Table 1Number of links (first number) and unique genes (second number) at different FBS (final Bayesian score) cutoffs in FunCoup, where c is the corresponding confidence then value of functional coupling.The proteins with the highest connectivity are mainly related to fundamental cellular processes such as protein synthesis and degradation, translation, and transcription (Table S2). Many of them are involved in multiple processes. The most connected protein in our chicken network is the RA-related nuclear protein (RAN). Due to its various functions in nuclear transport and cell cycle regulation, it acts as a major hub with a host of other proteins. Interestingly, RAN is highly differentially expressed between male and female chicken (i.e.

, sex biased) in the gonad (FDR P < 10?4 in the adult), which is actually less common for hubs as we show in the following.3.2. Sex Bias Depends on Network ConnectivityIs there a dependency between sex bias and network connectivity? To answer this question, we first grouped the genes in three sex bias categories: male biased, female biased, and unbiased. For this we used the MWT statistic of differential expression with an FDR P value cutoff of 0.1. This was done for all four tissue/stage conditions: the embryonic and adult gonad and brain. The number of sex-biased genes in the network for each category is shown in Table 2. Remarkably, the embryonic brain contained almost no sex-biased genes and was therefore left out of this analysis. The adult brain had more sex-biased genes, but these still represented only 3% of the genes in the network.

In contrast, the gonad abounded with sex-biased genes in the network: 43% in the embryo and 82% in the adult.Table 2Number of sex-biased and unbiased genes separated by the cut-off FDR<0.1 according to MWT.Sex-biased hub genes were thus frequent in the gonad, but not in the brain, and this may be due to the fact that the male and female gonads have extensive sex-specific functions, while the brain consists of many different tissues of which only small fractions of our microarray samples may be affected by the sex. The sex-specific expression signal in the brain will therefore be diluted by the nonaffected tissues until it is no longer statistically significant. Finer-scale analysis of specific brain tissues might reveal more dimorphism in gene expression, particularly those regions related to vocalization differences between male and female birds [32] or reproductive behavior [33].

We calculated Spearman’s rank correlation coefficient between FDR values from differential expression analysis and node degree (i.e., the number of connections a gene has in the network), for AV-951 each tissue/stage combination. As can be seen in Table 3, all but one of the sex-biased categories in the gonads had a significant positive correlation at FDR P < 0.