Supplementary Materials

Convergent transcriptional specializations in the brains of humans and song-learning birds

Andreas R. Pfenning, Erina Hara, Osceola Whitney, Miriam V. Rivas, Rui Wang, Petra L. Roulhac, Jason T. Howard, Morgan Wirthlin, Peter V. Lovell, Ganeshkumar Ganapathy, Jacquelyn Mouncastle, M. Arthur Moseley, J. Will Thompson, Erik J. Soderblom, Atsushi Iriki, Masaki Kato, M. Thomas P. Gilbert, Guojie Zhang, Trygve Bakken, Angie Bongaarts, Amy Bernard, Ed Lein, Claudio V. Mello, Alexander J. Hartemink, Erich D. Jarvis

Materials/Methods, Supplementary Text, Tables, Figures, and/or References

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  • Materials and Methods
  • Figs. S1 to S19
  • Captions for tables S1 to S8
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Additional Data

Tables S1 to S8
Table S1. (A) Anatomical abbreviations of human brain regions. The abbreviations come from the Allen Human Brain Atlas (, June 2013) (B) Anatomical abbreviations of macaque brain regions (available online).

Table S2. Gene annotations across species. For each species with an ENSEMBL annotated genome in the study (human, zebra finch, and macaque) the ENSEMBL ID is shown along with which microarray probes that map to those ENSEMBL IDs. The "avian_oligos" column only contains that subset of the zebra finch oligos that map reliably to the genomes of other avian species profiled (budgerigar, Anna's hummingbird, ring dove, and quail). When available, the corresponding zebra finch protein ID is shown that was used to map genes to proteins in the expression-proteomics comparison. The gene name, symbol, and mappings across species are obtained from BioMart (, ENSEMBL version 64 (available online).

Table S3. Genes with common specializations between zebra finch and human brain regions. (A) Zebra finch striatum (Area X + VS) specialized genes shared with different human telencephalic regions. (B) Zebra finch pallial specialized genes shared with different human telencephalic regions. (C) Zebra finch Area X specialized genes shared with different human
striatal regions. (D) Zebra finch RA specialized genes shared with different human cortical regions; genes highlighted in yellow are also shared with layer 5 motor cortex of macaques. (E)
Zebra finch HVC specialized genes shared with different human cortical regions. (F) Zebra finch LMAN specialized genes shared with different human cortical regions. For each sub-table, the first six columns contain gene and microarray oligo annotations. The "permutation p-value" gives the statistical significance for the gene being specialized in the best matched brain regions between zebra finch (column H) and human (column J). The remaining columns contain a summary of the linear model results conducted on the zebra finch brain region and multiple other human brain regions. "1" means that the gene was significantly (adj. p < 0.05) increased in the region compared to the entire telencephalon for finch or for human. "–1" corresponds to a decrease and "0" corresponds to a lack of a significant difference. Only genes with a common specialization between finch and any of the human sub-regions are listed (available online).

Table S4. The relationship between human LMC and avian RA across species. (A-C) The specialization of RA (or analogous region) in the vocal learning species (zebra finch, budgerigar, hummingbird) and vocal non-learning species (dove and quail), compared to the specialization of different human subregions (PrG-cs vs. cortex; LMC/dLSC vs. cortex; LMC/dLSC vs. PrG/PoG). (D-E) Comparison of the arcopallium regions of avian species with macaque layer 5 cells alone and human LMC/dLSC vs. macaque layer 5 cells. For each comparison, shown is the correlation of the specializations and the number of significantly overlapping specialized genes observed vs. the number expected by chance. Hypergeometric p-values give the significance of the overlap with the LMC/dLSC specializations. For the specializations within and relative to macaque we show the permutation p-value for how often that correlation is reached by chance. Significant values are shown in bold (available online).

Table S5. Gene ontology analyses of specialized genes. Analysis was conducted using ingenuity pathway analysis (IPA; Ingenuity® Systems, Only categories of at least 5 24 genes are presented. (A) Zebra finch (Area X + VS) and human striatum regions compared to different human telencephalic regions. (B) Zebra finch pallium and sub-regions of the human forebrain. (C) Zebra finch Area X and sub-regions of the human striatum. (D) Zebra finch RA and sub-regions of the human cortex. (E) Convergently specialized genes. (F) Proteins specialized in zebra finch RA. Because we did not discriminate between up- and downregulated genes, categories contain both and could represent a mix of present vs. absence of enriched functions (available online).

Table S6. Genes with common specializations between avian RA analogues and human LMC/LSC. The first five columns contain gene and microarray oligo annotations. Symbols for
genes that have been validated by in situ hybridization in zebra finch are highlighted in yellow. False positives are highlighted in orange. Remaining columns show p-values of the linear model for different brain region comparison for each specific gene (p < 0.1). The significance of genes is distinguished by their color (green: p < 0.05; blue: 0.05 < p <0.1; red: p > 0.1). A non-adjusted p-value was used because of the numerous criteria that were required to be included in the linear model list. The genes shown met criteria for convergent evolution described in (SM8). They are broken down into sections based on whether the genes are also significantly specialized in the hummingbird RA analogue relative to the neighboring arcopallium (column R; sections A, B), in the LMC/LSC samples relative to PrG and PoG (sections A, C), or for the combined set of all laryngeal cortex regions (vLMC, dLMC, and dLSC; section D). The direction of specialization and permutation false discovery rate between vocal learners and non-learners is shown in columns S and T (available online).

Table S7. Coordinated differences between mRNA and protein levels in RA versus the surrounding arcopallium. The first four columns contain gene and microarray oligo annotations. Column E list the protein expression difference (specialization) of the mean RA gene expression minus the mean arcopallium gene expression detected my mass spec analyses. Column F lists the difference in levels of the corresponding mRNAs detected by microarray analyses (available online).

Table S8. Accession numbers of clones used for in situ hybridizations (available online).