Enrichment analyses
To test our prediction that males would exhibit increased expression of
genes associated with vascularization, we conducted Gene Ontology (GO)
enrichment analyses using the R package topGO to identify
biological processes that were enriched in genes differentially
expressed between males and females
(Alexa & Rahnenfuhrer,
2020). We searched for terms related to angiogenesis, the formation of
new blood vessels, including the phrase “angio” and removed those not
involved in angiogenesis (i.e., “lymphangiogenesis”). We compared the
standardized effect of sex for these angiogenesis genes with the
standardized effect of sex for all other detectably expressed genes with
a Kolmogorov-Smirnov test.
To test our prediction that males would have higher expression of genes
associated with estrogen and androgen regulation, we used the Online
Predicted Human Interaction Database (OPHID) to assess protein-protein
interaction (PPI) networks associated with hormone receptor proteins of
interest: ER\(\alpha\), ER\(\beta\), and AR
(Brown & Jurisica, 2005).
We used ENSEMBL orthology information queried through the R packagebiomaRt (Durinck et
al., 2009) to identify one-to-one orthologs in the human genome, then
filtered and reindexed our expression matrix to detectably expressed
genes in the human genome. We retained a total of 8,255 human genes for
analysis. Next we ran a query in OPHID to identify the set of human
genes involved in PPI networks with each hormone receptor protein and
used biomaRt to convert genes names from UniProt to Ensembl
(The UniProt Consortium,
2021). We found the genes associated with PPI networks for
ER\(\alpha\), ER\(\beta\), and AR and compared the standardized effect
of sex for these genes with the standardized effect of sex for all other
detectably expressed genes with human orthologs with a
Kolmogorov-Smirnov test.