Spotlight pubs.acs.org/crt
■
BROAD-SPECTRUM ANTIBIOTICS CAN EXACERBATE GRAFT-VERSUS-HOST DISEASE Patients undergoing allogeneic hematopoietic stem cell transplantation are at risk for systemic bacterial infections, which are treated with empiric antibiotics. These patients are additionally at risk for graft-versus-host disease (GVHD), an inflammatory condition that arises from donor cells attacking recipient tissues. As intestinal commensal bacteria can modulate GVHD severity, Marcel R. M. van den Brink, Robert R. Jenq, Yusuke Shono, and their collaborators studied the downstream effects of broadspectrum antibiotics on GVHD severity ((2016) Sci. Transl. Med., 8:339ra71). The authors retrospectively examined records of 857 allogeneic hematopoietic stem cell transplant recipients treated over 23 years at one institution. Of 12 frequently administered antibiotics, two drugs were significantly associated with increased GVHD-related mortality: piperacillin-tazobactam, a standard first-line antibiotic, and imipenem-cilastatin, a second-line treatment. Both drugs are highly active against obligate anaerobes. Conversely, aztreonam and cefepime, two first-line antibiotics with reduced activity against anaerobes, were associated with a decreased risk of GVHD-related mortality. Systemically treating healthy mice for 2 days with either of the first two drugs, but not with the latter ones, resulted in a reduction of major anaerobic bacteria called Clostridiales, which prevent inflammation. Further murine experiments demonstrated that piperacillin-tazobactam or imipenem-cilastatin treatment aggravated GVHD severity after 2 weeks of administration. When compared to animals treated with aztreonam, those treated with imipenem-cilastatin had a shortened survival time along with increased GVHD pathology in the colon and elevated numbers of donor CD4+ T cells, granulocytes, and interleukin-23 in the colon. These mice showed lower abundance of Clostridiales and at the same time contained microbiota with enriched abilities to degrade mucus. Indeed, mice treated with imipenem-cilastatin showed a marked reduction in the thickness of the protective colonic inner mucus barrier. This study demonstrates that using antibiotics with more limited anaerobic activity to treat transplant patients could decrease injury to beneficial intestinal microbiota and, subsequently, reduce GVHD severity and mortality. Abigail Druck Shudofsky
■
IMMUNE RESPONSES TO ZIKA VIRUS MAY BE IMPACTED BY PREEXISTING IMMUNITY TO DENGUE VIRUS
specifically bind and potently neutralize Zika virus in vitro. To further examine the immune cross-reactivity, they analyzed 47 dengue virus-specific monoclonal antibodies isolated from patients during acute dengue virus infection for reactivity against Zika virus. While 26 of them bound with high affinity to Zika virus, just seven were able to neutralize it in vitro, and only one was able to do so potently. This suggests that while crossreactivity to Zika virus is more broad, the potential of dengueinduced antibodies to protect against Zika virus may be limited. The scientists next investigated whether, rather than providing protection, pre-existing dengue-induced antibodies could enhance Zika virus infection and lead to a higher disease severity. They found that, indeed, the dengue-immune sera and Zika-reactive monoclonal antibodies enhanced Zika virus infection in vitro. These results suggest that individuals who have been previously infected with dengue virus might have different immune responses to Zika virus infection than individuals who are ̈ As the current Zika outbreak is largely occurring flavivirus-naive. in dengue-endemic areas, it is critical that this issue be investigated further. Abigail Druck Shudofsky
Adapted from Priyamvada, L., Quicke, K.M., Hudson, W.H., Onlamoon, N., Sewatanon, J., Edupuganti, S., Pattanapanyasat, K., Chokephaibulkit, K., Mulligan, M.J., Wilson, P.C., Ahmed, R., Suthar, M.S., and Wrammert, J. Human antibody responses after dengue virus infection are highly cross-reactive to Zika virus. PNAS, 113 (28), 7852−7857.
■
NOVEL APPROACH FOR IDENTIFYING THE MECHANISTIC ACTIVITY OF ANTIMICROBIALS The increase in antibiotic resistance necessitates the development of antimicrobials with novel mechanisms of action. Previously, Joe Pogliano, Kit Pogliano, and their teams developed a rapid assay to identify the mechanism of action of antibiotics. This method, bacterial cytological profiling (BCP), uses fluorescent microscopy to compare cytological profiles of cells treated with new compounds to those treated with known antibiotics to determine the pathways targeted by the novel
Zika virus is a member of the Flaviviridae family. Prior exposure to other flaviviruses, such as dengue virus, makes serology-based diagnosis of Zika virus infection more difficult due to antibody cross-reactivity; the envelope protein of dengue virus, which is an antibody target, shares a high structural and sequence homology with the envelope protein of Zika virus (see figure). Rafi Ahmed, Mehul S. Suthar, Jens Wrammert, and their collaborators investigated whether pre-existing antibodies induced by dengue virus affect the immune response generated by Zika virus infection ((2016) PNAS, doi: 10.1073/pnas.1607931113). The authors collected sera from nine patients infected with dengue virus and found that the polyclonal sera could © 2016 American Chemical Society
Published: August 15, 2016 1223
DOI: 10.1021/acs.chemrestox.6b00251 Chem. Res. Toxicol. 2016, 29, 1223−1224
Chemical Research in Toxicology
Spotlight
inhibitor that reduced neuronal cholesterol concentrations. Additionally, monoacylation of PrPC prevented its localization to lipid rafts and its accumulation in the synapses. The researchers found that PrPC expressed by glial cells was not targeted to synapses. They investigated the composition of the GPI anchors attached to glial and neuronal PrPC and found that only neuronal PrPC was attached to a GPI containing a terminal sialic acid. After neuraminidase digestion, desialylated PrPC was distributed to neuronal lipid rafts but did not accumulate in the synapses. When the authors conjugated rabbit IgG to either GPI derived from neuronal PrPC or desialylated GPI, both fusion proteins bound to neurons but only IgG-GPI was found at the synapse, indicating that the sialic acid on the neuronal PrPC GPI anchor is responsible for synaptic targeting. This research demonstrates that PrPC synaptic targeting requires a normal concentration of cholesterol in the cell membrane and a GPI anchor containing two acyl chains and sialic acid. Abigail Druck Shudofsky
drugs. However, compounds that act against new targets will produce unique cytological profiles. As such, the researchers have now developed a rapid inhibition profiling (RIP) method that targets specific bacterial proteins, facilitating identification of the mechanistic activity of novel inhibitors ((2016) ACS Chem. Biol., doi: 10.1021/acschembio.5b01050). The authors found that degrading essential proteins in a pathway resulted in cytological profiles that mimic those produced by antibiotics targeting the same pathway. As such, they tagged target proteins with a proteasomal degradation sequence recognized by a protein regulated by an inducible promotor. They then generated postdegradation cytological profiles which they compared visually and quantitatively to other profiles using BCP. The researchers found that RIP profiles after degradation of proteins involved in DNA synthesis and transcription were similar, respectively, to the cytological profiles of cells treated with compounds that inhibit DNA replication and RNA polymerase activity. Similarly, the RIP profiles after degradation of different enzymes in the fatty acid and peptidoglycan biosynthesis pathways matched the cytological profiles seen after the use of antibiotics targeting steps in those processes. With this method, the profile of a cell treated with a compound that has an unknown mechanism of action can be matched to the degradation profile of a known protein, thus identifying the novel drug target. Combining RIP and BCP will enable researchers to identify the mechanistic activity of new antimicrobial molecules. Abigail Druck Shudofsky
■
■
CHEMICALLY REPROGRAMMING NONCARDIAC CELLS INTO FUNCTIONAL CARDIOMYOCYTES Sheng Ding, Deepak Srivastava, and their teams identified a combination of small molecules that efficiently reprograms human fibroblasts into functional cardiomyocytes ((2016) Science, 352, 1216−1220). This chemical reprogramming approach introduces compounds into a starting cell type where they interact with endogenous cellular factors without target cell type-specific proteins. As such, it may be modified to generate various cell types. The scientists screened 89 small molecules known to facilitate cellular reprogramming, adding them individually to a baseline four-compound cocktail which permits cardiac reprogramming in fibroblasts when combined with a specific gene. Fibroblast cells were treated with the five compounds for 6 days, then cultured in cardiac induction medium with cardiogenic molecules. The researchers identified compounds that enhanced cardiogenic gene expression and further assessed their efficacy in different combinations (ungrouping the baseline compounds). They identified seven indispensable compounds that, when combined, were sufficient for efficient induction of cardiac reprogramming; when used together on fibroblasts, they generated cellular clusters that beat spontaneously. Additionally, the authors identified two growth factor inhibitors that sped up fibroblast gene downregulation and yielded more clusters of beating cells. 97% of cardiomyocytes induced by these nine compounds were functional in vitro, contracting uniformly. After reprogramming, the chemically induced cardiomyocytes displayed an upregulation of genes related to heart development and cardiomyocyte function and downregulation of genes associated with fibroblast function. Their electrophysical and epigenetic properties were similar to those of human cardiomyocytes generated from pluripotent stem cells. Importantly, when transplanted into diseased mouse hearts, the reprogrammed cells matured into cardiomyocytes in vivo. The authors found that the nine compounds mechanistically facilitate cardiac reprogramming by decondensing chromatin at key regions for cardiogenesis, increasing the accessibility of DNA and allowing for cardiogenic signal effectors to bind to promotors and enhancers of heart development genes. Abigail Druck Shudofsky
SYNAPTIC TARGETING OF THE CELLULAR PRION PROTEIN DEPENDS ON THE STRUCTURE OF THE ATTACHED GLYCOSYLPHOSPHATIDYLINOSITOL ANCHOR
This research was originally published in Journal of Biological Chemistry. Bate, C., Nolan, W., McHale-Owen, H., and Williams, A. (2016) Sialic Acid within the Glycosylphosphatidylinositol Anchor Targets the Cellular Prion Protein to Synapses. JBC, doi: 10.1074/jbc.M116.731117. Copyright The American Society for Biochemistry and Molecular Biology.
The cellular prion protein (PrPC) is the normal isoform of the protein associated with transmissible spongiform encephalopathies. Neuronal PrPC is found in cholesterol-rich lipid rafts, where it is linked to the cellular membrane by a glycosylphosphatidylinositol (GPI) anchor (see figure). The protein is subsequently transported to the synapse, and a team led by Alun Williams investigated and identified the mechanism of PrPC synaptic targeting ((2016) J. Biol. Chem., doi: 10.1074/jbc.M116.731117). The scientists introduced PrPC into cultured neurons, where it concentrated in the synapses. They found that this synaptic targeting was reduced in a dose-dependent manner by use of an 1224
DOI: 10.1021/acs.chemrestox.6b00251 Chem. Res. Toxicol. 2016, 29, 1223−1224
