Selected Publications

EMBO J. Dec 2022

Han Groisman EMBO 2022

Weiwei Han, Bee-Zen Peng, Chunyan Wang, Guy E Townsend II, Natasha A Barry, Frank Peske, Andrew L Goodman, Jun Liu, Marina V Rodnina, Eduardo A Groisman.

Gut colonization by Bacteroides requires translation by an EF-G paralog lacking GTPase activity.

Protein synthesis is crucial for cell growth and survival yet one of the most energy-consuming cellular processes. How, then, do cells sustain protein synthesis under starvation conditions when energy is limited? To accelerate the translocation of mRNA–tRNAs through the ribosome, bacterial elongation factor G (EF-G) hydrolyzes energy-rich guanosine triphosphate (GTP) for every amino acid incorporated into a protein. Here, we identify an EF-G paralog—EF-G2—that supports translocation without hydrolyzing GTP in the gut commensal bacterium Bacteroides thetaiotaomicron. EF-G2’s singular ability to sustain protein synthesis, albeit at slow rates, is crucial for bacterial gut colonization. EF-G2 is ~10-fold more abundant than canonical EF-G1 in bacteria harvested from murine ceca and, unlike EF-G1, specifically accumulates during carbon starvation. Moreover, we uncover a 26-residue region unique to EF-G2 that is essential for protein synthesis, EF-G2 dissociation from the ribosome, and responsible for the absence of GTPase activity. Our findings reveal how cells curb energy consumption while maintaining protein synthesis to advance fitness in nutrient-fluctuating environments.

Nat Struct Mol Biol. Nov 2022

Jason Gorman, Chunyan Wang…Jun Liu, Mario Roederer, Peter D Kwong, et al.

Cryo-EM structures of prefusion SIV envelope trimer.

Simian immunodeficiency viruses (SIVs) are lentiviruses that naturally infect non-human primates of African origin and seeded cross-species transmissions of HIV-1 and HIV-2. Here we report prefusion stabilization and cryo-EM structures of soluble envelope (Env) trimers from rhesus macaque SIV (SIVmac) in complex with neutralizing antibodies. These structures provide residue-level definition for SIV-specific disulfide-bonded variable loops (V1 and V2), which we used to delineate variable-loop coverage of the Env trimer. The defined variable loops enabled us to investigate assembled Env-glycan shields throughout SIV, which we found to comprise both N- and O-linked glycans, the latter emanating from V1 inserts, which bound the O-link-specific lectin jacalin. We also investigated in situ SIVmac-Env trimers on virions, determining cryo-electron tomography structures at subnanometer resolutions for an antibody-bound complex and a ligand-free state. Collectively, these structures define the prefusion-closed structure of the SIV-Env trimer and delineate variable-loop and glycan-shielding mechanisms of immune evasion conserved throughout SIV evolution.

Cell. 2022 Nov

Hatzios Cell 2022

Daniel G. Dumitrescu, Stavroula K. Hatzios, et al.

A microbial transporter of the dietary antioxidant ergothioneine.

Low-molecular-weight (LMW) thiols are small-molecule antioxidants required for the maintenance of intracellular redox homeostasis. However, many host-associated microbes, including the gastric pathogen Helicobacter pylori, unexpectedly lack LMW-thiol biosynthetic pathways. Using reactivity-guided metabolomics, we identified the unusual LMW thiol ergothioneine (EGT) in H. pylori. Dietary EGT accumulates to millimolar levels in human tissues and has been broadly implicated in mitigating disease risk. Although certain microorganisms synthesize EGT, we discovered that H. pylori acquires this LMW thiol from the host environment using a highly selective ATP-binding cassette transporter—EgtUV. EgtUV confers a competitive colonization advantage in vivo and is widely conserved in gastrointestinal microbes. Furthermore, we found that human fecal bacteria metabolize EGT, which may contribute to production of the disease-associated metabolite trimethylamine N-oxide. Collectively, our findings illustrate a previously unappreciated mechanism of microbial redox regulation in the gut and suggest that inter-kingdom competition for dietary EGT may broadly impact human health.

Proc Natl Acad Sci U S A . 2022 Oct

See also: Yale News “Some facts about foreign genes”

PNAS Oct 2022 Choi Groisman

Jeongjoon Choi, Matias Schmukler, and Eduardo A. Groisman.

Degradation of gene silencer is essential for expression of foreign genes and bacterial colonization of the mammalian gut.

To express foreign genes, enteric bacteria must overcome the silencing effects caused by the heat-stable nucleoid structuring (H-NS) protein. Overcoming gene silencing by H-NS has been largely ascribed to proteins that compete with H-NS for binding to foreign DNA. However, we now report that H-NS proteolysis is a general mechanism of overcoming foreign gene silencing and essential for colonization of the murine gut by the enteric bacterium Escherichia coli. Both E. coli and Salmonella enterica serovar Typhimurium express foreign genes by degrading H-NS but do so under disparate conditions, likely reflecting their distinct lifestyles.

Front. Microbiol. 2022 May

Fig 1 Guo Frontiers

Shuaiqi Guo1,2 and Jun Liu1,2*.

The Bacterial Flagellar Motor: Insights Into Torque Generation, Rotational Switching, and Mechanosensing.

The flagellar motor is a bidirectional rotary nanomachine used by many bacteria to sense and move through environments of varying complexity. The bidirectional rotation of the motor is governed by interactions between the inner membrane-associated stator units and the C-ring in the cytoplasm. In this review, we take a structural biology perspective to discuss the distinct conformations of the stator complex and the C-ring that regulate bacterial motility by switching rotational direction between the clockwise (CW) and counterclockwise (CCW) senses. We further contextualize recent in situ structural insights into the modulation of the stator units by accessory proteins, such as FliL, to generate full torque. The dynamic structural remodeling of the C-ring and stator complexes as well as their association with signaling and accessory molecules provide a mechanistic basis for how bacteria adjust motility to sense, move through, and survive in specific niches both outside and within host cells and tissues.

Nat Commun. 2022 Apr

Fig Nat Commun (Liu)

Jun Liu # 1, Xiaoying Wang # 1 2, Ann T Chen # 1 3, Xingchun Gao # 1, et al

#Contributed equally.

ZNF117 regulates glioblastoma stem cell differentiation towards oligodendroglial lineage.

Glioblastoma (GBM) is a deadly disease without effective treatment. Because glioblastoma stem cells (GSCs) contribute to tumor resistance and recurrence, improved treatment of GBM can be achieved by eliminating GSCs through inducing their differentiation. Prior efforts have been focused on studying GSC differentiation towards the astroglial lineage. However, regulation of GSC differentiation towards the neuronal and oligodendroglial lineages is largely unknown. To identify genes that control GSC differentiation to all three lineages, we performed an image-based genome-wide RNAi screen, in combination with single-cell RNA sequencing, and identified ZNF117 as a major regulator of GSC differentiation. Using patient-derived GSC cultures, we show that ZNF117 controls GSC differentiation towards the oligodendroglial lineage via the Notch pathway. We demonstrate that ZNF117 is a promising target for GSC differentiation therapy through targeted delivery of CRISPR/Cas9 gene-editing nanoparticles. Our study suggests a direction to improve GBM treatment through differentiation of GSCs towards various lineages.

Nat Chem Biol. 2022 Mar

Figure Nat Chem Biol Hatzios

An infection-induced oxidation site regulates legumain processing and tumor growth.

Yekaterina Kovalyova, Daniel W. Bak, Elizabeth M. Gordon, Connie Fung, Jennifer H. B. Shuman, Timothy L. Cover, Manuel R. Amieva, Eranthie Weerapana & Stavroula K. Hatzios.

Oxidative stress is a defining feature of most cancers, including those that stem from carcinogenic infections. Reactive oxygen species can drive tumor formation, yet the molecular oxidation events that contribute to tumorigenesis are largely unknown. Here we show that inactivation of a single, redox-sensitive cysteine in the host protease legumain, which is oxidized during infection with the gastric cancer-causing bacterium Helicobacter pylori, accelerates tumor growth. By using chemical proteomics to map cysteine reactivity in human gastric cells, we determined that H. pylori infection induces oxidation of legumain at Cys219. Legumain oxidation dysregulates intracellular legumain processing and decreases the activity of the enzyme in H. pylori-infected cells. We further show that the site-specific loss of Cys219 reactivity increases tumor growth and mortality in a xenograft model. Our findings establish a link between an infection-induced oxidation site and tumorigenesis while underscoring the importance of cysteine reactivity in tumor growth.

Proc Natl Acad Sci. 2022 Mar

Fig 5 PNAS Shuaiqi Guo 2022

FliL ring enhances the function of periplasmic flagella.

Shuaiqi Guo 1 2, Hui Xu 3, Yunjie Chang 1 2, Md A Motaleb 3, Jun Liu 1 2.

Bacterial flagellar motors are rotary machines that can power motility in various fluid and surface environments, including within hosts. Activation of the stator complex MotA/MotB is required for torque generation and motor rotation. During activation, the stator complex is expected to undergo an extensive conformational change to allow ions to flow through its transmembrane channels to generate torque. However, the detailed mechanism underlying stator activation remains poorly understood. Here, we use the Lyme disease–causing spirochete Borrelia burgdorferi as the model system to reveal the stator complex and its interaction with the FliL ring, using cryo-electron tomography and subtomogram averaging of flagellar motors from wild-type, ΔmotB, ΔfliL, and ΔfliLmotAB mutants. Upon recruitment of stator units to the motor, FliL oligomerizes from a partial ring into a full ring, which wraps around the MotB periplasmic linkers and stabilizes the stator complex in an extended, active conformation, thus enabling a continuous influx of ions to generate higher torque. Furthermore, we provide evidence that FliL can mediate the assembly of stator complexes around the motor, thereby regulating stator and motor function. Given that FliL and the stator complex are ubiquitous in flagellated bacteria, these mechanisms may be utilized by various bacteria to modulate torque and motility in response to changing environmental conditions.

Nat Commun. 2022 Feb

Fig 1 Nat Commun (Malvankar)

Protein nanowires with tunable functionality and programmable self-assembly using sequence-controlled synthesis.

Daniel Mark Shapiro, Gunasheil Mandava, Sibel Ebru Yalcin, Pol Arranz-Gibert, Peter J. Dahl, Catharine Shipps, Yangqi Gu, Vishok Srikanth, Aldo I. Salazar-Morales, J. Patrick O’Brien, Koen Vanderschuren, Dennis Vu, Victor S. Batista, Nikhil S. Malvankar & Farren J. Isaacs.

Advances in synthetic biology permit the genetic encoding of synthetic chemistries at monomeric precision, enabling the synthesis of programmable proteins with tunable properties. Bacterial pili serve as an attractive biomaterial for the development of engineered protein materials due to their ability to self-assemble into mechanically robust filaments. However, most biomaterials lack electronic functionality and atomic structures of putative conductive proteins are not known. Here, we engineer high electronic conductivity in pili produced by a genomically-recoded E. coli strain. Incorporation of tryptophan into pili increased conductivity of individual filaments >80-fold. Computationally-guided ordering of the pili into nanostructures increased conductivity 5-fold compared to unordered pili networks. Site-specific conjugation of pili with gold nanoparticles, facilitated by incorporating the nonstandard amino acid propargyloxy-phenylalanine, increased filament conductivity ~170-fold. This work demonstrates the sequence-defined production of highly-conductive protein nanowires and hybrid organic-inorganic biomaterials with genetically-programmable electronic functionalities not accessible in nature or through chemical-based synthesis.

Proc Natl Acad Sci. 2022 Feb

Fig 1 PNAS Sanchez

Top-down and bottom-up cohesiveness in microbial community coalescence.

Juan Diaz-Colunga 1 2, Nanxi Lu 1 2, Alicia Sanchez-Gorostiaga 1 2 3, Chang-Yu Chang 1 2, Helen S Cai 1 2, Joshua E Goldford 4, Mikhail Tikhonov 5, Álvaro Sánchez 6 2.

Microbial communities frequently invade one another as a whole, a phenomenon known as community coalescence. Despite its potential importance for the assembly, dynamics, and stability of microbial consortia, as well as its prospective utility for microbiome engineering, our understanding of the processes that govern it is still very limited. Theory has suggested that microbial communities may exhibit cohesiveness in the face of invasions emerging from collective metabolic interactions across microbes and their environment. This cohesiveness may lead to correlated invasional outcomes, where the fate of a given taxon is determined by that of other members of its community—a hypothesis known as ecological coselection. Here, we have performed over 100 invasion and coalescence experiments with microbial communities of various origins assembled in two different synthetic environments. We show that the dominant members of the primary communities can recruit their rarer partners during coalescence (top-down coselection) and also be recruited by them (bottom-up coselection). With the aid of a consumer-resource model, we found that the emergence of top-down or bottom-up cohesiveness is modulated by the structure of the underlying cross-feeding networks that sustain the coalesced communities. The model also predicts that these two forms of ecological coselection cannot co-occur under our conditions, and we have experimentally confirmed that one can be strong only when the other is weak. Our results provide direct evidence that collective invasions can be expected to produce ecological coselection as a result of cross-feeding interactions at the community level.

Proc Natl Acad Sci. 2022 Jan

Fig 5 PNAS Tachiyama 2022

The flagellar motor protein FliL forms a scaffold of circumferentially positioned rings required for stator activation.

Shoichi Tachiyama 1, Kar L Chan 2, Xiaolin Liu 3, Skander Hathroubi 3, Briana Peterson 2, Mohammad F Khan 2, Karen M Ottemann 4, Jun Liu 5, Anna Roujeinikova 6 7.

The flagellar motor stator is an ion channel nanomachine that assembles as a ring of the MotA5MotB2 units at the flagellar base. The role of accessory proteins required for stator assembly and activation remains largely enigmatic. Here, we show that one such assembly factor, the conserved protein FliL, forms an integral part of the Helicobacter pylori flagellar motor in a position that colocalizes with the stator. Cryogenic electron tomography reconstructions of the intact motor in whole wild-type cells and cells lacking FliL revealed that the periplasmic domain of FliL (FliL-C) forms 18 circumferentially positioned rings integrated with the 18 MotAB units. FliL-C formed partial rings in the crystal, and the crystal structure–based full ring model was consistent with the shape of the rings observed in situ. Our data suggest that each FliL ring is coaxially sandwiched between the MotA ring and the dimeric periplasmic MotB moiety of the stator unit and that the central hole of the FliL ring has density that is consistent with the plug/linker region of MotB in its extended, active conformation. Significant structural similarities were found between FliL-C and stomatin/prohibitin/flotillin/HflK/C domains of scaffolding proteins, suggesting that FliL acts as a scaffold. The binding energy released upon association of FliL with the stator units could be used to power the release of the plug helices. The finding that isolated FliL-C forms stable partial rings provides an insight into the putative mechanism by which the FliL rings assemble around the stator units.

  • Annu Rev Microbiol. 2021 Oct. Cellular Adaptations to Cytoplasmic Mg 2+ Limitation.

  • Cell Syst. 2021 Oct. Functional attractors in microbial community assembly.

  • Nature. 2021 Sept. Structure of Geobacter pili reveals secretory rather than nanoqire behaviour.

  • Protein Sci. 2021 Aug. The regulation of DNA supercoiling across evolution.

  • Nat Ecol Evol. 2021 Jul. Engineering complex communities by directed evolution.

  • J Bacteriol. 2021 May. Low cytoplasmic magnesium increases the specificity of the Lon and ClpAP proteases.

  • Trends Microbiol. 2021 Apr. Structural basis of bacterial flagellar motor rotation and switching.

  • Elife. 2021 Apr. Nutrient dominance governs the assembly of microbial communities in mixed nutrient environments.

  • Nat Commun. 2021 Mar. Complex yeast-bacteria interactions affect the yield of industrial ethanol fermentation.

  • Sci Signal. 2021 Jan. Reduced ATP-dependent proteolysis of functional proteins during nutrient limitation speeds the return of microbes to a growth state.

  • Proc Natl Acad Sci USA. 2021 Jan. Intrinsic electronic conductivity of individual atomically resolved amyloid crystals reveals micrometer-long hole hopping via tyrosines.

  • PLoS Pathog. 2020 Dec. Chemical tools for decoding redox signaling at the host-microbe interface.

  • Biomolecules. 2020 Oct. Structural conservation and adaptation of the bacterial flagellar motor.

  • PLoS Genet. 2020 Oct. DNA supercoiling differences in bacteria result from disparate DNA gyrase activation by polyamines.

  • Nucleic Acids Res. 2020 Oct. Horizontally acquired regulatory gene activates ancestral regulatory system to promote Salmonella virulence.

  • Nat Struct Mol Biol. 2020 Sept. Molecular mechanism for rotational switching of the bacterial flagellar motor.

  • Proc Natl Acad Sci. 2020 Aug. Small proteins regulate Salmonella survival inside macrophages by controlling degradation of a magnesium transporter.

  • Nat Chem Biol. 2020 Aug. See also News and Views: Uncovering nature’s electronics and Live Science: Scientists find ‘secret molecule’ that allows bacteria to exhale electricityElectric field stimulates production of highly conductive microbial OmcZ nanowires.

  • Nat Struct Mol Biol. 2020 Jun. Subnanometer structures of HIV-1 envelope trimers on aldrithiol-2-inactivated virus particles.

  • Proc Natl Acad Sci. 2020 Apr. Salmonella expresses foreign genes during infection by degrading their silencer.

  • Elife. 2020 Mar. An asymmetric sheath controls flagellar supercoiling and motility in the leptospira spirochete.

  • PLoS Biol. 2019 Dec. High-order interactions distort the functional landscape of microbial consortia.

  • Proteopedia 2019 -  2014 Jun. Malvankar Lab OmcS nanowire structure named one of the highest impact structures of the century.

  •  2019 Jun. Functional characterization of a subtilisin-like serine protease from Vibrio cholerae.

  •  2019 Jun. Borrelia burgdorferi peptidoglycan is a persistent antigen in patients with Lyme arthritis.

  • Nature. 2019 Jun. Mapping human microbiome drug metabolism by gut bacteria and their genes.

  • Cell. 2019 May. Nucleoid Size Scaling and Intracellular Organization of Translation across Bacteria.

  • Cell. 2019 Apr. See also The New York TimesWired Bacteria Form Nature’s Power Grid: ‘We Have an Electric Planet’Structure of microbial nanowires reveals stacked hemes that transport electrons over micrometers.

  •  2019 Mar. Structural dynamics of bacteriophage P22 infection initiation revealed by cryo-electron tomography.

  •  2019 Feb. Available energy fluxes drive a transition in the diversity, stability, and functional structure of microbial communities.

  •  2019. Activity-Based Protein Profiling at the Host-Pathogen Interface.

  •  2019 Feb. Separating host and microbiome contributions to drug pharmacokinetics and toxicity.

  •  2018 Dec. Dietary sugar silences a colonization factor in a mammalian gut symbiont.

  •  2018 Nov. Cryo-electron tomography of periplasmic flagella in Borrelia burgdorferi reveals a distinct cytoplasmic ATPase complex.

  •  2018 Nov. De novo design of self-assembling helical protein filaments.

  •  2018 Nov. Mucosal homeostasis is altered in the ileum of gnotobiotic mice.
  •  2018 Nov. A tetratricopeptide repeat domain protein has profound effects on assembly of periplasmic flagella, morphology and motility of the lyme disease spirochete Borrelia burgdorferi.

  •  2018 Oct. On the deformability of an empirical fitness landscape by microbial evolution.

  •  2018 Oct. Visualization of the type III secretion mediated Salmonella-host cell interface using cryo-electron tomography.

  •  2018 Oct. Characterization of fluorescent proteins, promoters, and selectable markers for applications in the Lyme disease spirochete Borrelia burgdorferi.

  • Curr Top Microbiol Immunol. 2018 Sep. Activity-Based Protein Profiling at the Host-Pathogen Interface.

  • Elife.  2018 Sep. Human gut Bacteroides capture vitamin B12 via cell surface-exposed lipoproteins.

  •  2018 Aug. Emergent simplicity in microbial community assembly.

  •  2018 Jul. protein that controls the onset of a Salmonella virulence program.

  •  2018 Jul. The Stringent Response Determines the Ability of a Commensal Bacterium to Survive Starvation and to Persist in the Gut.

  •  2018 Jun. Genomewide phenotypic analysis of growthcell morphogenesis, and cell cycle events in Escherichia coli.

  •  2018 Jun. unique cytoplasmic ATPase complex defines the Legionella pneumophila type IV secretionchannel.

  •  2018 May. Reduction in adaptor amounts establishes degradation hierarchy among protease substrates.

  •  2018 May. Distribution of Initiation Times Reveals Mechanisms of Transcriptional Regulation in Single Cells.

  •  2018 May. Topical application of aminoglycoside antibiotics enhances host resistance to viral infections in a microbiota-independent manner.

  • Cell. 2018 Mar. Subcellular Organization: A Critical Feature of Bacterial Cell Replication.