Qiang Wei

Functional divergences of coding genes can be caused by divergences in their coding sequences and expression. However, whether and how expression divergences and coding sequence divergences coevolve is not clear. Gene expression divergences in differentiated cells and tissues recapitulate developmental models within a species, while gene expression divergences between analogous cells and tissues resemble traditional phylogenies in different species, suggesting that gene expression divergences are molecular traits that can be used for evolutionary studies. Using transcriptomes and evolutionary proxies to study gene expression divergences among differentiated cells and tissues in Arabidopsis, expression divergences of coding genes are shown to be strongly anti-correlated with phylostrata (gene ages), indicators of selective constraint Ka/Ks (nonsynonymous replacement rate/synonymous substitution rate) and indicators of positive selection (frequency of loci with Ka/Ks > 1), but only weakly or not correlated with indicators of neutral selection (Ks). Our results thus suggest that expression divergences largely coevolve with coding sequence divergences, suggesting that expression divergences of coding genes are selectively fixed by natural selection but not neutral selection, which provides a molecular framework for trait diversification, functional adaptation and speciation. Our findings therefore support that positive selection rather than negative or neutral selection is a major driver for the origin and evolution of Arabidopsis genes, supporting the Darwinian theory at molecular levels.

Whole genome duplication (WGD) events are widespread in plants and animals, thus their long-term evolutionary contribution has long been speculated, yet a specific contribution is difficult to verify. Here, we show that ɛ-WGD and ζ-WGD contribute to the origin and evolution of bona fide brassinosteroid (BR) signaling through the innovation of active BR biosynthetic enzymes and active BR receptors from their respective ancestors. We found that BR receptors BRI1 (BR Insensitive 1) and BRL1/3 (BRI1-likes 1/3) derived by ɛ-WGD and ζ-WGD, which occurred in the common ancestor of angiosperms and seed plants, respectively, while orphan BR receptor BRL2 first appeared in stomatophytes. Additionally, CYP85A enzymes synthesizing the bioactive BRs derived from a common ancestor of seed plants while its sister enzymes CYP90 synthesizing BR precursors presented in all land plants, implying possible ligand-receptor coevolution. Consistently, the island domains (IDs) responsible for BR perception in BR receptors were most divergent among different receptor branches, supporting ligand-driven evolution. As a result, BRI1 was the most diversified BR receptor in angiosperms. Importantly, relative to the BR biosynthetic DET2 gene presented in all land plants, BRL2, BRL1/3 and BRI1 had high expression in vascular plants ferns, gymnosperms and angiosperms, respectively. Notably, BRI1 is the most diversified BR receptor with the most abundant expression in angiosperms, suggesting potential positive selection. Therefore, WGDs initiate a neofunctionalization process diverged by ligand-perception and transcriptional expression, which might optimize both BR biosynthetic enzymes and BR receptors, likely contributing to the evolution of land plants, especially seed plants and angiosperms.

All biological functions evolve by fixing beneficial mutations and removing deleterious ones. Therefore, continuously fixing and removing the same essential function to separately diverge monophyletic gene families sounds improbable. Yet, here we report that brassinosteroid insensitive1 kinase inhibitor1 (BKI1)/membrane-associated kinase regulators (MAKRs) regulating a diverse function evolved into BKI1 and MAKR families from a common ancestor by respectively enhancing and losing ability to bind brassinosteroid receptor brassinosteroid insensitive1 (BRI1). The BKI1 family includes BKI1, MAKR1/BKI1-like (BKL) 1, and BKL2, while the MAKR family contains MAKR2-6. Seedless plants contain only BKL2. In seed plants, MAKR1/BKL1 and MAKR3, duplicates of BKL2, gained and lost the ability to bind BRI1, respectively. In angiosperms, BKL2 lost the ability to bind BRI1 to generate MAKR2, while BKI1 and MAKR6 were duplicates of MAKR1/BKL1 and MAKR3, respectively. In dicots, MAKR4 and MAKR5 were duplicates of MAKR3 and MAKR2, respectively. Importantly, BKI1 localized in the plasma membrane, but BKL2 localized to the nuclei while MAKR1/BKL1 localized throughout the whole cell. Importantly, BKI1 strongly and MAKR1/BKL1 weakly inhibited plant growth, but BKL2 and the MAKR family did not inhibit plant growth. Functional study of the chimeras of their N- and C-termini showed that only the BKI1 family was partially reconstructable, supporting stepwise evolution by a seesaw mechanism between their C- and N-termini to alternately gain an ability to bind and inhibit BRI1, respectively. Nevertheless, the C-terminal BRI1-interacting motif best defines the divergence of BKI1/MAKRs. Therefore, BKI1 and MAKR families evolved by gradually gaining and losing the same function, respectively, extremizing divergent evolution and adding insights into gene (BKI1/MAKR) duplication and divergence.

Summary: Brassinosteroid (BR) signaling has been identified from the ligand BRs sensed by the receptor Brassinosteroid Insensitive 1 (BRI1) to the final activation of Brassinozole Resistant 1/bri1 EMS‐Suppressor 1 through a series of transduction events. Extensive studies have been conducted to characterize the role of BR signaling in various biological processes. Our previous study has shown that Excess Microsporocytes 1 (EMS1) and BRI1 control different aspects of plant growth and development via conserved intracellular signaling. Here, we reveal that another receptor, NILR1, can complement the bri1 mutant in the absence of BRs, indicating a pathway that resembles BR signaling activated by NILR1. Genetic analysis confirms the intracellular domains of NILR1, BRI1 and EMS1 have a common signal output. Furthermore, we demonstrate that NILR1 and BRI1 share the coreceptor BRI1 Associated Kinase 1 and substrate BSKs. Notably, the NILR1‐mediated downstream pathway is conserved across land plants. In summary, we provide evidence for the signaling cascade of NILR1, suggesting pan‐brassinosteroid signaling initiated by a group of distant receptor–ligand pairs in land plants.