DOI: 10.5061/dryad.3bk3j9ktx
title: Data from: Ancient climate changes and relaxed selection shape cave colonization in North American cavefishes

description: Extreme environments serve as natural laboratories for studying
 evolutionary processes, with caves offering replicated instances of
 independent colonisations. The timing, mode, and genetic underpinnings
 underlying cave-obligate organismal evolution remain enigmatic. We
 integrate phylogenomics, fossils, paleoclimatic modeling, and newly
 sequenced genomes to elucidate the evolutionary history and adaptive
 processes of cave colonisation in the study group, the North American
 Amblyopsidae fishes. Amblyopsid fishes present a unique system for
 investigating cave evolution, encompassing surface, facultative
 cave-dwelling, and cave-obligate (troglomorphic) species. Using 1,105 exon
 markers and total-evidence dating, we reconstructed a robust phylogeny
 that supports the nested position of eyed, facultative cave-dwelling
 species within blind cavefishes. We identified three independent cave
 colonisations, dated to the Early Miocene (18.5 Mya), Late Miocene (10.0
 Mya), and Pliocene (3.0 Mya). Evolutionary model testing supported a
 climate-relict hypothesis, suggesting that global cooling trends since the
 Early–Middle Eocene may have influenced cave colonisation. Comparative
 genomic analyses of 487 candidate genes revealed both relaxed and
 intensified selection on troglomorphy-related loci. We found more loci
 under relaxed selection, supporting neutral mutation as a significant
 mechanism in cave-obligate evolution. Our findings provide empirical
 support for climate-driven cave colonisation and offer insights into the
 complex interplay of selective pressures in extreme environments.
description: We performed short-read shotgun genome sequencing on
 Percopsiformes fishes, then harvested exons using Hughes et al. (2018)
 FishLifeExonHarvesting pipeline. Exon markers were used to create a
 phylogeny, then extinct fossil taxa were added as tips in the phylogeny to
 time-calibrate the tree. 
description: # Ancient climate changes drive mode and tempo of cave colonization in
 North American cavefishes
 [https://doi.org/10.5061/dryad.3bk3j9ktx](https://doi.org/10.5061/dryad.3bk3j9ktx) ## Description of the data and file structure The dataset contains the aligned exons (fasta format) harvested from whole genome resequencing data for the dataset in Hart et al. (in review). Also included is the coding scheme for the morphological matrix used to incorporate fossil Percopsiformes taxa into the total-evidence divergence analyses. ### Folders, Files, and variables #### File: Percopsiformes\_exons\_NT\_Concatenated\_alignments.fasta **Description:** All exons harvested from whole genome resequencing data for all specimens aligned. #### File: Percopsiformes\_fossil\_coding.csv **Description:** Coding of fossil taxa into the morphological matrix from Armbruster et al. (2016). ##### Variables * Fossil_Taxa * Order * Family * Morphological characters from Armbruster et al. (2016) #### File: astral\_percopsiformes.tre **Description:** File containing the Percopsiformes phylogeny estimated using ASTRAL. #### File: iqtree\_percopsiformes.tre **Description:** File containing the Percopsiformes phylogeny estimated using IQTree. #### File: mrbayes\_tipdating\_percopsiformes.tre **Description:** File containing the tip-dated Percopsiformes phylogeny estimated using Mr. Bayes. #### Folder: gene\_alignments.zip **Description:** Folder containing all gene alignments used in this study in fasta format. #### File: nhmmer2fasta.py **Description:** Python script used to concatenated and convert nhmmer output to fasta. #### File: CDHit\_Exonerate.sh **Description:** Bash script used to run CDHIT and exonerate on the prealignments. #### File: run\_macse2.edit.sh **Description:** Bash script used to align our alignments using MACSE. #### File: Relax\_Intensify\_Selection.sh **Description:** Bash script used to test for relaxation and intensifying selection. #### File: SuppMat1\_Percopsiformes\_molecular-specimens.csv Description: Tissue specimens used for whole shotgun genome resequencing and their associated BioProject numbers. #### File: SuppMat2\_Percopsiformes.txt Description: Fossil calibration age priors & integration of fossil and extant species #### File: SuppMat3\_Percopsiformes\_genome-stats.xlsx **Description:** Genome statistics for the Percopsiformes used in this study. #### File: SuppMat4\_Percopsiformes\_SelectionAnalyses\_Results.xlsx **Description:** Results for the positive selection analyses performed for this study. #### File: SuppMat5\_Percopsiformes\_GUIS.xlsx **Description:** Genes found to be under intensified selection. #### File: SuppMat6\_Percopsiformes\_GURS.xlsx **Description:** Genes found to be under relaxed selection. #### File: SuppMat7\_Percopsiformes\_GURS\_Sources.txt **Description:** Sources used to determine the function for genes under relaxed selection. #### Folder: Pam\_Climatic.zip **Description:** Folder containing all data needed to replicate the paleoclimatic analyses used for this study. Folder contents include the R hisotry file (.Rhistory), a text file containing the trait data (Data_w_Out.txt), an Excel file containing the cave state for our taxa (pformes_cave-state_table) needed to run our analysis, tree file containing the phylogeny (Percopsiformes_TipDating_MrBayes.tre), the different scales used for the analyses (Scaled_Scotese_Deep.csv, Scaled_Scotese_GAT.csv, Scaled_Scotese_Polar.csv, Scaled_Scotese_Tropical.csv), and the results (Results.Pam). An additional subfolder contains the the facultative alternate scheme (Faculative_Alternate). This folder contains the the two functions used in all of our analyses (BM_OU_Lambda_ML_gauss.r and Climate_ML_LL_Gauss_ME.r), the submission script (Clim_Pam.sh), a text file containing the trait data (Faculative_Data_w_Out), a newick file containing the phylogeny (Newick_Pam_tree.nwk), R Script used for the analyses (Pam_Climatic.R), raw error and output files, result files (Results_Pam.csv), the different scales used for the analyses (Scaled_Scotese_GAT.csv, Scaled_Scotese_Tropical.csv), and the screen log outputs from R classified as either output (r_output_21248717) or error (r_error_21248717) to provide an all-encompassing and transparent view of the analysis.