Positively selected genes in the hoary bat (Lasiurus cinereus) lineage: prominence of thymus expression, immune and metabolic function, and regions of ancient synteny

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Zoological Science

Main article text

 

Introduction

Materials and Methods

DNA extraction and sequencing

Gene annotation

Evolutionary rate analysis

Results and discussion

Genes under positive selection

Genomic clusters of significant genes

Selected genes with immune function and identification of a Steep1 paralog

Energetics

Sensory and behavioral gene functions

Genes associated with craniodysmorphy

Expansion of a Tbx1-like gene in L. cinereus

Limitations and caveats of this study

Conclusions

Future directions

Supplemental Information

Alignment and selection test statistics for each orthogroup.

Round 1 values are for all tested orthogroup alignments whereas Round 2 values are after manual review of initially significant tests.

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Maker-derived gene models in gff format.

The gff format specification is detailed at https://en.wikipedia.org/wiki/General_feature_format and a number of software packages are available to extract or manipulate gff-formatted features.

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Orthogroup alignments in FASTA format.

Initially tested orthogroups that have been edge-trimmed as described in the Methods.

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Manually revised orthogroup alignments.

Likely alignment errors have been manually deleted between conserved codons. Recovery of the original codon positions requires alignment to the original accessions in each orthogroup.

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Qualitative comparison of PAML and aBSREL P-values for branch-specific diversifying selection in the hoary bat (Lasiurus cinereus) lineage.

(A) PAML and aBSREL P-values sorted by decreasing PAML value. Values are formatted as -log10(P). (B) PAML and aBSREL P-values sorted by decreasing aBSREL P-value. Values are formatted as -log10(P). (C) Scatterplots of PAML and aBSREL P-values by descending quintile of alignment length (longest alignments in the first panel). Arrows illustrate a qualitative dual-distribution pattern.

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Scatterplot illustrating the inverse relation between ω2and p2estimates for significant genes in the hoary bat (Lasiurus cinereus) lineage.

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Contingency table and statistics for frequency of positively selected single-exon genes in the hoary bat (Lasiurus cinereus) lineage.

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Gene expression values for mouse orthologs of tested orthogroups.

Mouse orthologs were extracted from the National Center for Biotechnology Information (NCBI) Gene database of the Myotis lucifugus gene in each orthogroup. Values are normalized to a maximum of 1.

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Top three physical clusters of postively selected Lasiurus cinereus genes and their inferred orthologs from model organisms.

Orthologies are pre-computed by Ensembl identifier and were extracted from each Myotis lucifiugus page when available, otherwise from the corresponding mouse gene page. Genes are listed in the form “Gene name:chromosme or scaffold:start coordinate:end coordinate:strand”. Colored blocks within species are colinear with the L. cinereus orthologs.

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Visualization of conserved synteny among mammals of positively selected genes clustered on Lasiurus cinereus Scaffold 133.

In each pairwise alignment, the same region of human chromosome 16 is compared with mouse, cow, M. lucifugus, L. cinereus, and zebrafish. Green bars indicate the positions of the human orthologs of the five positively selected L. cinereus genes (labeled). Dots represent pairwise protein similarity above threshold values in the same (red) or reverse (blue) orientation. Contiguous strings of protein similarity encompassing multiple genes are interpreted as evolutionary conservation of synteny.

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Table of features on Scaffold 533 of the Lasiurus cinereus assembly, in the vicinity of the region that is syntenic with the cat-eye critical region (CECR).

Features are ordered by coordinates and the orthogroup in which the feature was placed is given, along with the total number of orthogroup elements for each bat species. Green shaded cells highlight perfect or near-perfect orthogroups, whereas yellow shaded cells highlight orthogroups with a high copy number in L. cinereus. Features in bold font are landmarks of the human CECR or adjacent DiGeorge critical region (DGCR).

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Visualization of conserved synteny among tetrapods of positively selected genes clustered on Lasiurus cinereus Scaffold 111.

In each pairwise alignment, the same region of human chromosome 16 is compared with frog, chicken, cow, mouse, Myotis lucifugus, and L. cinereus. Green bars indicate the positions of the human orthologs of the four positively selected L. cinereus genes (labeled). Dots represent pairwise protein similarity above threshold values in the same (red) or reverse (blue) orientation. Contiguous strings of protein similarity encompassing multiple genes are interpreted as evolutionary conservation of synteny.

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Annotation summaries of positively selected genes, including orthologs, orthogroup properties, expression data, annotation data, and functional classifications.

Gene summaries, GeneRIFs, GO terms, and other controlled-vocabulary descriptions of gene function were extracted from the NCBI Gene database. Other annotations drawn from the literature are cited.

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Location, structure, and expression of genes inferred to be orthologous to the Steep1-like paralog of Lasiurus cinereus..

Each image is a genome browser view of the corresponding Gene database entry. The source scaffold is displayed in the upper left corner of each window, with coordinate tracks at the top and bottom of each window. The exon structure of each gene is shown in the first track, with subsequent tracks showing relative RNAseq-based expression and support for individual splicing events. Complete descriptions of track icons and data sources are available from the NCBI Gene database (https://www.ncbi.nlm.nih.gov/gene).

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A human Steep1 ‘pseudogene’ is well conserved and expressed in New World and Old World primates.

(A) Location, structure, and expression of the human locus and its orthologs in New and Old World primates. Each image is a genome browser view of the corresponding Gene database entry. The source scaffold is displayed in the upper left corner of each window, with a coordinates track at the base of each window. The exon structure of each gene is shown in the first track, with subsequent tracks showing relative RNAseq-based expression and support for individual splicing events. Complete descriptions of track icons and data sources are available from the NCBI Gene database (https://www.ncbi.nlm.nih.gov/gene). (B) Dendrogram of the Steep1 paralog in representative primates, illustrating a high-level of protein-sequence conservation.

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Expansion of a novel T-box transcription factor (Tbx)-like gene family in Lasiurus cinereus.

(A) Alignment of orthogroup OG0000803 sequences with Tbx family homologs of the Vespertilionid species L. cinereus, Eptesicus fuscus, and Myotis lucifugus. The alignment region containing the conserved Tbx domain is indicated by the blue box above the alignment, whereas the portion of the domain present in the novel orthogroup sequences is outlined in yellow. (B) Neighbor-joining dendrogram of the alignment in panel A. The two halves of the dendrogram, split here for clarity, are joined on the dotted lines. The left half contains the single Tbx-like gene present in E. fuscus (XP_027987819) and annotated as Tbx1-like, along with the fifteen copies annotated in L. cinereus. Not all annotations are complete and whether all are expressed is unknown. The right half of the dendrogram contains all identified Tbx homologs with complete domains identified in these three species. Note that an unannotated Tbx1 ortholog is likely present in L. cinereus, as a TBLASTN search with the E. fuscus ortholog (XP_028003188) has a strong match in the syntenic genomic location on scaffold 586. The Tbx family appears to be approximately complete in L. cinereus and shows an approximately 1-to-1 relationship with M. lucifugus and E. fuscus genes. Therefore, the Tbx1-like sequences are not simply incomplete annotation artifacts. The phylogeny used the JTT amino-acid distance measure with pairwise deletion of gaps. Rate variation among sites was modeled as a five-category gamma distribution with a shape parameter of one.

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Additional Information and Declarations

Competing Interests

The authors declare that they have no competing interests.

Author Contributions

Robert S. Cornman conceived and designed the experiments, performed the experiments, analyzed the data, prepared figures and/or tables, authored or reviewed drafts of the paper, and approved the final draft.

Paul M. Cryan conceived and designed the experiments, authored or reviewed drafts of the paper, and approved the final draft.

Data Availability

The following information was supplied regarding data availability:

The hoary bat genome sequence that was annotated is available from the ScienceBase repository: Cornman, R.S., Fike, J.M., Oyler-McCance, S.J., and Cryan, P.M., 2021, Genetic variation in hoary bats (Lasiurus cinereus) assessed from archived samples: U.S. Geological Survey data release, https://doi.org/10.5066/P9VSG54Z.

The gene annotations in gff format and orthogroup sequence alignments in FASTA format are available in the Supplemental Files.

Funding

The authors received no external funding for this work.

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