Transcript profiling provides evidence of functional divergence and expression networks among ribosomal protein gene paralogs in Brassica napus

Abstract

The plant ribosome is composed of 80 distinct ribosomal (r)-proteins. In Arabidopsis thaliana, each r-protein is encoded by two or more highly similar paralogous genes, although only one copy of each r-protein is incorporated into the ribosome. Brassica napus is especially suited to the comparative study of r-protein gene paralogs due to its documented history of genome duplication as well as the recent availability of large EST data sets. We have identified 996 putative r-protein genes spanning 79 distinct r-proteins in B. napus using EST data from 16 tissue collections. A total of 23,408 tissue-specific r-protein ESTs are associated with this gene set. Comparative analysis of the transcript levels for these unigenes reveals that a large fraction of r-protein genes are differentially expressed and that the number of paralogs expressed for each r-protein varies extensively with tissue type in B. napus. In addition, in many cases the paralogous genes for a specific r-protein are not transcribed in concert and have highly contrasting expression patterns among tissues. Thus, each tissue examined has a novel r-protein transcript population. Furthermore, hierarchical clustering reveals that particular paralogs for nonhomologous r-protein genes cluster together, suggesting that r-protein paralog combinations are associated with specific tissues in B. napus and, thus, may contribute to tissue differentiation and/or specialization. Altogether, the data suggest that duplicated r-protein genes undergo functional divergence into highly specialized paralogs and coexpression networks and that, similar to recent reports for yeast, these are likely actively involved in differentiation, development, and/or tissue-specific processes.

Figure 1.

Expression of r-Protein ESTs and Genes across Tissues. (A) The percentages of the transcriptome composed of r-protein ESTs for each of the 16 tissues examined in B. napus. (B) The number of r-protein genes expressed per 10,000 ESTs for each tissue type (includes all paralogs per gene family).

Figure 2.

Expression of r-Protein Paralogs Relative to Tissue Type and Library Size. (A) The number of paralogs expressed per r-protein (i.e., the number of r-protein genes [all paralogs] with transcripts/number of r-proteins with transcripts) for each of 16 tissue collections in B. napus. (B) The number of paralogs expressed per r-protein versus EST data set size for each tissue (Pearson correlation coefficient, R = 0.82, P = 5 × 10⁻⁵). EST data set size was divided by 2 for libraries sequenced in both directions. Anther (A), apical meristem (AM), bud (B), early embryo (EE), embryos and seeds (ES), endosperm (E), leaves (L), MDEs, microspores (M), ovules (O), pollen (P), pollen-in vitro (PIV), root (R), seed coats (SC), seedlings (S), and stem (ST).

Figure 3.

Clustering of r-Protein Genes across Tissues Based on EST Frequencies. Hierarchical clustering of 532 r-protein genes (representing 79 r-proteins) that are differentially expressed among the 16 tissue types examined based on the transcript level per gene per tissue (transcript levels are standardized by EST data set size). Nine distinct gene clusters (A to I) were identified. The percentage of r-proteins represented by a paralog(s) in each cluster that also have paralogs in other clusters is shown. Yellow, high expression; black, low expression. Anther (A), apical meristem (AM), bud (B), early embryo (EE), embryos and seeds (ES), endosperm (E), leaves (L), MDEs, microspores (M), ovules (O), pollen (P), pollen-in vitro (PIV), root (R), seed coats (SC), seedlings (S), and stem (ST).