DOI: 10.25338/b8jm11
title: Data from: Cyanobacteriochromes from Gloeobacterales provide new insight into the diversification of cyanobacterial photoreceptors

description: The phytochrome superfamily comprises three groups of photoreceptors
 sharing a conserved GAF (cGMP-specific phosphodiesterases, cyanobacterial
 adenylate cyclases, and formate hydrogen lyase transcription activator
 FhlA) domain that uses a covalently attached linear tetrapyrrole (bilin)
 chromophore to sense light. Knotted red/far-red phytochromes are
 widespread in both bacteria and eukaryotes, but cyanobacteria also contain
 knotless red/far-red phytochromes and cyanobacteriochromes (CBCRs). Unlike
 phytochromes, CBCRs require only the GAF domain for bilin binding,
 chromophore ligation, and full, reversible photoconversion. CBCRs can
 sense a wide range of wavelengths (ca. 330-750 nm) and can regulate
 phototaxis, second messenger metabolism, and optimization of the
 cyanobacterial light-harvesting apparatus. However, the origin and
 diversification of CBCRs are not well understood. In the current work, we
 use the increasing availability of genomes and
 metagenome-assembled-genomes from early-branching cyanobacteria to
 identify the earliest branches in CBCR evolution. Our analyses also show
 that early-branching cyanobacteria contain more recently evolved CBCRs,
 implicating significant diversification of CBCRs very early in
 cyanobacterial evolution. Moreover, we show that early-branching CBCRs
 behave as integrators of light and pH, providing a potential unique
 function for early CBCRs that could have led to their retention and
 subsequent diversification. Our results thus provide new insight into the
 origins of these diverse cyanobacterial photoreceptors.
description: Data are deposited as a gzipped tarball containing three types of
 data, each in its own directory: Phylogenies, Genome Analysis, and
 Spectroscopic data. Deposited data are all flat text files with UNIX
 newlines. 1. Phylogenies are presented for the CBCR
 (cyanobacteriochrome) domain, the histidine kinase bi-domain, for a
 catenation of taxis proteins, and for a catenation of translation proteins
 (ribosomal proteins + elongation factor 4). In each case, the initial
 sequence alignment was generated using MAFFT v7.450 with the following
 command-line settings:  --genafpair --maxiterate 16
 --clustalout –reorder The resulting alignments are
 deposited in CLUSTAL format (indicated by the .aln extension). For
 phylogenetic analysis, each alignment was processed with an in-house
 script to remove positions having ≥5% gaps. The resulting alignments are
 deposited in PHYLIP format (indicated by the .phy extension) and were used
 to infer maximum-likelihood phylogenies in PhyML-3.1 with 100 bootstraps,
 using the following command-line settings: m WAG -d aa
 -s SPR -a e -c 4 -v e -o tlr -b 100 Support was
 evaluated using the transfer bootstrap expectation (TBE) as implemented in
 booster, and the resulting trees are deposited in Newick format with TBE
 as support values. 2. Genome Analysis.
 Cyanobacterial genomes and metagenome-assembled metagenomes were
 evaluated for assembly size, for the number of candidate phytochrome or
 CBCR open reading frames (ORFs) found in the assembly, and for the total
 number of candidate bilin-binding GAF domains in the assembly. Three files
 are deposited from this analysis, each as a tab-delimited text
 file. ORF_assembly_plot.txt is a spreadsheet for
 2-dimensional (x-y) scatter plotting of the number of ORFs vs. the
 assembly size for Prochlorococcaceae, Gloeobacter spp., and other
 cyanobacteria. ORF_density_plot.txt is a spreadsheet
 for 1-dimensional plotting (e.g., box/whisker) of the ORF density for a
 series of assemblies belonging to cyanobacterial lineages: Gloeobacter
 spp., all other members of the Gloeobacterales, Thermostichales,
 Pseudanabaenales, Gloeomargaritales, and higher crown cyanobacteria. ORF
 density was calculated for each assembly as (number of candidate
 ORFs)/(assembly size). tandem_index_plot.txt is a
 spreadsheet for 1-dimensional plotting of the tandem index for the same
 assemblies as the ORF density and is organized into the same lineages. The
 tandem index was calculated for each assebmly as (number of candidate
 bilin-binding GAF domains)/(number of candidate ORFs).
 3. Spectroscopic data. Six types of
 spectroscopic data are presented, each in its own subdirectory. All
 absorption spectra were acquired on Cary 50 or Cary 60 spectrophotometers;
 circular dichroism (CD) spectra were acquired on an Applied Photophysics
 Chirascan. Raw files were processed with an in-house script to convert to
 tab-delimited format and remove user metadata. The resulting files were
 used for analysis and figure preparation. The contents of each
 subdirectory are as follows: Absorption spectra pH
 7.5 This set comprises a series of 19 text files, one
 for each protein characterized, and each filename matches the name of the
 protein in the manuscript. Spectra were acquired in TKKG buffer (25 mM
 TES-KOH pH 7.5, 100 mM KCl, 10% (v/v) glycerol). Spectra are presented for
 the 15Z dark-adapted state and the 15E photoproduct, along with the
 photochemical difference spectrum (calculated as 15Z – 15E).
 Normalized spectra This set has a similar series of files, each containing
 normalized data for samples
 to facilitate comparison. Files are named by the proteins
 that are compared in each
 case, with three exceptions. These three files present normalized  photochemical
 difference spectra for various proteins at pH 6, at pH 9, and at pH 10. These files are named
 by the pH value instead (for
 example, "pH6_difference" has the difference spectra at pH
 6). static pH spectra
 The pH response was examined for a series of proteins, each in
 either the 15Z or 15E state. In this experiment, 100 µl of protein in TKKG
 buffer was diluted into 1 ml of 0.4 M buffer at different pH values (e.g.,
 0.4M MES, pH 6). This subdirectory contains one file for each protein,
 with the filename containing the name of the protein and the configuration
 (for example, "AnPixJg2_15Z.txt" indicates that the protein is
 AnPixJg2 and it is in the 15Z state). photoconversion
 pH spectra Photoconversion was evaluated at different
 pH values for twelve of the proteins examined under "static pH
 spectra" (above). The resulting photochemical difference spectra at
 pH values of interest are deposited here, with each file named by the
 protein (for example, "AnPixJg2.txt" contains photochemical
 difference spectra for AnPixJg2 at a range of pH values).
 pKa plotting This set has a series of files, each of which
 contains titration data (pH
 and absorbance data as x-y pairs) for one protein. Some files contain data
 for multiple wavelengths
 and/or datasets. CD
 spectra This set has a series of files, each containing
 circular dichroism spectra for a single protein in two photostates. Each file
 is named by the protein. All
 spectra are deposited after baseline subtraction.
description: All files are flat text files. Alignments can
 be opened in text editors or word processors.
 Newick-format tree files were examined using FigTree.
 All other data can be examined using spreadsheet/plotting
 software such as xmgrace, Excel, etc. For this work, data were analyzed
 and plotted using Kaleidagraph.
description: ## Title: Underlying data for "Cyanobacteriochromes from
 Gloeobacterales provide new insight into the diversification of
 cyanobacterial photoreceptors" ## Description of the Data and file
 structure Deposited data are in three folders: Phylogenies Genome_Analysis
 Spectroscopic_data Phylogenies: For each phylogeny, three files are
 deposited: the original multiple sequence alignment in CLUSTAL format, the
 input file for PHYML in PHYLIP format after removal of gap-enriched
 columns, and the final tree with transfer bootstrap expectation values in
 Newick format. As an example, files for the CBCR phylogeny are the
 original alignment (CBCR_domain.aln), the PHYLIP format input file
 (CBCR_domain.phy), and the final output tree (CBCR_domain.TBE.txt). The
 different phylogenetic analyses are described as follows: CBCR_domain
 files are for analysis of the CBCR domain. HisKinase files are for
 analysis of histidine kinase bidomains. Taxis_catenation files are for
 catenated PtxBCDE/HmpBCDE and paralogs. Translation_catenation files are
 for catenated EF4 and ribosomal proteins. Genome Analysis: Three files are
 provided. ORF_assembly_plot.txt number of phytochrome/CBCR open reading
 frames vs. assembly size. ORF_density_plot.txt number of open reading
 frames per Mb for different cyanobacterial lineages. tandem_index_plot.txt
 number of bilin-binding GAF domains per open reading frame for different
 cyanobacterial lineages. Spectroscopic data: Data are organized in six
 subdirectories. "Absorption spectra pH 7.5" has one file for
 each protein, containing spectra for that protein in the 15Z and 15E
 states as well as the (15Z - 15E) photochemical difference spectrum.
 "Normalized spectra" has a similar series of files, each
 containing normalized data for samples to facilitate comparison. Files are
 named by the proteins that are compared in each case, with three
 exceptions. These three files present normalized photochemical difference
 spectra for various proteins at pH 6, at pH 9, and at pH 10. These files
 are named by the pH value instead (for example, "pH6_difference"
 has the difference spectra at pH 6). "pKa plotting" has a series
 of files, each of which contains titration data (pH and absorbance data as
 x-y pairs) for one protein. Some files contain data for multiple
 wavelengths and/or datasets. "photoconversion pH spectra" has a
 series of files, each containing difference spectra for one protein at a
 range of pH values. Difference spectra have been normalized by the peak
 15Z absorbance of that sample at that pH. Each file is named by the
 protein. "static pH spectra" has a series of files, each
 containing spectra for one protein in a single photostate at a range of pH
 values. Each file is named by the file and photostate. "CD
 spectra" has a series of files, each containing circular dichroism
 spectra for a single protein in two photostates. Each file is named by the
 protein. All spectra are deposited after baseline subtraction. CHANGES IN
 SECOND RELEASE: All changes in the second release are in the
 “Spectroscopic Data” directory. First, an additional subdirectory
 containing circular dichroism (CD) data has been added. Additionally,
 spectroscopic data have been added to the other subdirectories, as
 follows: subdirectory “Absorption spectra pH 7.5” Data have been added for
 four additional proteins (HGZ86378, WP_083263321, NpR4776g1, and
 NpR4776g1-PADCIP). subdirectory “Normalized spectra” Data have been added
 for six additional case-by-case comparisons, indicated by the filenames
 (see above). subdirectory “photoconversion pH spectra: Data have been
 added for one additional protein (HGZ86378). subdirectory “pKa plotting”
 Data have been added for eight additional analyses. Each filename
 indicates the photostate and name of the protein in that file (e.g.,
 15Z_HGZ86378.txt has the data for HGZ86378 in the 15Z state). subdirectory
 “static pH spectra” Data have been added for one additional protein
 (HGZ86378). ## Sharing/access Information Links to other publicly
 accessible locations of the data: none Was data derived from another
 source? no If yes, list source(s): n/a