Genome-Wide Comprehensive Analysis of PtLACs: Prediction and Verification of the Functional Divergence of Tandem-Duplicated Genes

Laccases (EC 1.10.3.2) have been widely considered to participate in the metabolic processes of lignin synthesis, osmotic stress response, and flavonoid oxidation in higher plants. The research into Populus trichocarpa laccase focused on the synthesis of lignin in the past few years. In this study, for the first time, a comprehensive analysis of 53 laccase copies in the P. trichocarpa genome was conducted. Positive selection analysis using the branch-site model indicated that LAC genes in terrestrial plants have undergone selective pressure for adaptive evolution. On the basis of the phylogenetic relationship, we reconstructed the evolutionary process of terrestrial plant laccase and found that this gene family began to expand during the evolution of angiosperms. Tandem duplication is the main form of expansion of the PtLAC gene family. The analysis of the sequence characteristics, gene structure, expression pattern, and gene synonymous mutation rate of PtLACs provided a theoretical basis for the functional divergence of tandem duplicated genes. The synonymous mutation rate was used to quantify the divergence time of 11 tandem duplicated gene clusters. Cluster 2, with the earliest divergence time and lower share of sequence similarity, and cluster 5, with the latest divergence time and higher share of similarity, were selected in this study to explore the functional divergence of tandem-duplicated gene clusters. Tobacco subcellular localization and Arabidopsis transgenes verified the functional differentiation of PtLAC genes in cluster 2 and the functional non-differentiation of PtLAC genes in cluster 5. The results of this study provide a reference for the functional differentiation of tandem-duplicated PtLAC.