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DOE OSTI2021

The Role of Quantum Science Concepts in Enhancing Sensing and Imaging Technologies: Applications for Biology: Proceedings of a Workshop

Quantum concepts hold the potential to enable significant advances in sensing and imaging technologies that could be vital to the study of biological systems. The workshop Quantum Science Concepts in Enhancing Sensing and Imaging Technologies: Applications for Biology, held online March 8–10, 2021, was organized to examine the research and development needs to advance biological applications of quantum technology. Hosted by the National Academies of Sciences, Engineering, and Medicine, the event brought together experts working on state-of-the-art, quantum-enabled technologies and scientists who are interested in applying these technologies to biological systems. Through talks, panels, and discussions, the workshop facilitated a better understanding of the current and future biological applications of quantum-enabled technologies in fields such as microbiology, molecular biology, cell biology, plant science, mycology, and many others. The workshop was organized around three main themes. The first, quantum in biology, examined quantum concepts that are hypothesized to be important for life processes and that researchers are working to observe through biological imaging and sensing. The second, quantum for biology, addressed ways to use quantum concepts to enhance technologies for biological imaging and sensing. The third, biology for quantum, offered a wider discussion of how the frontiers of biological imaging and sensing could enable future study using quantum concepts, tools, or technologies. Throughout the workshop, participants identified a wide range of emerging approaches and opportunities at the intersection of quantum physics and biological sensing and imaging. During the workshop, there were some differences in how each speaker defined the term quantum. During one of the panels, Prem Kumar offered thoughts on what phenomena are classical versus quantum, explaining that techniques get progressively more quantum as you move from just having superposition to having superposition with measurement and entanglement. Another explanation from Clarice Aiello delineates the definition into several levels. This includes a base level of “quantum-ness,” which reflects that all matter is made of atoms, and when these particles are isolated they behave based on quantum mechanical principles. A second level is related to quantum coherence, where a single quantum object might be found in a coherent superposition state. A final level, which she described as the quantum-entangled level, involves multiple quantum systems which are entangled among themselves. Overall, the workshop touched on concepts such as superposition, entanglement, and squeezing and their potential implications for communication, computing, and simulation, in addition to the workshop’s main focal area, biological sensing and imaging. At the opening of the workshop, Thorsten Ritz of the University of California, Irvine, identified two questions at the heart of quantum biology: Is the machinery of life quantum mechanical, and can quantum mechanics be used to study the machinery of life in new ways? Participants highlighted systems in which researchers have explored these questions, from the vast array of molecular interactions involved in biological processes such as photosynthesis, to the mechanics involved in cellular functions such as differentiation and aggregation, to the role of oscillating magnetic fields in flight orientation among birds. Sensing and imaging technologies are crucial to biological research; these technologies could both enhance the study of quantum effects and be enhanced by quantum concepts. A critical challenge in biological research is to develop imaging and sensing tools that do not damage or interfere with the often fragile and fleeting systems being studied. Attendees discussed a variety of established and emerging technologies that could enhance noninvasive biological imaging, including single- and two-photon spectroscopy, single-molecule spectroscopy, quantum illumination, ghost imaging, and cryo-electron microscopy. One example came from Marlan Scully who gave a keynote address on the first day of the workshop. Scully emphasized the use of different laser technologies, which exhibit coherence and other quantum properties, in moving toward real-world biological applications, such as the detection of SARS-CoV-2. Both tools and theory will play an important role in advancing quantum biology research and applications. Several participants suggested theorists and experimentalists should work in tandem to understand and model biological processes. While physics often reduces systems to their simplest forms for fundamental insights, participants also noted the value of observing and understanding biological systems in all their “messiness,” capturing both the inner workings of biological systems and the complex interactions that occur within and between organisms. In discussions among participants, several attendees stressed the need to match emerging tools with the right scientific questions. Rather than developing quantum technologies as “a hammer looking for a nail,” participants emphasized a focus on exploring the problems these technologies are best suited to address. For example, it is important to consider the size of the phenomenon being studied, the timescales that are important in answering the scientific question, and other relevant considerations. Every tool along the spectrum from classical to quantum involves its own set of trade-offs. For example, Ted Laurence of the Lawrence Livermore National Laboratory said that quantum measurements, such as single photon counting, fluorescence transitions, and lasers, can take longer to produce the same results as classical measurements. These quantum measurements, however, do not require calibration and can enable new research questions to be answered. Prem Kumar, Northwestern University, noted that, despite their promise, quantum approaches should not be used simply for the sake of using quantum, especially in situations where classical approaches better meet the needs of the researcher. Understanding and applying quantum concepts could enable advances in a wide range of application areas including energy, synthetic biology, medicine, and sustainability. For example, Michelle O’Malley, University of California, Santa Barbara, described how improved noninvasive imaging approaches could help capture the complex interactions and functions involved in the breakdown of organic matter by microbial communities and lead to new technologies for capturing valuable products from plant waste. Several other participants discussed needs in tracking the movement of metabolites and molecules in microbial communities for insights into nutrient cycling in environments such as soil. Margaret Ahmad, Sorbonne University, discussed potential opportunities to leverage the magnetic properties of cryptochromes to advance new treatment approaches for diseases such as COVID-19 and cancer. Looking toward the future development of the field, participants discussed challenges to advancing quantum biology that arise from disciplinary disconnects between physicists and biologists. The siloing of academic research disciplines represents a significant barrier to progress. Disconnects in terminology, motivations and priorities, and structural barriers to collaborative work underscore the need for concerted efforts to bridge these divides. Attendees and speakers offered suggestions for resolving these divergences, establishing a shared language, moving the field forward, and fostering meaningful feedback between disciplines. Overall, participants stressed a need for balance, open communication, collaboration, unity, and clear dialogue on trade-offs between quantum and classical approaches. Keiko Torii, The University of Texas at Austin, said that the best collaborations happen when the project provides mutual advantages that can show off everyone’s talents, each team finds the work interesting, and partners develop a camaraderie to pursue new knowledge. To enable near- and long-term opportunities in this space, participants suggested that exploratory, high-risk funding could improve existing instrumentation to explore quantum enhancement collaboratively. They also emphasized the need for collaboration between quantum physicists and sensing/imaging scientists, which could be advanced through a dedicated quantum biology investigator program. People, even more than technology, will be crucial to the future of quantum biology. Participants explored training, education, and workforce needs to further develop this burgeoning field and cultivate the next generation of scientists. While many programs are still in their nascent stages, participants highlighted examples of approaches and programs being developed at various types of institutions to engage students and professional scientists in quantum biology research. Several participants stressed the need for an inclusive approach, spanning disciplines as well as communities to foster a diverse field fueled by the intellectual contributions of a wide range of people, including historically under-resourced schools and students. To increase awareness and excitement about quantum physics and related areas of biology, attendees suggested capitalizing on the “buzz” around quantum. Several participants emphasized the need to start early, introducing students to quantum concepts and their appealing “weirdness” in K–12 education. Engaging students early—before they become entrenched in traditional disciplinary siloes as typically happens in graduate school—could help to galvanize interest in the area and foster a generation of scientists with the interdisciplinary mindset and skills needed to advance this interdisciplinary field. While these efforts could be advanced at many levels and across multiple sectors, several participants suggested a national quantum biology center could be a valuable hub to coordinate and support quantum biology education and workforce development across academia, industry, and government.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

Lake Arrowhead Microbial Genomics Conference

The Lake Arrowhead Microbial Genomics Conference (formerly the E. coli and Small Genomes Conference, and the Microbial Genomes Conference) was held September 11-15, 2022, at the UCLA Conference Center at Lake Arrowhead, California. This conference is part of a yearly meeting initiated in 1991 to bring together genome sequencers, bioinformatics specialists, biologists, and geneticists, to forge interactions that would result in meaningful functional genomics. The goal of the meeting was to translate the influx of new genome sequencing information into useful biological studies. The Lake Arrowhead 2022 meeting had a major focus on microbial communities, the human microbiome, pathogens, phage therapy, bioinformatic methods of analysis, bioenergenics, and environmental microbiology. The field of genomics has reached the point where deriving the sequence of an organism’s entire genome is now seen as a beginning rather than an endpoint. Moreover, understanding the sequences of whole communities, and particularly those that make up different microbiomes, is now a central point of the field. The 2022 meeting covered microorganisms for which extensive analyses exist, and those for which new biological and technical strategies are being developed. The focus on biodiversity, the human microbiome, pathogenic organisms and methods of countering them, and bioenergetics added special significance to this meeting. This meeting was designed to have a mix of 36 invited presentations and poster sessions with more than 80 posters, and had 147 participants. This meeting has become established as a major annual microbial genomics conference during the last 29 years and has assured status and quality. It offers a major opportunity for young investigators and students to attend, present posters, and give talks. In fact, 50% of the invited speakers were early career stage scientists. An important impact of the conference was the forging of new collaborations between scientists and particularly young scientists with the array of multidisciplinary researchers at the meeting.

Miller, Jeffrey H.↗
DOE OSTI2023

Coupling Waste Feedstocks to Microbial Protein for a Circular Food System

The global food system is responsible for approximately 34% of annual greenhouse gas (GHG) emissions and up to 85% of water consumption. This critical sector suffers from intensive and inefficient land and water use, the generation of multiple (solid, liquid, and gaseous) waste streams, and high fuel, fertilizer, and pesticide consumption. The production of waste-derived microbial protein (MP) represents a promising alternative for reducing the environmental impacts of protein production relative to conventional agriculture. MP can be mass-produced in volumetrically scalable cultivation processes on short timescales, enabling facile up-scaling with lower greenhouse gas emissions, land use, and water impacts than animal and, in some cases, plant protein production. MP can also be produced from waste feedstocks, diverting waste from landfills or the natural environment. Here, we present the availability and suitability of waste feedstocks for MP production, as well as the fermentation and downstream processes required to convert MP into human food products. We discuss the challenges and opportunities facing waste-derived bacterial MP and highlight key areas for innovation in both the microbiology and process design space for a more sustainable and circular food system.

BASIC BIOLOGICAL SCIENCES,ENERGY PLANNING, POLICY,↗
DOE OSTI2020

Pump-and-Treat Impacts on Aquifer Conditions and the Development of System-Wide Performance Strategies - 20377

The pump-and-treat (P and T) system on the Hanford Site's Central Plateau currently treats multiple contaminants within the 200-ZP-1 operable unit, including nitrate through its biological treatment processes. Although it was initially designed to support the 200-ZP-1 Remedial Action Objectives, the P and T system has evolved in use to address contaminants from other regions of the aquifer as well. The P and T influents are biogeochemically distinct, treating groundwaters with elevated concentrations of cyanide, uranium, technetium-99, and nitrate. Given the varying geochemical signatures from current and potential future sources, operational conditions, and amendments considered for treatment processes, there is a need to identify the impacts that the P and T effluent has on the aquifer, and how the aquifer affects P and T performance. To this end, laboratory efforts have focused on characterizing P and T effluent and its biogeochemical effects on the sediments through a series repacked column experiments. The P and T effluent is rich in particulate matter. A systematic inspection of this particulate matter in the P and T effluent showed significant levels of manganese/iron oxides, calcite crystals, relatively large organic matter molecules, and biomass. Therefore, the column experiments were designed and conducted for four different treatment systems: artificial groundwater, filtered P and T effluent, non-filtered P and T effluent, and chlorinated P and T effluent. Different levels of microbiologically- and geochemically-induced changes in the columns occurred and modified the flow systems. For example, physical filtration occurred in the sediments where the particulate material had accumulated in the top portion of the column. Reactive transport modeling of the laboratory experiments is being carried out so that the reaction network can be extrapolated to the field scale. These simulations will provide predictive evaluations of system performance based on P and T operational changes and aquifer mass loading and capacity estimates. This information can be used to address combined subsurface and operational challenges, such as the need for frequent well redevelopment to maintain injection capacity. This information also provides the technical basis for decisions related to the P and T system operations that support remedy optimization efforts and short- and long-term remedy decisions related to multiple operable units on the Hanford Site Central Plateau. (authors)

12 MANAGEMENT OF RADIOACTIVE AND NON-RADIOACTIVE W↗
DOE OSTI2020

Assessment of the Corrosion Risks to a Double Shell Tank During the Selective Removal of a Cesium-rich Supernate Layer - 20227

The Direct Feed Low Activity Waste (DF-LAW) activity will provide the initial feed to the Waste Treatment and Immobilization Plant at the Hanford Tank Farm facility. Feed preparation for DF-LAW will be carried out in multiple double shell waste tanks (DST) utilizing the Tank Side Cesium Removal (TSCR) process. Tank AP-106 will be an integral part of the TSCR process and hence key to the DFLAW activity. Prior to being utilized for this purpose, the Cs-137 concentration levels in the AP-106 supernate must be reduced significantly. The AP-106 re-purposing process manages normal waste tank addition procedures such that waste acceptance criteria objectives for Cs-137 removal are achieved. As this process was being developed, several concerns regarding tank integrity during and after repurposing of Tank AP-106, and the associated tanks (i.e., AP-102, AN-101, and AP-108), were addressed. The potential corrosion degradation mechanisms identified based on process knowledge included: general and pitting corrosion, caustic stress corrosion cracking, concentration cell corrosion, and microbiologically induced corrosion (MIC). A systematic approach was undertaken to assess the risk of corrosion during the AP-106 re-purposing process. The assessment considered the initial and present condition of the tank, the service history of the tank, the environmental conditions created in each tank during each stage of the process, process timing, and corrosion testing and modeling that approximated tank conditions. Based on this information, recommendations were provided to minimize the exposure of the tank to corrosive conditions. These recommendations included time limits for each stage of the process, staging of corrosion inhibitor solutions should process delays occur, and waste sampling with potential chemistry adjustments at defined steps in the process. Additionally, recommendations for post-process supernate chemistry adjustments and tank inspections were put forth. The greatest risk for this process occurs during the manipulation of large quantities of Columbia River water (i.e., greater than 450,000 gallons) within AP-106, the associated tanks and the supporting infrastructure (i.e., transfer line piping). This risk becomes more pronounced if there is a process upset and the tank walls remain exposed to a corrosive condition longer than intended. As long as the river water is blended into the supernate in a timely fashion, and the waste is adjusted appropriately with corrosion inhibitors, corrosion may be minimized and the final condition of the supernate in AP-106 and the associated tanks is expected to be in compliance with corrosion chemistry requirements. Contingency plans will be made to minimize the impact of a process upset. (authors)

12 MANAGEMENT OF RADIOACTIVE AND NON-RADIOACTIVE W↗
DOE OSTI2020

Study of an Unrefined Humate Solution as a Possible Attenuation-based Remedy for Uranium Contamination in Acidic Groundwater - 20351

The Savannah River Site (SRS), joined the environmental cleanup program in 1981 after the Cold War, and was designated as a hazardous waste management facility. About 1.8 billion gallons of acid wastes were disposed into the F/H Area Seepage Basin that included many radionuclides and dissolved metals, resulting in highly contaminated groundwater plumes with pH of 3 - 5.5. The acidity of the plumes contributes to the mobility of several constituents of concern (COC) such as H-3, U-238, I-129, and Sr-90 for the F-Area plume and H-3, Sr-90 and mercury for the H-Area plume. An active treatment (pump-and-treat and re-injection) system was constructed and put in place in 1997 to address the removal of metals and radionuclides from the contaminated groundwater. As remediation projects advanced, active treatments were transitioned to more passive or enhanced-passive approaches such as the subsurface barrier with base injection system, which has been controlling the mobility of contaminants. However, new and more efficient attenuation-based remedies are always sought. Several studies have proposed that humic substances can be used to remediate sites contaminated with heavy metals by creating a permeable reactive barrier. Humic substances (HS) are major components of soil organic matter. HS are polyfunctional organic macromolecules formed by the chemo-microbiological decomposition of biomass or dead organic matter. These substances are usually divided into three main fractions: humin (insoluble at all pH values), humic acid (soluble at pH greater than 3.5), and fulvic acid (soluble at all pH values). Humic substances are major components of the soil at SRS and, in addition, are also helpful in the removal process of uranium. This research focuses on uranium (VI), which is a key contaminant of concern in the F-Area groundwater plume. The interaction of uranium with sediment in the presence or absence of humic substances involves complex mechanisms that are not yet well understood. The interaction of U(VI) with humic substances can affect the adsorption of U on sediment, altering its mobility in the subsurface. The objective of this study is to study the mobility of uranium in the presence of humic acid and to determine if sediments amended by humic acid can enhance the sorption of heavy metals onto sediments. In this research, experiments were conducted using background (clean) F-Area aquifer sediments to understand and predict uranium mobility. A blended material, humate, containing both humic and fulvic acids and a chemically modified humate (KW15 modified humic or mod-HA) was studied as a possible amendment for uranium remediation in SRS groundwater. Batch experiments were conducted in triplicates using 20 ml of aqueous suspension in DI water containing SRS sediment and mod-HA and studied the effect of pH, kinetics of humate sorption onto SRS sediment and also studied the sorption of uranium onto mod-HA amended SRS sediment. Results indicate that mod-HA amended sediment increases uranium removal significantly. (authors)

12 MANAGEMENT OF RADIOACTIVE AND NON-RADIOACTIVE W↗
DOE OSTI2024

Effectiveness of Vapor Corrosion Inhibitors Under Bacterial Activity for Aboveground Storage Tank Application

Bacterial activity is known to influence corrosion and can result in increasing soil-side corrosion of the aboveground storage tanks (ASTs) floor plates. Association for Materials Protection and Performance (AMPP) defines this phenomenon as Microbiologically Influenced Corrosion (MIC). It has been hypothesized that vapor corrosion inhibitors (VCIs) can not only function in presence of bacterial activity but can also function as biocides. Experiments were conducted to study the effect of bacterial activity on corrosion in the sand pad conditions and effect of VCIs in mitigating MIC. Various bacteria cultures were grown using commercially available culture media, and coupon exposure tests were performed with field sand mixed with the culture media. To this end, control and VCI-dosed electrolytes were prepared using the field sand plus bacteria cultures. Pre- and post-test bacteria concentration estimation and measurements, respectively, in the VCI-dosed electrolytes indicated that VCIs do not function as biocides, however, VCIs are highly effective for mitigating corrosion in presence of bacterial activity.

Shukla, Pavan K.↗
DOE OSTI2023

CO2 Valorization via Rewiring Carbon Metabolic Network

Energizing the world sustainably requires revolutionizing the way we harness natural resources. New renewable technologies include the microbiological upgrading of CO2, the primary greenhouse gas that causes climate change. This project aims to develop a new bioeconomy by taking up CO2 and converting it into high-value products using an autotrophic microbe. Specifically, the team constructed an acetogenic bacterium, Clostridium ljungdahlii, and developed gas fermentation processes that convert syngas (CO2/H2/CO) to the platform compound 3-hydroxybutyrate (3HB). The project has achieved several key successes, including the design and construction of 3HB genes in the gas-fermenting bacterium, the identification of a native 3HB pathway, the conversion of syngas to 3HB through strain engineering, and the improvement of the conversion through rational and iterative strain development. The optimized strain has achieved industry-relevant production rates (0.1 g/L/h) and titers (9.2 g/L), and sustained continuous gas fermentation for 3HB production for > 50 days in a bench-top gas bioreactor. The project has both industrial and scientific implications, including the exploitation of host microbes and processes to improve industrial-scale production of value-added compounds from CO2, and the promotion of gas fermentation technology. The team is also strengthening R&D synergies with other teams and industrial partners in the CO2 consortium, developing cutting-edge approaches for rapid and rational engineering of gas-fermenting hosts, and dissecting the thermodynamics and kinetics of processes for biological syngas valorization. So far, one patent application and one research article have resulted from this work, and the team anticipates additional significant outcomes from this project.

3-hydroxybutyrate↗
DOE OSTI2024

MIBiG 4.0: advancing biosynthetic gene cluster curation through global collaboration

Specialized or secondary metabolites are small molecules of biological origin, often showing potent biological activities with applications in agriculture, engineering and medicine. Usually, the biosynthesis of these natural products is governed by sets of co-regulated and physically clustered genes known as biosynthetic gene clusters (BGCs). To share information about BGCs in a standardized and machine-readable way, the Minimum Information about a Biosynthetic Gene cluster (MIBiG) data standard and repository was initiated in 2015. Since its conception, MIBiG has been regularly updated to expand data coverage and remain up to date with innovations in natural product research. Here, we describe MIBiG version 4.0, an extensive update to the data repository and the underlying data standard. In a massive community annotation effort, 267 contributors performed 8304 edits, creating 557 new entries and modifying 590 existing entries, resulting in a new total of 3059 curated entries in MIBiG. Particular attention was paid to ensuring high data quality, with automated data validation using a newly developed custom submission portal prototype, paired with a novel peer-reviewing model. MIBiG 4.0 also takes steps towards a rolling release model and a broader involvement of the scientific community. MIBiG 4.0 is accessible online at https://mibig.secondarymetabolites.org/.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2026

Heterotrophic respiration by soil microbes in a changing climate

Soil microbes strongly influence the soil organic carbon (SOC) pool, which globally stores ~2,000 PgC. Specifically, the balance between microbial heterotrophic respiration (R H ), which degrades SOC, and plant–microbe interactions that stabilize SOC determines whether terrestrial ecosystems are a net source or sink of CO 2 to the atmosphere. Here, in this Review, we evaluate how climate change alters these competing processes. R H is approximately half of total soil respiration, at ~50 PgC yr −1 , with 70% occurring in topsoils. Warming accelerates microbial metabolism, with a 10 °C temperature increase estimated to raise R H by ~50%, an effect that is particularly strong in Arctic soils. Warming also reduces soil moisture, further modulating R H , which responds nonlinearly to soil moisture, being limited by saturation and desiccation and meeting a maximum at intermediate levels. Consequently, R H is highly sensitive to future precipitation changes and drought. However, soil management strategies could enhance SOC stocks and persistence under climate change. Bacterial and fungal inoculants can promote SOC production and stabilization, while deep-rooting plants increase SOC inputs to deeper layers that experience lower R H . Agricultural practices and biochar amendments can also enhance SOC and reduce R H . Expanding field trials across regions, climates and soil types would improve empirical understanding of these responses and support better representation of R H in predictive models, enabling more accurate assessments of climate impacts on SOC storage.

Jansson, Janet K. [Pacific Northwest National Labo↗
DOE OSTI2025

Torsional twist of the SARS ‐ CoV and SARS ‐ CoV ‐2 SUD ‐N and SUD ‐M domains

Abstract Coronavirus non‐structural protein 3 (nsp3) forms hexameric crowns of pores in the double membrane vesicle that houses the replication–transcription complex. Nsp3 in SARS‐like viruses has three unique domains absent in other coronavirus nsp3 proteins. Two of these, SUD‐N (Macrodomain 2) and SUD‐M (Macrodomain 3), form two lobes connected by a peptide linker and an interdomain disulfide bridge. We resolve the first complete x‐ray structure of SARS‐CoV SUD‐N/M as well as a mutant variant of SARS‐CoV‐2 SUD‐N/M modified to restore cysteines for interdomain disulfide bond naturally lost by evolution. Comparative analysis of all structures revealed SUD‐N and SUD‐M are not rigidly associated but rather have significant rotational flexibility. Phylogenetic analysis supports that the potential to form the disulfide bond is common across betacoronavirus isolates from many bat species and civets, but also one or both of the cysteines that form the disulfide bond are absent across isolates from bats and pangolins. The absence of these cysteines does not impact viral replication or protein translation.

Rosas‐Lemus, Monica [Department of Microbiology‐Im↗
DOE OSTI2025

De Novo Design of High‐Affinity Miniprotein Binders Targeting Francisella Tularensis Virulence Factor

Abstract Francisella tularensis poses considerable public health risk due to its high infectivity and potential for bioterrorism. Francisella‐like lipoprotein (Flpp3), a key virulence factor unique to Francisella, plays critical roles in infection and immune evasion, making it a promising target for therapeutic development. However, the lack of well‐defined binding pockets and structural information on native interactions has hindered structure‐guided ligand discovery against Flpp3. Here, we used a combination of physics‐based and deep‐learning methods to design high‐affinity miniprotein binders targeting two distinct sites on Flpp3. We identified four binders for site I with binding affinities ranging between 24–110 nM. For the second site, an initial binder showed a dissociation constant ( K D ) of 81 nM, and subsequent site saturation mutagenesis yielded variants with sub‐nanomolar affinities. Circular dichroism confirmed the topology of designed miniproteins. The X‐ray crystal structure of Flpp3 in complex with a site I binder is nearly identical to the design model (Cα root‐mean‐square deviation (RMSD): 0.9 Å). These designed miniproteins provide research tools to explore the roles of Flpp3 in tularemia and should enable the development of new therapeutic candidates.

Gokce‐Alpkilic, Gizem [Molecular Engineering and S↗
DOE OSTI2025

CpoS-Inc interactions facilitate host cell modulation during Chlamydia trachomatis infection

ABSTRACT Chlamydia trachomatis ( C.t .), the leading bacterial cause of sexually transmitted infections, replicates within a unique intracellular compartment called the inclusion, which is modified by secreted proteins known as inclusion membrane (Inc) proteins. Here, we further characterize CpoS, an Inc protein previously shown to be critical for bacterial replication and inclusion development. We demonstrate that CpoS directly binds multiple coiled-coil region-containing Incs and engages Rab GTPases at a separate site. Notably, CpoS-InaC interactions facilitate the recruitment of select Arf GTPases to the inclusion membrane, while Rab recruitment occurs independently of these interactions. Biochemical and biophysical analyses revealed that Incs self-oligomerize to form higher-ordered structures, with CpoS adopting a tetrameric conformation resembling that of eukaryotic SNARE proteins. We propose that these assemblies serve as scaffolds to orchestrate vesicle docking, tethering, and fusion. Our findings highlight the intricate interplay between bacterial and host factors, revealing how C.t . leverages both Inc-Inc interactions and host protein engagement to manipulate vesicular trafficking and sustain infection.

Tijerina, Xavier [Department of Microbiology and I↗