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Structures of honeybee-infecting Lake Sinai virus reveal domain functions and capsid assembly with dynamic motions.

ABSTRACT: Understanding the structural diversity of honeybee-infecting viruses is critical to maintain pollinator health and manage the spread of diseases in ecology and agriculture. We determine cryo-EM structures of T = 4 and T = 3 capsids of virus-like particles (VLPs) of Lake Sinai virus (LSV) 2 and delta-N48 LSV1, belonging to tetraviruses, at resolutions of 2.3-2.6 Å in various pH environments. Structural analysis shows that the LSV2 capsid protein (CP) structural features, particularly the protruding domain and C-arm, differ from those of other tetraviruses. The anchor loop on the central β-barrel domain interacts with the neighboring subunit to stabilize homo-trimeric capsomeres during assembly. Delta-N48 LSV1 CP interacts with ssRNA via the rigid helix α1', α1'-α1 loop, β-barrel domain, and C-arm. Cryo-EM reconstructions, combined with X-ray crystallographic and small-angle scattering analyses, indicate that pH affects capsid conformations by regulating reversible dynamic particle motions and sizes of LSV2 VLPs. C-arms exist in all LSV2 and delta-N48 LSV1 VLPs across varied pH conditions, indicating that autoproteolysis cleavage is not required for LSV maturation. The observed linear domino-scaffold structures of various lengths, made up of trapezoid-shape capsomeres, provide a basis for icosahedral T = 4 and T = 3 architecture assemblies. These findings advance understanding of honeybee-infecting viruses that can cause Colony Collapse Disorder.

SUBMITTER: Chen NC

PROVIDER: S-EPMC9892032 | biostudies-literature | 2023 Feb

REPOSITORIES: biostudies-literature

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Structures of honeybee-infecting Lake Sinai virus reveal domain functions and capsid assembly with dynamic motions.

Chen Nai-Chi NC Wang Chun-Hsiung CH Yoshimura Masato M Yeh Yi-Qi YQ Guan Hong-Hsiang HH Chuankhayan Phimonphan P Lin Chien-Chih CC Lin Pei-Ju PJ Huang Yen-Chieh YC Wakatsuki Soichi S Ho Meng-Chiao MC Chen Chun-Jung CJ

Nature communications 20230201 1

Understanding the structural diversity of honeybee-infecting viruses is critical to maintain pollinator health and manage the spread of diseases in ecology and agriculture. We determine cryo-EM structures of T = 4 and T = 3 capsids of virus-like particles (VLPs) of Lake Sinai virus (LSV) 2 and delta-N48 LSV1, belonging to tetraviruses, at resolutions of 2.3-2.6 Å in various pH environments. Structural analysis shows that the LSV2 capsid protein (CP) structural features, particularly the protrudi ...[more]

PMID: 36726015

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Project description:Lake Sinai Viruses (Sinaivirus) are commonly detected in honey bees (Apis mellifera) but no disease phenotypes or fitness consequences have yet been demonstrated. This viral group is genetically diverse, lacks obvious geographic structure, and multiple lineages can co-infect individual bees. While phylogenetic analyses have been performed, the molecular evolution of LSV has not been studied extensively. Here, I use LSV isolates from GenBank as well as contigs assembled from honey bee Sequence Read Archive (SRA) accessions to better understand the evolutionary history of these viruses. For each ORF, substitution rate variation, codon usage, and tests of positive selection were evaluated. Outlier regions of high or low diversity were sought with sliding window analysis and the role of recombination in creating LSV diversity was explored. Phylogenetic analysis consistently identified two large clusters of sequences that correspond to the current LSV1 and LSV2 nomenclature, however lineages sister to LSV1 were the most frequently detected in honey bee SRA accessions. Different expression levels among ORFs suggested the occurrence of subgenomic transcripts. ORF1 and RNA-dependent RNA polymerase had higher evolutionary rates than the capsid and ORF4. A hypervariable region of the ORF1 protein-coding sequence was identified that had reduced selective constraint, but a site-based model of positive selection was not significantly more likely than a neutral model for any ORF. The only significant recombination signals detected between LSV1 and LSV2 initiated within this hypervariable region, but assumptions of the test (single-frame coding and independence of substitution rate by site) were violated. LSV codon usage differed strikingly from that of honey bees and other common honey-bee viruses, suggesting LSV is not strongly co-evolved with that host. LSV codon usage was significantly correlated with that of Varroa destructor, however, despite the relatively weak codon bias exhibited by the latter. While codon usage between the LSV1 and LSV2 clusters was similar for three ORFs, ORF4 codon usage was uncorrelated between these clades, implying rapid divergence of codon use for this ORF only. Phylogenetic placement and relative abundance of LSV isolates reconstructed from SRA accessions suggest that detection biases may be over-representing LSV1 and LSV2 in public databases relative to their sister lineages.

Project description:BackgroundLake Sinai Viruses (LSV) are common RNA viruses of honey bees (Apis mellifera) that frequently reach high abundance but are not linked to overt disease. LSVs are genetically heterogeneous and collectively widespread, but despite frequent detection in surveys, the ecological and geographic factors structuring their distribution in A. mellifera are not understood. Even less is known about their distribution in other species. Better understanding of LSV prevalence and ecology have been hampered by high sequence diversity within the LSV clade.MethodsHere we report a new polymerase chain reaction (PCR) assay that is compatible with currently known lineages with minimal primer degeneracy, producing an expected 365 bp amplicon suitable for end-point PCR and metagenetic sequencing. Using the Illumina MiSeq platform, we performed pilot metagenetic assessments of three sample sets, each representing a distinct variable that might structure LSV diversity (geography, tissue, and species).ResultsThe first sample set in our pilot assessment compared cDNA pools from managed A. mellifera hives in California (n = 8) and Maryland (n = 6) that had previously been evaluated for LSV2, confirming that the primers co-amplify divergent lineages in real-world samples. The second sample set included cDNA pools derived from different tissues (thorax vs. abdomen, n = 24 paired samples), collected from managed A. mellifera hives in North Dakota. End-point detection of LSV frequently differed between the two tissue types; LSV metagenetic composition was similar in one pair of sequenced samples but divergent in a second pair. Overall, LSV1 and intermediate lineages were common in these samples whereas variants clustering with LSV2 were rare. The third sample set included cDNA from individual pollinator specimens collected from diverse landscapes in the vicinity of Lincoln, Nebraska. We detected LSV in the bee Halictus ligatus (four of 63 specimens tested, 6.3%) at a similar rate as A. mellifera (nine of 115 specimens, 7.8%), but only one H. ligatus sequencing library yielded sufficient data for compositional analysis. Sequenced samples often contained multiple divergent LSV lineages, including individual specimens. While these studies were exploratory rather than statistically powerful tests of hypotheses, they illustrate the utility of high-throughput sequencing for understanding LSV transmission within and among species.

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Structures of honeybee-infecting Lake Sinai virus reveal domain functions and capsid assembly with dynamic motions.

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Structures of honeybee-infecting Lake Sinai virus reveal domain functions and capsid assembly with dynamic motions.

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