Human genome-wide RNAi screen identifies an essential role for inositol pyrophosphates in Type-I interferon response
Authors: Andrew M Riley
Jeremy D Weaver
Sajith Nair
Solomon H Snyder
Ramnik J Xavier
Kathleen McCaffrey
Pradeep D Uchil
Adolfo García-Sastre
Pradeep Bist
John D York
Rongtuan Lin
Manoj N Krishnan
Richa Tyagi
Stephen B Shears
Aleksandar D Kostic
Barry V L Potter
Niyas Kudukkil Pulloor
Journal: PLoS Pathogens (2014)
Discipline: MICROBIOLOGY
Keywords: RIG-I; interferon production; RNAi-screen; inositol pyrophosphate; IPPK
Affiliations: Program on Emerging Infectious Diseases, DUKE-NUS Graduate Medical School, Singapore., Center for Computational and Integrative Biology, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts, United States of America., Inositol Signaling Group, Laboratory of Signal Transduction, National Institute of Environmental Health Sciences, NIH, DHHS, Research Triangle Park, North Carolina, United States of America., Department of Pharmacy and Pharmacology, University of Bath, Claverton Down, Bath, United Kingdom., Solomon H. Snyder Department of Neuroscience and Departments of Psychiatry and Behavioral Sciences, The Johns Hopkins University School of Medicine, Baltimore, Maryland, United States of America., Section of Microbial Pathogenesis, Yale University School of Medicine, New Haven, Connecticut, United States of America., Department of Biochemistry, Vanderbilt University Medical Center, Nashville, Tennessee, United States of America., Department of Microbiology, Global Health and Emerging Pathogens Institute, Department of Medicine, Division of Infectious Diseases, Icahn School of Medicine at Mount Sinai, New York, New York, United States of America., Lady Davis Institute for Medical Research, Jewish General Hospital, Montreal, Canada.

Abstract:  The pattern recognition receptor RIG-I is critical for Type-I interferon production. However, the global regulation of RIG-I signaling is only partially understood. Using a human genome-wide RNAi-screen, we identified 226 novel regulatory proteins of RIG-I mediated interferon-β production. Furthermore, the screen identified a metabolic pathway that synthesizes the inositol pyrophosphate 1-IP7 as a previously unrecognized positive regulator of interferon production. Detailed genetic and biochemical experiments demonstrated that the kinase activities of IPPK, PPIP5K1 and PPIP5K2 (which convert IP5 to1-IP7) were critical for both interferon induction, and the control of cellular infection by Sendai and influenza A viruses. Conversely, ectopically expressed inositol pyrophosphate-hydrolases DIPPs attenuated interferon transcription. Mechanistic experiments in intact cells revealed that the expression of IPPK, PPIP5K1 and PPIP5K2 was needed for the phosphorylation and activation of IRF3, a transcription factor for interferon. The addition of purified individual inositol pyrophosphates to a cell free reconstituted RIG-I signaling assay further identified 1-IP7 as an essential component required for IRF3 activation. The inositol pyrophosphate may act by β-phosphoryl transfer, since its action was not recapitulated by a synthetic phosphonoacetate analogue of 1-IP7. This study thus identified several novel regulators of RIG-I, and a new role for inositol pyrophosphates in augmenting innate immune responses to viral infection that may have therapeutic applications. 

Retracted: No

PubPeer comments:
 The authors investigated wettability control of silica nanoparticles by first synthesizing silica nanoparticles using the Stu00f6ber method, followed by modifying it with different amounts of dichlorodimethylsilane (DCDMS) in the presence of water. The degree of modification (grafting) was evaluated by inferring OH concentration from titration and thermogravimetric analysis. A film was formed from these modified particles and water contact angle (CA) on these surfaces were measured, where the results indicate strong dependence of CA on the degree of modification. Here are a number of comments and questions. 1) The major contribution of this work is surface wettability control via modification. The water CA was measured on the thin films formed from the modified particles but details were not given. How the films were fabricated ? drop-casted ? What is the solvent . The major issue is that roughness of surfaces also has significant effect on the CA, as indicated by Cassie-Baxter equations. Please refer to this reference, especially Table 2. https: doi.org 10.1016 j.jcis.2010.08.003 2) With regard to inferring the silanol concentration via titration. Sodium hexametophosphate was added, how much? Does not it also react with HCl, blur the titration result? The surface area of the particles are also invoked in the equation (1). How was it measured ? using BET ? what is the number ? How does the dispersity of the particles affect the extrapolated OH number concentration ? Either acidic or basic solutions can reverse the grafting reaction. How does these impact the measured OH number ? 3) The authors claimed that the particle size is 30 nm from SEM images of Figure 1. It is not possible from this image, an TEM imaged is needed. 4) The authors used FTIR spectrum to indicate modification. The most significant peak and change is between 3600 - 3800 wavenumber, the OH vibration of silanol, and vibration of water (product). However, the spectral data the authors provided do not show any change. Then how do you claim successful modification ? Please refer to the following reference for the FTIR spectra. https: doi.org 10.1021 acs.langmuir.9b03223 5) The proposed mechanism is flawed. The hydrolyzed DCDMS has OH group at both ends. Whether it reacts via self-condensation or graft to silica, one end always would be bare OH. Then how do you explain the wettability change from hydrophilic to hydrophobic, not to mention to superhydrophobic ? Besides, even if a hydrophobic molecule, such as CH3(CH2)nCOOH, is grafted to a surface, the alkyl chain length (n) has significant impact on the hydrophobicity. Please refer to the following reference. https: doi.org 10.1039 C6PY01596D ,