Spotlight pubs.acs.org/acschemicalbiology
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NOVEL MECHANISM FOR QUORUM SENSING INHIBITION
Pqs pathway, which had the downstream effect of suppressing pyocyanin production. The authors conclude that because of the crosstalk between the Rh1 and Pqs pathways, it may be necessary to target both pathways simultaneously to effectively control P. aeruginosa virulence. Heidi A. Dahlmann, Ph.D.
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Safavi-Hemami, H., et al. Proc. Natl. Acad. Sci. U.S.A., DOI: 10.1073/pnas.1423857112. Copyright 2015 National Academy of Sciences, U.S.A.
Reprinted with permission from Welsh, M. A. et al. J. Am. Chem. Soc., DOI: 10.1021/ja5110798. Copyright 2015 American Chemical Society.
Safavi-Hemami, H., et al. Proc. Natl. Acad. Sci. U.S.A., DOI: 10.1073/pnas.1423857112. Copyright 2015 National Academy of Sciences, U.S.A.
Microorganisms produce small molecules that mediate intercellular bacterial communication, a process also known as quorum sensing, which in turn controls the behavior of the entire bacterial population, especially with respect to biofilm production and bacterial virulence. Currently, quorum sensing is of particular interest as a possible target for controlling bacterial infections in immunocompromised patients. However, efforts to manipulate quorum sensing are complicated because quorum sensing is mediated by a variety of intracellular biochemical pathways that can simultaneously induce or inhibit each other. Recently, Helen E. Blackwell and co-workers studied the effect of small molecules on the virulence phenotype of the opportunistic pathogen Pseudomonas aeruginosa and uncovered new facets of how complementary quorum sensing systems in P. aeruginosa interact (J. Am. Chem. Soc. 2014, DOI: 10.1021/ja5110798). In P. aeruginosa, a protein called Rh1 produces N-butyryl-Lhomoserine lactone (BHL) to act as a ligand for the P. aeruginosa Rhl receptor protein (Rh1R). Upon ligand binding, Rh1R acts as transcription factor for genes controlling biosynthesis of pyocyanin, an inducer of oxidative stress and inflammation, and rhamnolipid, which facilitates biofilm formation and immune evasion. Knowing that rh1R-knockdown or mutation abolishes pyocyanin and rhamnolipid biosynthesis, respectively, Blackwell and coworkers hypothesized that Rh1R antagonists would inhibit pyocyanin and rhamnolipid production while Rh1R agonists would induce their production. However, when the team exposed P. aeruginosa to a series of BHL analogs comprised of Rh1R agonists and antagonists, they obtained conflicting results: while their hypothesis proved true for the effect of Rh1R agonists and antagonists on rhamnolipid biosynthesis, the converse was true for pyocyanin production. In fact, Rh1R agonists unexpectedly downregulated an enzyme belonging to a separate quorum-sensing system, the © 2015 American Chemical Society
INSULINA CHEMICAL WEAPON
Insulin proteins are well-known to regulate metabolism and affect cognition in vertebrates. For invertebrates, insulins are involved in not just metabolic control but also in neuronal signaling, growth, and reproduction. A research team led by Helena Safavi-Hemami and Baldomero M. Olivera has just added a new role for invertebrate insulin to the list: chemical warfare (Proc. Natl. Acad. Sci. U.S.A., 2014, DOI: 10.1073/ pnas.1423857112). The team discovered insulins expressed in the venom glands of Conus geographus and Conus tulipa, two related species of “net-hunting” cone snails that release venom into their environment to dull the senses of small fish prior to their capture. Based on the tested activity of one of the insulins, they are presumed to mimic fish insulin, aiding in the cone snails’ hunting process. Among the newly discovered insulins, Con-Ins G1 (derived from C. geographus) and Con-Ins T1, T2, and T3 (derived from C. tulipa) were closely related to zebrafish insulin. In contrast, insulin could not be detected in five “hook and line” fishhunting cone snails that inject venom as they snag their prey. Six other mollusc- and worm-hunting cone snails were found to produce mollusc-like but no fish-like insulins in their venom glands. Altogether, the results indicated that venom-gland insulins appear to be targeted to each cone snail species’ mode of hunting and preferred prey. Mass-spectrometry studies of Con-Ins G1 revealed that the protein was subjected to post-translation modifications never before seen on insulins but similar to those observed on other C. geographus (or “cone snail venom peptides”) venom peptides. Furthermore, synthetically prepared Con-Ins G1 significantly lowered blood glucose levels following injection into hyperglycemic adult zebrafish and reduced locomotor activity in zebrafish upon direct application into the water column. The authors suggest that net-hunting cone Published: February 20, 2015 340
DOI: 10.1021/acschembio.5b00083 ACS Chem. Biol. 2015, 10, 340−342
ACS Chemical Biology
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snails release fish-like insulins to induce hypoglycemia and subsequent weakness in their prey, facilitating the capture of their next meal. Heidi A. Dahlmann, Ph.D.
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Spotlight
PHOSPHORYLATE AND FOLD
ZINC SPARKS EMBRYO DEVELOPMENT
Adapted by permission from Macmillan Publishers Ltd.: Nature Chemistry, Woodruff et al. 7, 130−139, Copyright 2014.
The maturation of female reproductive cells begins in the ovary when immature germinal vesical (GV) oocytes arrested in the prophase stage of meiosis I transform to fertilization-ready MII eggs arrested in metaphase of meiosis II. In mouse oocytes, this transformation is accompanied by a greater than 50% increase in total zinc content within several hours, a process that is necessary for maturation to occur. The massive buildup of zinc precedes a subsequent massive release of zinc from egg cells, areas that are fertilized or otherwise activated to divide. These zinc efflux events, which may occur several times per cell, are known as “zinc sparks.” Recently, Teresa K. Woodruff and co-workers used a novel fluorescent probe molecule and complementary spectroscopic techniques to pinpoint the exact source of zinc released during a zinc spark (Nature Chemistry 2014, 7, 130−139). The research team had previously observed that the spatial distribution of zinc sparks coincided with regions of zinc accumulation in the cellular periphery of unfertilized MII eggs, prompting the team to hypothesize that the zinc was released from discrete cortical compartments as opposed to being ejected from the cytosol through a membrane-bound transporter protein. Utilizing their fluorescent zinc probe (ZincBY-1) to detect labile zinc in live GV and MII cells, the team observed that over 90% of zinc-containing compartments were within 5 μm of the plasma membranes. These compartments appeared to colocalize with cortical granules, a type of exocyctotic vesicles that releases enzymes to block polyspermy after fertilization. Furthermore, by simultaneously monitoring intracellular and extracellular zinc levels during fertilization, the team confirmed that the disappearance of localized zinc compartments in oocyte membranes during a zinc spark occurred concomitantly with a sudden increase of zinc in the extracellular space immediately adjacent to the regions of zinc compartmentalization, confirming the hypothesis that zinc sparks emanate from zinc-rich cortical vesicles. Heidi A. Dahlmann, Ph.D.
Reprinted with permission from Macmillan Publishers Ltd.: Nature DOI: 10.1038/nature13999, Copyright 2015.
Reprinted with permission from Macmillan Publishers Ltd.: Nature DOI: 10.1038/nature13999, Copyright 2015.
Proteins can sometimes adopt two or more conformations, often acting as an elegant switch to turn the activity of a protein on or off. The switch can be triggered by many mechanisms, including binding of a small molecule, protein partner, or posttranslational modifications. The downstream effects of multiple protein folds are as diverse as their trigger, ranging from basic metabolism to the regulation of gene expression. Altering an already synthesized protein’s function has important implications for responding to cellular needs in a rapid and efficient manner. Now, Bah et al. (Nature 2014, DOI: 10.1038/nature13999) report a dramatic structural transition, switching an intrinsically disordered protein (IDP) to a folded domain in response to phosphorylation. The protein of interest, mammalian 4E-BP2, is a neural protein that serves a regulatory role by binding to the translation factor eIF4E, in turn suppressing translation initiation. Prior studies showed that 4E-BP2 phosphorylated at multiple sites is quite stable while the protein lacking these post-translational modifications is targeted for degradation, probably due to its IDP character. Using a combination of NMR and isothermal calorimetry, along with mutagenesis, the new study demonstrated that phosphorylation of the disordered 4E-BP2 induces folding of a four-stranded β-domain. The change was triggered by two threonine phosphorylations, with the modifications stabilizing multiple hydrogen bonds at the sharp β-turns. The net result of the structural acrobatics was a reduction in 4E-BP2’s affinity for eIF4E by nearly 2 orders of magnitude fold. Additional serine and threonine phosphorylation events then add another order of magnitude. The structural data indicate that ordering of the protein sequesters the canonical YXXXXLφ motif, critical to eIF4E binding, into a stable β-strand. While phosphorylation and other post-translational modifications often play a 341
DOI: 10.1021/acschembio.5b00083 ACS Chem. Biol. 2015, 10, 340−342
ACS Chemical Biology
Spotlight
critical role in the regulation of protein structure and function, this study unlocks a more dramatic effect than is usually seen in textbooks. Jason G. Underwood, Ph.D.
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DIGGING FOR NEW ANTIBIOTICS
Reprinted with permission from Macmillan Publishers Ltd.: Nature, DOI: 10.1038/nature14098, Copyright 2015.
The first antibiotics, many of which are still used in the clinic, were natural products isolated from bacteria or fungi. Taking inspiration from nature was a distinct advantage because these compounds had already evolved to penetrate the cell wall and kill a wide variety of bacteria. The disadvantage to this approach was that it usually required that the antibiotic-encoding organisms be readily grown under lab culture conditions, a tall order given that the vast majority of bacteria have not been successfully cultured. Now, Ling et al. (Nature 2015, DOI: 10.1038/nature14098) have searched for new therapeutics using an approach that is more friendly to unculturable bacteria. The multichannel iChip device was used to encapsulate single bacterial cells from a soil preparation. Then, instead of culturing in artificial media, the device was buried in soil, allowing colony formation inside the device but in more native surroundings. The extracts of 10 000 isolates were screened for antimicrobial properties using a plate of the notorious Staphylococcus aureus, a bacterium known to develop antibiotic resistance. One extract displayed good activity against this and many other pathogenic Grampositive species, so it was chosen for further analysis. The result was a novel Gram-negative bacterium related to the genus Aquabacteria, a group not previously known to produce antibiotics. The group solved the primary structure of the antibiotic, dubbed teixobactin, demonstrating that it is a depsipeptide, meaning that it has both amide and ester linkages. Genome sequencing of the isolate identified its 11 module biosynthetic cluster. Additionally, the mechanism of action was investigated and yielded a specific interaction with peptidoglycan precursors, the building blocks of the cell wall. This led to the group to postulate that evolving antibioticresistance to teixobactin would be particularly difficult, and they showed that no resistant strains of S. aureus arise over the course of 25 days cultured in low levels of the antibiotic. Teixobactin showed potent activity in three mouse models of infection, so this compound or a relative could someday make it into human care as well. Jason G. Underwood, Ph.D.
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DOI: 10.1021/acschembio.5b00083 ACS Chem. Biol. 2015, 10, 340−342
