Publications by authors named "Henry H Le"

15 Publications

  • Page 1 of 1

Dietary sphinganine is selectively assimilated by members of the mammalian gut microbiome.

J Lipid Res 2021 Feb 6:100034. Epub 2021 Feb 6.

Division of Nutritional Sciences, Cornell University, Ithaca, NY, USA. Electronic address:

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http://dx.doi.org/10.1194/jlr.RA120000950DOI Listing
February 2021

An Untargeted Approach for Revealing Electrophilic Metabolites.

ACS Chem Biol 2020 11 19;15(11):3030-3037. Epub 2020 Oct 19.

Boyce Thompson Institute and Department of Chemistry and Chemical Biology, Cornell University, Ithaca, New York 14853, United States.

Reactive electrophilic intermediates such as coenzyme A esters play central roles in metabolism but are difficult to detect with conventional strategies. Here, we introduce hydroxylamine-based stable isotope labeling to convert reactive electrophilic intermediates into stable derivatives that are easily detectable LC-MS. In the model system , parallel treatment with NHOH and NHOH revealed >1000 labeled metabolites, e.g., derived from peptide, fatty acid, and ascaroside pheromone biosyntheses. Results from NHOH treatment of a pheromone biosynthesis mutant, , suggested upregulation of thioesterase activity, which was confirmed by gene expression analysis. The upregulated thioesterase contributes to the biosynthesis of a specific subset of ascarosides, determining the balance of dispersal and attractive signals. These results demonstrate the utility of NHOH labeling for investigating complex biosynthetic networks. Initial results with and human cell lines indicate applicability toward uncovering reactive metabolomes in diverse living systems.
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http://dx.doi.org/10.1021/acschembio.0c00706DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7852410PMC
November 2020

Modular metabolite assembly in depends on carboxylesterases and formation of lysosome-related organelles.

Elife 2020 10 16;9. Epub 2020 Oct 16.

Boyce Thompson Institute and Department of Chemistry and Chemical Biology, Cornell University, Ithaca, United States.

Signaling molecules derived from attachment of diverse metabolic building blocks to ascarosides play a central role in the life history of and other nematodes; however, many aspects of their biogenesis remain unclear. Using comparative metabolomics, we show that a pathway mediating formation of intestinal lysosome-related organelles (LROs) is required for biosynthesis of most modular ascarosides as well as previously undescribed modular glucosides. Similar to modular ascarosides, the modular glucosides are derived from highly selective assembly of moieties from nucleoside, amino acid, neurotransmitter, and lipid metabolism, suggesting that modular glucosides, like the ascarosides, may serve signaling functions. We further show that carboxylesterases that localize to intestinal organelles are required for the assembly of both modular ascarosides and glucosides via ester and amide linkages. Further exploration of LRO function and carboxylesterase homologs in and other animals may reveal additional new compound families and signaling paradigms.
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http://dx.doi.org/10.7554/eLife.61886DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7641594PMC
October 2020

Deep Interrogation of Metabolism Using a Pathway-Targeted Click-Chemistry Approach.

J Am Chem Soc 2020 10 14;142(43):18449-18459. Epub 2020 Oct 14.

Boyce Thompson Institute and Department of Chemistry and Chemical Biology, Cornell University, Ithaca, New York 14853, United States.

Untargeted metabolomics indicates that the number of unidentified small-molecule metabolites may exceed the number of protein-coding genes for many organisms, including humans, by orders of magnitude. Uncovering the underlying metabolic networks is essential for elucidating the physiological and ecological significance of these biogenic small molecules. Here we develop a click-chemistry-based enrichment strategy, DIMEN (deep interrogation of metabolism via enrichment), that we apply to investigate metabolism of the ascarosides, a family of signaling molecules in the model organism . Using a single alkyne-modified metabolite and a solid-phase azide resin that installs a diagnostic moiety for MS/MS-based identification, DIMEN uncovered several hundred novel compounds originating from diverse biosynthetic transformations that reveal unexpected intersection with amino acid, carbohydrate, and energy metabolism. Many of the newly discovered transformations could not be identified or detected by conventional LC-MS analyses without enrichment, demonstrating the utility of DIMEN for deeply probing biochemical networks that generate extensive yet uncharacterized structure space.
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http://dx.doi.org/10.1021/jacs.0c06877DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7875204PMC
October 2020

Identification of Uric Acid Gluconucleoside-Ascaroside Conjugates in by Combining Synthesis and MicroED.

Org Lett 2020 09 21;22(17):6724-6728. Epub 2020 Aug 21.

Boyce Thompson Institute and Department of Chemistry and Chemical Biology, Cornell University, Ithaca, New York 14853, United States.

Few nucleoside-derived natural products have been identified from animals, despite the ubiquity of nucleosides in living organisms. Here, we use a combination of synthesis and the emerging electron microscopy technique microcrystal electron diffraction to determine the structures of several -(β-glucopyranosyl)uric acid derivatives in . These noncanonical gluconucleosides further integrate an ascaroside moiety, for which we present a shortened synthetic route. The production of a phosphorylated gluconucleoside is influenced by evolutionarily conserved insulin signaling.
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http://dx.doi.org/10.1021/acs.orglett.0c02038DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7526323PMC
September 2020

Dietary sphinganine is selectively assimilated by members of the mammalian gut microbiome.

J Lipid Res 2020 Jul 9. Epub 2020 Jul 9.

Division of Nutritional Sciences, Cornell University, United States

Functions of the gut microbiome have a growing number of implications for host metabolic health, with diet being one of the most significant influences on microbiome composition. Compelling links between diet and the gut microbiome suggest key roles for various macronutrients, including lipids, yet how individual classes of dietary lipids interact with the microbiome remains largely unknown. Sphingolipids are bioactive components of most foods and are also produced by prominent gut microbes. This makes sphingolipids intriguing candidates for shaping diet-microbiome interactions. Here, we used a click chemistry-based approach to track the incorporation of bioorthogonal dietary omega-alkynyl sphinganine (sphinganine alkyne [SAA]) into the murine gut microbial community (Bioorthogonal labeling). We identified microbial and SAA-specific metabolic products through fluorescence-based sorting of SAA-containing microbes (Sort), 16S rRNA gene sequencing to identify the sphingolipid-interacting microbes (Seq), and comparative metabolomics to identify products of SAA assimilation by the microbiome (Spec). Together, this approach, termed Bioorthogonal labeling-Sort-Seq-Spec (BOSSS), revealed that SAA assimilation is nearly exclusively performed by gut Bacteroides, indicating that sphingolipid-producing bacteria play a major role in processing dietary sphinganine. Comparative metabolomics of cecal microbiota from SAA-treated mice revealed conversion of SAA to a suite of dihydroceramides, consistent with metabolic activities of Bacteroides and Bifidobacterium. Additionally, other sphingolipid-interacting microbes were identified with a focus on an uncharacterized ability of Bacteroides and Bifidobacterium to metabolize dietary sphingolipids. We conclude that BOSSS provides a platform to study the flux of virtually any alkyne-labeled metabolite in diet-microbiome interactions.
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http://dx.doi.org/10.1194/jlr.RA120000950DOI Listing
July 2020

Going Keto? Say βHB-ye Bye to Your Gut Bifidobacteria.

Cell Host Microbe 2020 07;28(1):3-5

Division of Nutritional Sciences, Cornell University, Ithaca, NY 14853, USA. Electronic address:

Ketogenic diets (KDs) are popularly used to aid a myriad of conditions. KDs induce metabolic changes, but how microbiome alterations contribute to these changes remains unexplored. In a recent Cell paper, Ang et al. identify KD-specific changes to the gut microbiota linked to a reduction in pro-inflammatory Th17 cells.
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http://dx.doi.org/10.1016/j.chom.2020.06.012DOI Listing
July 2020

Liver-Type Glutaminase GLS2 Is a Druggable Metabolic Node in Luminal-Subtype Breast Cancer.

Cell Rep 2019 10;29(1):76-88.e7

Department of Molecular Medicine, Cornell University, Ithaca, NY 14853, USA; Department of Chemistry and Chemical Biology, Cornell University, Ithaca, NY 14853, USA. Electronic address:

Efforts to target glutamine metabolism for cancer therapy have focused on the glutaminase isozyme GLS. The importance of the other isozyme, GLS2, in cancer has remained unclear, and it has been described as a tumor suppressor in some contexts. Here, we report that GLS2 is upregulated and essential in luminal-subtype breast tumors, which account for >70% of breast cancer incidence. We show that GLS2 expression is elevated by GATA3 in luminal-subtype cells but suppressed by promoter methylation in basal-subtype cells. Although luminal breast cancers resist GLS-selective inhibitors, we find that they can be targeted with a dual-GLS/GLS2 inhibitor. These results establish a critical role for GLS2 in mammary tumorigenesis and advance our understanding of how to target glutamine metabolism in cancer.
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http://dx.doi.org/10.1016/j.celrep.2019.08.076DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6939472PMC
October 2019

Diketopiperazine Formation in Fungi Requires Dedicated Cyclization and Thiolation Domains.

Angew Chem Int Ed Engl 2019 10 28;58(41):14589-14593. Epub 2019 Aug 28.

Boyce Thompson Institute and Department of Chemistry and Chemical Biology, Cornell University, Ithaca, NY, USA.

Cyclization of linear dipeptidyl precursors derived from nonribosomal peptide synthetases (NRPSs) into 2,5-diketopiperazines (DKPs) is a crucial step in the biosynthesis of a large number of bioactive natural products. However, the mechanism of DKP formation in fungi has remained unclear, despite extensive studies of their biosyntheses. Here we show that DKP formation en route to the fungal virulence factor gliotoxin requires a seemingly extraneous couplet of condensation (C) and thiolation (T) domains in the NRPS GliP. In vivo truncation of GliP to remove the CT couplet or just the T domain abrogated production of gliotoxin and all other gli pathway metabolites. Point mutation of conserved active sites in the C and T domains diminished cyclization activity of GliP in vitro and abolished gliotoxin biosynthesis in vivo. Verified NRPSs of other fungal DKPs terminate with similar CT domain couplets, suggesting a conserved strategy for DKP biosynthesis by fungal NRPSs.
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http://dx.doi.org/10.1002/anie.201909052DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6764874PMC
October 2019

Linking Genomic and Metabolomic Natural Variation Uncovers Nematode Pheromone Biosynthesis.

Cell Chem Biol 2018 06 17;25(6):787-796.e12. Epub 2018 May 17.

Department for Evolutionary Biology, Max Planck Institute for Developmental Biology, 72076 Tübingen, Germany. Electronic address:

In the nematodes Caenorhabditis elegans and Pristionchus pacificus, a modular library of small molecules control behavior, lifespan, and development. However, little is known about the final steps of their biosynthesis, in which diverse building blocks from primary metabolism are attached to glycosides of the dideoxysugar ascarylose, the ascarosides. We combine metabolomic analysis of natural isolates of P. pacificus with genome-wide association mapping to identify a putative carboxylesterase, Ppa-uar-1, that is required for attachment of a pyrimidine-derived moiety in the biosynthesis of ubas#1, a major dauer pheromone component. Comparative metabolomic analysis of wild-type and Ppa-uar-1 mutants showed that Ppa-uar-1 is required specifically for the biosynthesis of ubas#1 and related metabolites. Heterologous expression of Ppa-UAR-1 in C. elegans yielded a non-endogenous ascaroside, whose structure confirmed that Ppa-uar-1 is involved in modification of a specific position in ascarosides. Our study demonstrates the utility of natural variation-based approaches for uncovering biosynthetic pathways.
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http://dx.doi.org/10.1016/j.chembiol.2018.04.004DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6014897PMC
June 2018

Antimicrobial Activity of Ibuprofen against Cystic Fibrosis-Associated Gram-Negative Pathogens.

Antimicrob Agents Chemother 2018 03 23;62(3). Epub 2018 Feb 23.

Department of Microbial Pathogenesis and Immunology, Texas A&M University Health Science Center, College Station, Texas, USA

Clinical trials have demonstrated the benefits of ibuprofen therapy in cystic fibrosis (CF) patients, an effect that is currently attributed to ibuprofen's anti-inflammatory properties. Yet, a few previous reports demonstrated an antimicrobial activity of ibuprofen as well, although none investigated its direct effects on the pathogens found in the CF lung, which is the focus of this work. Determination of ibuprofen's antimicrobial activity against and species strains through measurements of the endpoint number of CFU and growth kinetics showed that ibuprofen reduced the growth rate and bacterial burden of the tested strains in a dose-dependent fashion. In an biofilm model, a reduction in the rate of biomass accumulation over 8 h of growth with ibuprofen treatment was observed. Next, an acute pneumonia model was used to test this antimicrobial activity after the oral delivery of ibuprofen. Following intranasal inoculation, ibuprofen-treated mice exhibited lower CFU counts and improved survival compared with the control animals. Preliminary biodistribution studies performed after the delivery of ibuprofen to mice by aerosol demonstrated a rapid accumulation of ibuprofen in serum and minimum retention in lung tissue and bronchoalveolar lavage fluid. Therefore, ibuprofen-encapsulated polymeric nanoparticles (Ibu-NPs) were formulated to improve the pharmacokinetic profile. Ibu-NPs formulated for aerosol delivery inhibited the growth of and may provide a convenient dosing method. These results provide an additional explanation for the previously observed therapeutic effects of ibuprofen in CF patients and further strengthen the argument for its use by these patients.
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http://dx.doi.org/10.1128/AAC.01574-17DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5826130PMC
March 2018

Biosynthesis of Modular Ascarosides in C. elegans.

Angew Chem Int Ed Engl 2017 04 28;56(17):4729-4733. Epub 2017 Mar 28.

Boyce Thompson Institute and Department of Chemistry and Chemical Biology, Cornell University, Ithaca, NY, USA.

The nematode Caenorhabditis elegans uses simple building blocks from primary metabolism and a strategy of modular assembly to build a great diversity of signaling molecules, the ascarosides, which function as a chemical language in this model organism. In the ascarosides, the dideoxysugar ascarylose serves as a scaffold to which diverse moieties from lipid, amino acid, neurotransmitter, and nucleoside metabolism are attached. However, the mechanisms that underlie the highly specific assembly of ascarosides are not understood. We show that the acyl-CoA synthetase ACS-7, which localizes to lysosome-related organelles, is specifically required for the attachment of different building blocks to the 4'-position of ascr#9. We further show that mutants lacking lysosome-related organelles are defective in the production of all 4'-modified ascarosides, thus identifying the waste disposal system of the cell as a hotspot for ascaroside biosynthesis.
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http://dx.doi.org/10.1002/anie.201700103DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5486983PMC
April 2017

Plant-like biosynthesis of isoquinoline alkaloids in Aspergillus fumigatus.

Nat Chem Biol 2016 06 11;12(6):419-24. Epub 2016 Apr 11.

Boyce Thompson Institute and Department of Chemistry and Chemical Biology, Cornell University, Ithaca, New York, USA.

Natural product discovery efforts have focused primarily on microbial biosynthetic gene clusters (BGCs) containing large multimodular polyketide synthases and nonribosomal peptide synthetases; however, sequencing of fungal genomes has revealed a vast number of BGCs containing smaller NRPS-like genes of unknown biosynthetic function. Using comparative metabolomics, we show that a BGC in the human pathogen Aspergillus fumigatus named fsq, which contains an NRPS-like gene lacking a condensation domain, produces several new isoquinoline alkaloids known as the fumisoquins. These compounds derive from carbon-carbon bond formation between two amino acid-derived moieties followed by a sequence that is directly analogous to isoquinoline alkaloid biosynthesis in plants. Fumisoquin biosynthesis requires the N-methyltransferase FsqC and the FAD-dependent oxidase FsqB, which represent functional analogs of coclaurine N-methyltransferase and berberine bridge enzyme in plants. Our results show that BGCs containing incomplete NRPS modules may reveal new biosynthetic paradigms and suggest that plant-like isoquinoline biosynthesis occurs in diverse fungi.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5049701PMC
http://dx.doi.org/10.1038/nchembio.2061DOI Listing
June 2016

Single-cell control of initial spatial structure in biofilm development using laser trapping.

Langmuir 2014 Apr 9;30(15):4522-30. Epub 2014 Apr 9.

Center for Nonlinear Dynamics and Department of Physics and ‡School of Biological Sciences, The University of Texas at Austin , Austin, Texas 78712, United States.

Biofilms are sessile communities of microbes that are spatially structured by an embedding matrix. Biofilm infections are notoriously intractable. This arises, in part, from changes in the bacterial phenotype that result from spatial structure. Understanding these interactions requires methods to control the spatial structure of biofilms. We present a method for growing biofilms from initiating cells whose positions are controlled with single-cell precision using laser trapping. The native growth, motility, and surface adhesion of positioned microbes are preserved, as we show for model organisms Pseudomonas aeruginosa and Staphylococcus aureus. We demonstrate that laser-trapping and placing bacteria on surfaces can reveal the effects of spatial structure on bacterial growth in early biofilm development.
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http://dx.doi.org/10.1021/la500128yDOI Listing
April 2014

The extracellular polysaccharide Pel makes the attachment of to surfaces symmetric and short-ranged.

Soft Matter 2013 Apr;9(14):3871-3876

Center for Nonlinear Dynamics and Department of Physics, University of Texas at Austin, 2515 Speedway, C1610, Austin, TX 78712, USA.

Biofilms are surface-mounted, multicellular communities of microbes. Biofilms are often associated with chronic infections that resist treatment, evade the immune system, and damage host tissue. An essential characteristic of the biofilm state is that constituent organisms are bound in a polymeric matrix. This matrix gives the system spatial structure and clusters bacteria near each other, facilitating intercellular interactions. The strain PAO1 is widely studied as a model biofilm-forming organism. The polymeric matrix of PAO1 biofilms is dominated by two bacteria-produced extracellular polymers, Pel and Psl. We use a combination of optical and atomic force microscopy to examine the roles of these polymers in very early biofilm development. In agreement with other researchers, we find that Psl mediates strong attachment to a glass surface. We find that Pel alone can mediate some attachment, but not as permanent as that mediated by Psl. Unexpectedly, we find that Pel promotes symmetric attachment, in the form of rod-shaped bacteria lying down flat on the surface, and that the presence of Pel makes attachment forces more short-ranged than they are with Psl alone. We suggest that these effects may result from synergistic interactions of Pel with the Psl polymeric matrix.
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http://dx.doi.org/10.1039/C3SM27638DDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3719985PMC
April 2013