homeostasis. Thus we hypothesize that after radiation exposure, microRNAs acting as hub negative regulators for unique signaling pathways needed to be down-regulated so as to de-repress their target genes for the proper cellular responses, including DNA repair and cell maintenance. The unique microRNAs up-regulated at 2 hr after 2 Gy suggest the cellular response to functionally suppress the apoptotic death signaling reflex after exposure to high dose radiation. Further analyses with transcriptome and global proteomic profiling will validate the reciprocal expression of signature microRNAs selected in our radiation-exposed cells, and their candidate target gene families, and test our hypothesis that unique radiation-specific microRNAs are keys in governing signaling responses for damage control of this environmental hazard.

Maes, Olivier C.↗
DOE OSTI2023

Manipulation of glycogen and sucrose synthesis increases photosynthetic productivity in cyanobacteria

Photosynthetic productivity is limited by low energy conversion efficiency in naturally evolved photosynthetic organisms, via multiple mechanisms that are not fully understood. Here we show evidence that extends recent findings that cyanobacteria use “futile” cycles in the synthesis and degradation of carbon compounds to dissipate ATP. Reduction of the glycogen cycle or the sucrose cycle in the model cyanobacterium Synechocystis 6803 led to redirection of cellular energy toward faster growth under simulated outdoor light conditions in photobioreactors that was accompanied by higher energy charge [concentration ratio of ATP/(ATP + ADP)]. Such manipulation of energy metabolism may have potential in engineering microalgal chassis cells to increase productivity of biomass or target metabolites.

09 BIOMASS FUELS↗
DOE OSTI2020

Paraburkholderia madseniana sp. nov., a phenolic acid-degrading bacterium isolated from acidic forest soil

RP11 T was isolated from forest soil following enrichment with 4-hydroxybenzoic acid. Cells of RP11 T are aerobic, non-sporulating, exhibit swimming motility, and are rods (0.8 µm by 1.4 µm) that often occur as diplobacillus or in short chains (3–4 cells). Optimal growth on minimal media containing 4-hydroxybenzoic acid (µ=0.216 hr -1 ) occurred at 30 °C, pH 6.5 or 7.0 and 0% salinity. Comparative chemotaxonomic, genomic and phylogenetic analyses revealed the isolate was distinct from its closest relative type strains identified as Paraburkholderia aspalathi LMG 27731 T , Paraburkholderia fungorum LMG 16225 T and Paraburkholderia caffeinilytica CF1 T . Strain RP11 T is genetically distinct from P. aspalathi , its closest relative, in terms of 16S rRNA gene sequence similarity (98.7%), genomic average nucleotide identity (94%) and in silico DNA–DNA hybridization (56.7%±2.8). The composition of fatty acids and substrate utilization pattern differentiated strain RP11 T from its closest relatives, including growth on phthalic acid. Strain RP11 T encoded the greatest number of aromatic degradation genes of all eleven closely related type strains and uniquely encoded a phthalic acid dioxygenase and paralog of the 3-hydroxybenzoate 4-monooxygenase. The only ubiquinone detected in strain RP11 T was Q-8, and the major cellular fatty acids were C 16:0 , 3OH-C 16:0 , C 17:0 cyclo, C 19:0 cyclo ω8c, and summed feature 8 (C 18:1 ω7c/ω6c). On the basis of this polyphasic approach, it was determined that strain RP11 T represents a novel species from the genus Paraburkholderia for which the name Paraburkholderia madseniana sp. nov. is proposed. The type strain is RP11 T (=DSM 110123 T =LMG 31517 T ).

Microbiology↗
DOE OSTI2022

Radiation-Hardened GaN-Based Wireless Communications Architectures for Terrestrial Nuclear Reactor Sensing and Instrumentation

Wireless technologies have become increasingly common in applications, ranging from close proximity inductive communication links in medical devices to short-range Bluetooth and WiFi communications and longer-range cellular communications. Nonetheless, these technologies are unsuitable for use in or around nuclear reactors due to the extreme radiation and temperatures inherent in these environments and their associated significant degradative effects on electronics hardware. However, recent work in wide bandgap– based electronics has shown promise for gallium nitride (GaN) as an emerging technology for the realization of practical wireless communications systems for use in harsh environments. This report presents initial progress on the development of wireless communications architectures designed specifically for nuclear reactor application, based on inherently radiation-hardened (rad-hard) GaN technology. Prior work on rad-hard analog communications topologies is reviewed, and several digital modulation and encoding schemes that utilize GaN-based electronics devices are newly proposed for application in reactor environments. Continuous-time and discrete-time system simulations were performed, and results are presented for the preliminary transmitter and receiver designs, respectively. In addition, an overview of a software-defined radio testbed, designed for communications protocol development, is provided. Future work will focus on implementing candidate radiation-resistant wireless architectures using a research AlGaN/GaN high electron mobility transistor (HEMT) integrated circuit process, which is available at the Ohio State University, and irradiation studies will be carried out to assess the true potential of this technology for reactor application.

22 GENERAL STUDIES OF NUCLEAR REACTORS↗
DOE OSTI2020

A global lipid map defines a network essential for Zika virus replication

Zika virus (ZIKV), an emerging mosquito-borne pathogen of global concern, remodels intra-cellular lipid membranes to form replication factories. How ZIKV regulates lipid networks to allow infection, and consequences for disease is poorly understood. Here, we performed comprehensive lipidomics to create a lipid network map during ZIKV infection. We found that ZIKV significantly alters host lipid composition with striking changes seen within sub-classes of sphingolipids. We also demonstrated that ectopic expression of NS4B, one of the non-structural proteins of ZIKV with known roles in ZIKV replication resulted in similar changes on cellular lipidome. Ceramide, a key bioactive molecule with functions in signaling and apoptosis, redistributes to ZIKV replication factories. Increase of ceramide through knockout of sphingomyelin synthase or via sphingomyelinase-catalyzed degradation of sphingomyelin to ceramide sensitizes cells to ZIKV infection. Remarkably, we observed an enrichment of pro-apoptotic ceramide sub-species in infected cells, providing a link between ZIKV biogenesis and apoptosis. Thus, we identify a sphingolipid metabolic network with a critical role in ZIKV replication and show that ceramide flux is a key mediator of ZIKV infection.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2024

Analysis of 26S Proteasome Activity across Arabidopsis Tissues

Plants utilize the ubiquitin proteasome system (UPS) to orchestrate numerous essential cellular processes, including the rapid responses required to cope with abiotic and biotic stresses. The 26S proteasome serves as the central catalytic component of the UPS that allows for the proteolytic degradation of ubiquitin-conjugated proteins in a highly specific manner. Despite the increasing number of studies employing cell-free degradation assays to dissect the pathways and target substrates of the UPS, the precise extraction methods of highly potent tissues remain unexplored. Here, we utilize a fluorogenic reporting assay using two extraction methods to survey proteasomal activity in different Arabidopsis thaliana tissues. This study provides new insights into the enrichment of activity and varied presence of proteasomes in specific plant tissues.

26S proteasome↗
DOE OSTI2021

Fusion and fission events regulate endosome maturation and viral escape

Endosomes are intracellular vesicles that mediate the communication of the cell with its extracellular environment. They are an essential part of the cell’s machinery regulating intracellular trafficking via the endocytic pathway. Many viruses, which in order to replicate require a host cell, attach themselves to the cellular membrane; an event which usually initiates uptake of a viral particle through the endocytic pathway. In this way viruses hijack endosomes for their journey towards intracellular sites of replication and avoid degradation without host detection by escaping the endosomal compartment. Recent experimental techniques have defined the role of endosomal maturation in the ability of enveloped viruses to release their genetic material into the cytoplasm. Endosome maturation depends on a family of small hydrolase enzymes (or GTPases) called Rab proteins, arranged on the cytoplasmic surface of its membrane. Here, we model endosomes as intracellular compartments described by two variables (its levels of active Rab5 and Rab7 proteins) and which can undergo coagulation (or fusion) and fragmentation (or fission). The key element in our approach is the “per-cell endosomal distribution” and its dynamical (Boltzmann) equation. The Boltzmann equation allows us to derive the dynamics of the total number of endosomes in a cell, as well as the mean and the standard deviation of its active Rab5 and Rab7 levels. We compare our mathematical results with experiments of Dengue viral escape from endosomes. The relationship between endosomal active Rab levels and pH suggests a mechanism that can account for the observed variability in viral escape times, which in turn regulate the viability of a viral intracellular infection.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2021

Metabolic Interactions between Brachypodium and Pseudomonas fluorescens under Controlled Iron-Limited Conditions

Plant iron (Fe) nutritional status has a significant impact on rhizosphere microbial communities. While this effect has been attributed to alterations in the composition of root exudates, the underlying mechanisms are not known. Here, we investigated the effect of Fe deficiency on the interactions between the grass Brachypodium distachyon and the common soil bacteria Pseudomonas fluorescens SBW25. Increased phytosiderophore production was the most significant response of Brachypodium root exudation to Fe deficiency. We observed upregulated expression of phytosiderophore biosynthesis genes but lower accumulated exudate concentrations in the presence of Pseudomonas, indicating that the bacteria degrade these phytosiderophores. Pseudomonas did not produce endogenous siderophores under the low Fe conditions. Rather, the bacterial transcriptome revealed upregulation of four genes in response to Fe deficiency that contain motifs associated with cellular uptake of small molecules. Collectively, these results suggest that rhizosphere Pseudomonas fluorescens may take up phytosiderophores produced by grasses in response to Fe deficiency, and we propose target genes that may be involved. Our findings provide insight into the molecular basis for the exchange of C, N, and Fe between plants and bacteria under Fe deficient conditions. These interactions may contribute to differences in the rhizosphere microbial community structure across soil pH regimes.

Boiteau, Rene M.↗
DOE OSTI2020

The Antioxidant Enzyme Methionine Sulfoxide Reductase A (MsrA) Interacts with Jab1/CSN5 and Regulates Its Function

Methionine sulfoxide (MetO) is an oxidative posttranslational modification that primarily occurs under oxidative stress conditions, leading to alteration of protein structure and function. This modification is regulated by MetO reduction through the evolutionarily conserved methionine sulfoxide reductase (Msr) system. The Msr type A enzyme (MsrA) plays an important role as a cellular antioxidant and promotes cell survival. The ubiquitin- (Ub) like neddylation pathway, which is controlled by the c-Jun activation domain-binding protein-1 (Jab1), also affects cell survival. Jab1 negatively regulates expression of the cell cycle inhibitor cyclin-dependent kinase inhibitor 1B (P27) through binding and targeting P27 for ubiquitination and degradation. Here we report the finding that MsrA interacts with Jab1 and enhances Jab1's deneddylase activity (removal of Nedd8). In turn, an increase is observed in the level of deneddylated Cullin-1 (Cul-1, a component of E3 Ub ligase complexes). Furthermore, the action of MsrA increases the binding affinity of Jab1 to P27, while MsrA ablation causes a dramatic increase in P27 expression. Thus, an interaction between MsrA and Jab1 is proposed to have a positive effect on the function of Jab1 and to serve as a means to regulate cellular resistance to oxidative stress and to enhance cell survival.

59 BASIC BIOLOGICAL SCIENCES↗
NASA NTRS1990

Skeletal muscle fiber, nerve, and blood vessel breakdown in space-flown rats

Histochemical and ultrastructural analyses were performed postflight on hind limb skeletal muscles of rats orbited for 12.5 days aboard the unmanned Cosmos 1887 biosatellite and returned to Earth 2 days before sacrifice. The antigravity adductor longus (AL), soleus, and plantaris muscles atrophied more than the non-weight-bearing extensor digitorum longus, and slow muscle fibers were more atrophic than fast fibers. Muscle fiber segmental necrosis occurred selectively in the AL and soleus muscles; primarily, macrophages and neutrophils infiltrated and phagocytosed cellular debris. Granule-rich mast cells were diminished in flight AL muscles compared with controls, indicating the mast cell secretion contributed to interstitial tissue edema. Increased ubiquitination of disrupted myofibrils implicated ubiquitin in myofilament degradation. Mitochondrial content and succinic dehydrogenase activity were normal, except for subsarcolemmal decreases. Myofibrillar ATPase activity of flight AL muscle fibers shifted toward the fast type. Absence of capillaries and extravasation of red blood cells indicated failed microcirculation. Muscle fiber regeneration from activated satellite cells was detected. About 17% of the flight AL end plates exhibited total or partial denervation. Thus, skeletal muscle weakness associated with spaceflight can result from muscle fiber atrophy and segmental necrosis, partial motor denervation, and disruption of the microcirculation.

NASA Experiment Number COS 1887-6↗
DOE OSTI2022

A distinct RNA recognition mechanism governs Np 4 decapping by RppH

Dinucleoside tetraphosphates, often described as alarmones because their cellular concentration increases in response to stress, have recently been shown to function in bacteria as precursors to nucleoside tetraphosphate (Np 4 ) RNA caps. Removal of this cap is critical for initiating 5' end-dependent degradation of those RNAs, potentially affecting bacterial adaptability to stress; however, the predominant Np 4 decapping enzyme in proteobacteria, ApaH, is inactivated by the very conditions of disulfide stress that enable Np 4 -capped RNAs to accumulate to high levels. Here, in this study, we show that, in Escherichia coli cells experiencing such stress, the RNA pyrophosphohydrolase RppH assumes a leading role in decapping those transcripts, preferring them as substrates over their triphosphorylated and diphosphorylated counterparts. Unexpectedly, this enzyme recognizes Np 4 -capped 5' ends by a mechanism distinct from the one it uses to recognize other 5' termini, resulting in a one-nucleotide shift in substrate specificity. The unique manner in which capped substrates of this kind bind to the active site of RppH positions the δ-phosphate, rather than the β-phosphate, for hydrolytic attack, generating triphosphorylated RNA as the primary product of decapping. Consequently, a second RppH-catalyzed deprotection step is required to produce the monophosphorylated 5' terminus needed to stimulate rapid RNA decay. The unconventional manner in which RppH recognizes Np 4 -capped 5' ends and its differential impact on the rates at which such termini are deprotected as a prelude to RNA degradation could have major consequences for reprogramming gene expression during disulfide stress.

59 BASIC BIOLOGICAL SCIENCES↗