Screening for Novel Small-Molecule Inhibitors Targeting the Assembly of Influenza Virus Polymerase Complex by a Bimolecular Luminescence Complementation-Based Reporter System

ABSTRACT Influenza virus RNA-dependent RNA polymerase consists of three viral protein subunits: PA, PB1, and PB2. Protein-protein interactions (PPIs) of these subunits play pivotal roles in assembling the functional polymerase complex, which is essential for the replication and transcription of influenza virus RNA. Here we developed a highly specific and robust bimolecular luminescence complementation (BiLC) reporter system to facilitate the investigation of influenza virus polymerase complex formation. Furthermore, by combining computational modeling and the BiLC reporter assay, we identified several novel small-molecule compounds that selectively inhibited PB1-PB2 interaction. Function of one such lead compound was confirmed by its activity in suppressing influenza virus replication. In addition, our studies also revealed that PA plays a critical role in enhancing interactions between PB1 and PB2, which could be important in targeting sites for anti-influenza intervention. Collectively, these findings not only aid the development of novel inhibitors targeting the formation of influenza virus polymerase complex but also present a new tool to investigate the exquisite mechanism of PPIs. IMPORTANCE Formation of the functional influenza virus polymerase involves complex protein-protein interactions (PPIs) of PA, PB1, and PB2 subunits. In this work, we developed a novel BiLC assay system which is sensitive and specific to quantify both strong and weak PPIs between influenza virus polymerase subunits. More importantly, by combining in silico modeling and our BiLC assay, we identified a small molecule that can suppress influenza virus replication by disrupting the polymerase assembly. Thus, we developed an innovative method to investigate PPIs of multisubunit complexes effectively and to identify new molecules inhibiting influenza virus polymerase assembly.

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PL-15 Modulation of influenza virus replication and virulence by viral-host protein interactions

Cytokine ◽

10.1016/j.cyto.2008.07.329 ◽

2008 ◽

Vol 43 (3) ◽

pp. 302

Author(s):

Adolfo García-Sastre

Keyword(s):

Influenza Virus ◽

Virus Replication ◽

Protein Interactions ◽

Host Protein ◽

Influenza Virus Replication

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A small molecule screen in yeast identifies inhibitors targeting protein–protein interactions within the vaccinia virus replication complex

Antiviral Research ◽

10.1016/j.antiviral.2012.07.010 ◽

2012 ◽

Vol 96 (2) ◽

pp. 187-195 ◽

Cited By ~ 8

Author(s):

Olivier Flusin ◽

Laurent Saccucci ◽

Céline Contesto-Richefeu ◽

Amel Hamdi ◽

Carine Bardou ◽

...

Keyword(s):

Vaccinia Virus ◽

Virus Replication ◽

Small Molecule ◽

Protein Interactions ◽

Protein Protein Interactions ◽

Replication Complex ◽

Small Molecule Screen

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Faculty Opinions recommendation of Drosophila RNAi screen identifies host genes important for influenza virus replication.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1117828.573915 ◽

2008 ◽

Author(s):

Adolfo Garcia-Sastre

Keyword(s):

Influenza Virus ◽

Virus Replication ◽

Rnai Screen ◽

Influenza Virus Replication ◽

Host Genes

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Faculty Opinions recommendation of The lipid mediator protectin D1 inhibits influenza virus replication and improves severe influenza.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.718018306.793478156 ◽

2013 ◽

Author(s):

Shizuo Akira ◽

Tatsuya Saitoh

Keyword(s):

Influenza Virus ◽

Virus Replication ◽

Lipid Mediator ◽

Severe Influenza ◽

Influenza Virus Replication

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Faculty Opinions recommendation of Importance of Rigidity in Designing Small Molecule Drugs To Tackle Protein-Protein Interactions (PPIs) through Stabilization of Desired Conformers.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.732129727.793539183 ◽

2017 ◽

Author(s):

John Lowe

Keyword(s):

Small Molecule ◽

Protein Interactions ◽

Protein Protein Interactions ◽

Small Molecule Drugs

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Exploring Proteomic Drug Targets, Therapeutic Strategies and Protein - Protein Interactions in Cancer: Mechanistic View

Current Cancer Drug Targets ◽

10.2174/1568009618666180803104631 ◽

2019 ◽

Vol 19 (6) ◽

pp. 430-448 ◽

Cited By ~ 1

Author(s):

Khalid Bashir Dar ◽

Aashiq Hussain Bhat ◽

Shajrul Amin ◽

Syed Anjum ◽

Bilal Ahmad Reshi ◽

...

Keyword(s):

Protein Interactions ◽

High Throughput Screening ◽

Critical Role ◽

Cancer Therapeutics ◽

Adaptor Proteins ◽

Therapeutic Strategies ◽

Small Gtpases ◽

Beta Catenin ◽

Protein Protein Interactions ◽

Apoptosis Protein

Protein-Protein Interactions (PPIs) drive major signalling cascades and play critical role in cell proliferation, apoptosis, angiogenesis and trafficking. Deregulated PPIs are implicated in multiple malignancies and represent the critical targets for treating cancer. Herein, we discuss the key protein-protein interacting domains implicated in cancer notably PDZ, SH2, SH3, LIM, PTB, SAM and PH. These domains are present in numerous enzymes/kinases, growth factors, transcription factors, adaptor proteins, receptors and scaffolding proteins and thus represent essential sites for targeting cancer. This review explores the candidature of various proteins involved in cellular trafficking (small GTPases, molecular motors, matrix-degrading enzymes, integrin), transcription (p53, cMyc), signalling (membrane receptor proteins), angiogenesis (VEGFs) and apoptosis (BCL-2family), which could possibly serve as targets for developing effective anti-cancer regimen. Interactions between Ras/Raf; X-linked inhibitor of apoptosis protein (XIAP)/second mitochondria-derived activator of caspases (Smac/DIABLO); Frizzled (FRZ)/Dishevelled (DVL) protein; beta-catenin/T Cell Factor (TCF) have also been studied as prospective anticancer targets. Efficacy of diverse molecules/ drugs targeting such PPIs although evaluated in various animal models/cell lines, there is an essential need for human-based clinical trials. Therapeutic strategies like the use of biologicals, high throughput screening (HTS) and fragment-based technology could play an imperative role in designing cancer therapeutics. Moreover, bioinformatic/computational strategies based on genome sequence, protein sequence/structure and domain data could serve as competent tools for predicting PPIs. Exploring hot spots in proteomic networks represents another approach for developing targetspecific therapeutics. Overall, this review lays emphasis on a productive amalgamation of proteomics, genomics, biochemistry, and molecular dynamics for successful treatment of cancer.

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Small Molecule Inhibitors of Influenza Virus Entry

Pharmaceuticals ◽

10.3390/ph14060587 ◽

2021 ◽

Vol 14 (6) ◽

pp. 587

Author(s):

Zhaoyu Chen ◽

Qinghua Cui ◽

Michael Caffrey ◽

Lijun Rong ◽

Ruikun Du

Keyword(s):

Influenza Virus ◽

Membrane Fusion ◽

Small Molecule ◽

Drug Target ◽

Small Molecule Inhibitors ◽

Virus Entry ◽

Critical Role ◽

Structural Basis ◽

Future Directions ◽

The Past

Hemagglutinin (HA) plays a critical role during influenza virus receptor binding and subsequent membrane fusion process, thus HA has become a promising drug target. For the past several decades, we and other researchers have discovered a series of HA inhibitors mainly targeting its fusion machinery. In this review, we summarize the advances in HA-targeted development of small molecule inhibitors. Moreover, we discuss the structural basis and mode of action of these inhibitors, and speculate upon future directions toward more potent inhibitors of membrane fusion and potential anti-influenza drugs.

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Coiled-coil networking shapes cell molecular machinery

Molecular Biology of the Cell ◽

10.1091/mbc.e12-05-0396 ◽

2012 ◽

Vol 23 (19) ◽

pp. 3911-3922 ◽

Cited By ~ 44

Author(s):

Yongqiang Wang ◽

Xinlei Zhang ◽

Hong Zhang ◽

Yi Lu ◽

Haolong Huang ◽

...

Keyword(s):

Protein Interactions ◽

Critical Role ◽

Coiled Coil ◽

Coiled Coils ◽

Protein Protein Interactions ◽

Full Spectrum ◽

Kinetochore Assembly ◽

Genome Wide ◽

Molecular Machinery ◽

Systematic Understanding

The highly abundant α-helical coiled-coil motif not only mediates crucial protein–protein interactions in the cell but is also an attractive scaffold in synthetic biology and material science and a potential target for disease intervention. Therefore a systematic understanding of the coiled-coil interactions (CCIs) at the organismal level would help unravel the full spectrum of the biological function of this interaction motif and facilitate its application in therapeutics. We report the first identified genome-wide CCI network in Saccharomyces cerevisiae, which consists of 3495 pair-wise interactions among 598 predicted coiled-coil regions. Computational analysis revealed that the CCI network is specifically and functionally organized and extensively involved in the organization of cell machinery. We further show that CCIs play a critical role in the assembly of the kinetochore, and disruption of the CCI network leads to defects in kinetochore assembly and cell division. The CCI network identified in this study is a valuable resource for systematic characterization of coiled coils in the shaping and regulation of a host of cellular machineries and provides a basis for the utilization of coiled coils as domain-based probes for network perturbation and pharmacological applications.

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