Structural and Functional Characterization of Host FHL1 Protein Interaction with Hypervariable Domain of Chikungunya Virus nsP3 Protein

• Published in: Journal of virology Vol. 95; no. 1
• Main Authors: Lukash, Tetyana, Agback, Tatiana, Dominguez, Francisco, Shiliaev, Nikita, Meshram, Chetan, Frolova, Elena I, Agback, Peter, Frolov, Ilya
• Format: Journal Article
• Language: English
• Published: United States American Society for Microbiology 09-12-2020
• Subjects: Adaptor Proteins, Signal Transducing - chemistry; Adaptor Proteins, Signal Transducing - metabolism; Allosteric Site; Animals; Binding Sites; Cell Line; Chikungunya virus - metabolism; Cytoskeletal Proteins - chemistry; Cytoskeletal Proteins - metabolism; Host-Pathogen Interactions; Humans; Intracellular Signaling Peptides and Proteins - chemistry; Intracellular Signaling Peptides and Proteins - genetics; Intracellular Signaling Peptides and Proteins - metabolism; LIM Domain Proteins - chemistry; LIM Domain Proteins - genetics; LIM Domain Proteins - metabolism; LIM-Homeodomain Proteins - chemistry; LIM-Homeodomain Proteins - genetics; LIM-Homeodomain Proteins - metabolism; Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy); Medicinsk bioteknologi (med inriktning mot cellbiologi (inklusive stamcellsbiologi), molekylärbiologi, mikrobiologi, biokemi eller biofarmaci); Muscle Proteins - chemistry; Muscle Proteins - genetics; Muscle Proteins - metabolism; Mutation; Protein Binding; Protein Domains; Transcription Factors - chemistry; Transcription Factors - genetics; Transcription Factors - metabolism; Viral Nonstructural Proteins - chemistry; Viral Nonstructural Proteins - genetics; Viral Nonstructural Proteins - metabolism; Virus Replication; Virus-Cell Interactions; viral replication; FHL1; FHL2; alphaviruses; FHL3; chikungunya virus; nsP3; intrinsically disordered proteins; nuclear magnetic resonance; CD2AP
• ISSN: 0022-538X; 1098-5514; 1098-5514
• DOI: 10.1128/JVI.01672-20

Decades of insufficient control have resulted in unprecedented spread of chikungunya virus (CHIKV) around the globe, and millions have suffered from the highly debilitating disease. Nevertheless, the current understanding of CHIKV-host interactions and adaptability of the virus to replication in mosquitoes and mammalian hosts is still elusive. Our new study shows that four-and-a-half LIM domain protein (FHL1) is one of the host factors that interact with the hypervariable domain (HVD) of CHIKV nsP3. Unlike G3BPs, FHL1 is not a prerequisite of CHIKV replication, and many commonly used cell lines do not express FHL1. However, its expression has a detectable stimulatory effect(s) on CHIKV replication, and
knockout (KO) cell lines demonstrate slower infection spread. Nuclear magnetic resonance (NMR)-based studies revealed that the binding site of FHL1 in CHIKV nsP3 HVD overlaps that of another proviral host factor, CD2AP. The structural data also demonstrated that FHL1-HVD interaction is mostly determined by the LIM1 domain of FHL1. However, it does not mirror binding of the entire protein, suggesting that other LIM domains are involved. In agreement with previously published data, our biological experiments showed that interactions of CHIKV HVD with CD2AP and FHL1 have additive effects on the efficiency of CHIKV replication. This study shows that CHIKV mutants with extensive modifications of FHL1- or both FHL1- and CD2AP-binding sites remain viable and develop spreading infection in multiple cell types. Our study also demonstrated that other members of the FHL family can bind to CHIKV HVD and thus may be involved in viral replication.
Replication of chikungunya virus (CHIKV) is determined by a wide range of host factors. Previously, we have demonstrated that the hypervariable domain (HVD) of CHIKV nsP3 contains linear motifs that recruit defined families of host proteins into formation of functional viral replication complexes. Now, using NMR-based structural and biological approaches, we have characterized the binding site of the cellular FHL1 protein in CHIKV HVD and defined the biological significance of this interaction. In contrast to previously described binding of G3BP to CHIKV HVD, the FHL1-HVD interaction was found to not be a prerequisite of viral replication. However, the presence of FHL1 has a stimulatory effect on CHIKV infectivity and, subsequently, the infection spread. FHL1 and CD2AP proteins were found to have overlapping binding sites in CHIKV HVD and additive proviral functions. Elimination of the FHL1-binding site in the nsP3 HVD can be used for the development of stable, attenuated vaccine candidates.