Broad Spectrum Anti-Influenza Agents by Inhibiting Self-Association of Matrix Protein 1

Mosier, Philip D.; Chiang, Meng-Jung; Lin, Zhengshi; Gao, Yamei; Althufairi, Bashayer; Zhou, Qibing; Musayev, Faik N.; Safo, Martin K.; Xie, Hang; Desai, Umesh R.

doi:10.1038/srep32340

Cited by 10 publications

(9 citation statements)

References 57 publications

(82 reference statements)

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“… 30 , 31 Impairing M1 oligomerization through the use of a small molecule ‘wedge’ has also proven to be a viable approach to develop broad-spectrum anti-IAV agents. 42 Besides IAV, many enveloped viruses such as Flaviviridae (Dengue and Zika viruses) and Filoviridae (Ebola and Marburg viruses) also have similar M1 structure in their viral particles. Thus, our current study offers new insights into developing M1-based countermeasures beyond IAV.…”

Section: Discussionmentioning

confidence: 99%

Maintaining pH-dependent conformational flexibility of M1 is critical for efficient influenza A virus replication

Chiang

Musayev

Kosikova

et al. 2017

Emerging Microbes & Infections

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The M gene segment of influenza A virus has been shown to be a contributing factor to the high growth phenotype. However, it remains largely unknown why matrix protein 1 (M1), the major structural protein encoded by M gene, exhibits pH-dependent conformational changes during virus replication. Understanding the mechanisms underlying efficient virus replication can help to develop strategies not only to combat influenza infections but also to improve vaccine supplies. M(NLS-88R) and M(NLS-88E) are two M1 mutants differing by only a single amino acid: G88R vs G88E. G88R but not G88E was the compensatory mutation naturally selected by the virus after its nuclear localization signal was disrupted. Our study shows that, compared with M(NLS-88E) M1, M(NLS-88R) M1 dissociated quickly from viral ribonucleoproteins (vRNPs) at higher pH and took less time to dissemble in vitro, despite forming thicker matrix layer and having stronger association with vRNP in assembled virions. Correspondingly, M(NLS-88R) replicated more efficiently and was genetically more stable than M(NLS-88E). Crystallographic analysis indicated that M(NLS-88R) M1, like wild-type M1, is able to switch from a face-to-back-oriented conformation to a face-to-face-oriented conformation when pH drops from neutral to acidic, whereas G88E mutation causes M(NLS-88E) M1 to be trapped in a face-to-face-arranged conformation regardless of environmental pH. Our results suggest that maintaining M1 pH-dependent conformational flexibility is critical for efficient virus replication, and position 88 is a key residue controlling M1 pH-dependent conformational changes. Our findings provide insights into developing M1-based antiviral agents.

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Section: Discussionmentioning

confidence: 99%

Maintaining pH-dependent conformational flexibility of M1 is critical for efficient influenza A virus replication

Chiang

Musayev

Kosikova

et al. 2017

Emerging Microbes & Infections

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“…Such multifaceted functional roles impose significant constraints on M1’s ability to evolve. In addition to its role in recruiting multiple viral components at assembly budding sites, M1 plays pivotal roles in determining particle morphology, promoting an orderly sequence to membrane fusion, and trafficking vRNP following cell entry (24–32). Additionally, M1 interacts with a variety of host proteins, which phosphorylate and SUMOylate several sites in M1.…”

Section: Introductionmentioning

confidence: 99%

Deep Mutational Scan of the Highly Conserved Influenza A Virus M1 Matrix Protein Reveals Substantial Intrinsic Mutational Tolerance

Hom

Gentles

Bloom

et al. 2019

J Virol

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Influenza A virus matrix protein M1 is involved in multiple stages of the viral infectious cycle. Despite its functional importance, our present understanding of this essential viral protein is limited. The roles of a small subset of specific amino acids have been reported, but a more comprehensive understanding of the relationship between M1 sequence, structure, and virus fitness remains elusive. In this study, we used deep mutational scanning to measure the effect of every amino acid substitution in M1 on viral replication in cell culture. The map of amino acid mutational tolerance we have generated allows us to identify sites that are functionally constrained in cell culture as well as sites that are less constrained. Several sites that exhibit low tolerance to mutation have been found to be critical for M1 function and production of viable virions. Surprisingly, significant portions of the M1 sequence, especially in the C-terminal domain, whose structure is undetermined, were found to be highly tolerant of amino acid variation, despite having extremely low levels of sequence diversity among natural influenza virus strains. This unexpected discrepancy indicates that not all sites in M1 that exhibit high sequence conservation in nature are under strong constraint during selection for viral replication in cell culture. IMPORTANCE The M1 matrix protein is critical for many stages of the influenza virus infection cycle. Currently, we have an incomplete understanding of this highly conserved protein’s function and structure. Key regions of M1, particularly in the C terminus of the protein, remain poorly characterized. In this study, we used deep mutational scanning to determine the extent of M1’s tolerance to mutation. Surprisingly, nearly two-thirds of the M1 sequence exhibits a high tolerance for substitutions, contrary to the extremely low sequence diversity observed across naturally occurring M1 isolates. Sites with low mutational tolerance were also identified, suggesting that they likely play critical functional roles and are under selective pressure. These results reveal the intrinsic mutational tolerance throughout M1 and shape future inquiries probing the functions of this essential influenza A virus protein.

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“…Conservation of M1 suppressor mutation sequences and M1-M2 cooperativity in virus budding. There is a high degree of conservation in natural isolates of influenza virus from animals and humans at the amino acids corresponding to the M1 suppressor mutations (43). This suggests a highly conserved function for these residues in the influenza virus life cycle.…”

Section: Discussionmentioning

confidence: 98%

Mutations in the Influenza A Virus M1 Protein Enhance Virus Budding To Complement Lethal Mutations in the M2 Cytoplasmic Tail

Liu

Grantham

Pekosz

2018

J Virol

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The influenza A virus M1 and M2 proteins play important roles in virus assembly and in the morphology of virus particles. Mutations in the distal cytoplasmic tail region of M2, and in particular a tyrosine-to-alanine mutation at residue 76 (Y76A), were essential for infectious virus production and filament formation while having limited effects on total virus particle budding. Using a novel selection method, mutations at seven different M1 amino acids (residue 73, 94, 135, 136, or 138 or a double mutation, 93/244) that are not found in circulating influenza virus strains or have not been previously identified to play a role in influenza A virus assembly were found to complement the lethal M2Y76A mutation. These M1 suppressor mutations restored infectious virus production in the presence of M2Y76A and mediated increased budding and filament formation even in the absence of M2. However, the efficiency of infectious virus replication was still dependent on the presence of the distal region of the M2 cytoplasmic tail. The data suggest that influenza A virus budding and genome incorporation can occur independently and provide further support for complementary roles of the M1 and M2 proteins in virus assembly. Influenza virus particle assembly involves the careful coordination of various viral and host factors to optimally produce infectious virus particles. We have previously identified a mutation at position 76 of the influenza A virus M2 protein that drastically reduces infectious virus production and filament formation with minimal effects on virus budding. In this work, we identified suppressor mutations in the M1 protein which complement this lethal M2 mutation by increasing the efficiency with which virus particles bud from infected cells and restoring filament formation at the infected-cell surface. M2 distal cytoplasmic domain sequences were still required for optimal infectivity. This indicates that M1 and M2 can functionally replace each other in some, but not all, aspects of virus particle assembly.

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Broad Spectrum Anti-Influenza Agents by Inhibiting Self-Association of Matrix Protein 1

Cited by 10 publications

References 57 publications

Maintaining pH-dependent conformational flexibility of M1 is critical for efficient influenza A virus replication

Maintaining pH-dependent conformational flexibility of M1 is critical for efficient influenza A virus replication

Deep Mutational Scan of the Highly Conserved Influenza A Virus M1 Matrix Protein Reveals Substantial Intrinsic Mutational Tolerance

Mutations in the Influenza A Virus M1 Protein Enhance Virus Budding To Complement Lethal Mutations in the M2 Cytoplasmic Tail

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