One consequence of gene duplication is that it allows the two copies to subdivide the functions of the ancestral copy (functional specialization or sub-functionalization), or for one copy to acquire a new function (neo-functionalization) while the other retains the original function, thus preserving fitness; but is there evidence for either functional specialization or acquisition of novel functions among PRR genes during evolution of the angiosperms? The strongest evidence comes from Arabidopsis, where clock function is best studied. TOC1 and four other PRR genes each show circadian oscillations in transcript abundance, with peak abundance occurring at intervals spanning the day starting at dawn with PRR9, followed by PRR7, PRR5, PRR3, and finally at dusk with TOC1 (PRR1) [4, 5]. As shown in Figure 2, TOC1 is recruited to the CCA1 promoter and is a positive regulator of CCA1 expression, although the molecular details remain incompletely described [1]. PRR9, PRR7, and PRR5 are recruited to the promoters of CCA1 and LHY and negatively regulate their expression [10]. It is likely that the sequential expression of PRR9, PRR7, and PRR5 contributes to sustained repression of CCA1 and LHY expression throughout the day. This indicates that, while the function of these three genes is partially redundant, with normal expression of the three genes the temporal window of CCA1/LHY repression is extended. Thus PRR9, PRR7, and PRR5 offer an example of sub-functionalization in the temporal domain. Although the function of the rice (Oryza sativa) orthologs of PRR9, PRR7, and PRR5 has not been established, there is a similar sequential pattern of expression of OsPRR73/OsPRR37 and then OsPRR95/OsPRR59, followed by OsTOC1 (OsPRR1) [11].
In Arabidopsis, the PRR3 gene offers an example of acquisition of a novel function. PRR9, PRR7, and PRR5 all have a similar role in negatively regulating CCA1 and LHY, suggesting that this represents the ancestral function (Figure 2). PRR3 appears, instead, to have acquired a novel and specialized function in the vascular tissue, where PRR3 binds to TOC1 and, in doing so, blocks the interaction of TOC1 with ZTL, the F-box protein that targets TOC1 for proteasomal degradation [12]. Thus, PRR3 exhibits a restricted domain of expression and has acquired a novel function, the regulation of TOC1 stability through protein-protein interaction (Figure 2). In Arabidopsis, loss of PRR3 function confers only a very small shortening of circadian period [13], which is consistent with the apparent loss of PRR3 in poplar, without concomitant perturbation of clock function.
There are additional suggestions of evolving function in the PRR7 lineage. In Arabidopsis, PRR7 contributes to the determination of flowering time, although the effects are not large and PRR7 is not a major determinant of flowering time among natural populations [14]. In contrast, in the monocots barley and wheat, PRR7 (Ppd-H1 and Ppd-D1, respectively) is one of the major determinants of photoperiod sensitivity and flowering time [15, 16]. Whether this represents a true acquisition of novel function in the monocots or a loss of function in the eudicots remains uncertain and will require more detailed dissection of the roles of PRR7 in the flowering pathways of monocots and eudicots.