Structural Determinants of the Interactions Between Influenza A Virus Matrix Protein M1 and Lipid Membranes

Abstract

13 Influenza A virus is a pathogen responsible for severe seasonal epidemics threatening human 14 and animal populations every year. One of the ten proteins encoded by the viral genome, the 15 matrix protein M1, is abundantly produced in infected cells and plays a structural role in 16 determining the morphology of the virus. During assembly of new viral particles, M1 is 17recruited to the host cell membrane where it associates with lipids and other viral proteins. 18The structure of M1 is only partially known. In particular, structural details of M1 interactions 19with the cellular plasma membrane as well as M1-protein interactions and multimerization 20have not been clarified, yet. 21In this work, we employed a set of complementary experimental and theoretical tools to 22 tackle these issues. Using raster image correlation, surface plasmon resonance and circular 23 dichroism spectroscopies, we quantified membrane association and oligomerization of full-24 length M1 and of different genetically engineered M1 constructs (i.e., N-and C-terminally 25 truncated constructs and a mutant of the polybasic region, residues 95-105). Furthermore, we 26 report novel information on structural changes in M1 occurring upon binding to membranes. 27 Our experimental results are corroborated by an all-atom model of the full-length M1 protein 28 bound to a negatively charged lipid bilayer. 29 30 31 Influenza A viruses (IAV) circulate in human and animal populations, posing a significant 32 threat to global health. They are responsible for severe pathologies with high mortality during 33 seasonal epidemics and occasional pandemic outbreaks [1]. IAVs belong to the family of 34 Orthomyxoviridae and are negative-sense single-stranded RNA viruses [2]. The segmented 35 viral genome, in form of eight viral ribonucleoprotein complexes (vRNPs), is enclosed by the 36 viral lipid envelope [3]. The matrix layer underneath the viral membrane is formed by the 37 matrix protein M1, which consists of 252 amino acid (aa) residues [4-8]. M1 is the most 38 abundant protein in virus particles, stabilizes the virion structure and is considered to be a 39 determinant of viral morphology [9-13]. Electron micrographs demonstrate tight association 40 of the matrix layer with the viral lipid membrane [5]. M1 is abundantly produced in infected 41 cells and distributes throughout cytoplasm and nuclear compartment. In addition to its role as 42 a structural protein, M1 has other functions during the viral replication cycle. It is involved in 43 nucleocytoplasmic transport of the viral genome and, therefore, plays a role in coordinating 44 nuclear-localized replication and assembly of progeny viruses at the plasma membrane (PM) 45 [14-19]. M1 is furthermore considered to be a key player during virus assembly because of its 46 ability to associate with all other structural components of virus particles, including the 47 genomic vRNP complexes [20-22], the three viral transmembrane proteins (i.e. hemagglutinin 48 (HA), neuraminidase (NA) and the proton chan...

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

“…We find that arginines at positions 76 and 77 are essential, while a relatively conservative lysine substitution is permissible in place of R78. We further note that this triplet is buttressed by highly preferred hydrophobic, likely membrane-interacting residues L74, F79, and V80 (67).…”