Does P protein require a partner, as C1 protein does?
The P gene encodes a myb-domain protein, P, required for the transcriptional regulation of a subset of the flavonoid biosynthetic genes regulated by C1 (Grotewold et al., Cell 76:543, 1994), including C2, Chi1 and A1. C1, however, does not regulate transcription by itself and requires a member of the R or B gene families (encoding proteins with bHLH domains) for its activity. In agreement with the genetic evidence, P does not require R protein for the regulation of A1. Either P protein alone, or C1 protein (in the presence of R/B) are able to activate efficiently a minimal 35S promoter containing the high-affinity P binding sites identified in the A1 promoter, when transiently expressed in embryonic callus or BMS cells (Grotewold et al., 1994). In addition to these P-binding sites, other A1-promoter elements are required for normal regulation of A1 by P and by C1 proteins (Grotewold et al., 1994; Tuerck and Fromm, Plant Cell 6:1655, 1994), yet those promoter elements are not recognized by P in vitro.
P and C1/Pl1 share over 83% identity in their Myb-domains, the region responsible for the interaction between C1 and R (Goff et al., Genes Deve. 6:864, 1992). Does P require a partner, as C1 does? I decided to approach this problem by investigating the possibility that P would function in a heterologous system, and for that purpose I have chosen yeast. I introduced a reporter LacZ gene downstream of a dimeric high-affinity P-binding site (with a cyc TATA box) into the yeast chromosome. When P was expressed in yeast cells carrying this reporter construct, a very efficient activation of the reporter was obtained (over 130-fold). There was no activation when the plasmid carrying the P cDNA was absent (1-fold), nor when the P-binding sites were replaced by a dimer of a sequence to which P does not bind in vitro.
These observations are a strong indication that P is sufficient to activate transcription from promoters carrying high-affinity P-binding sites. These results are not in contradiction with the observation that other promoter elements are important for the regulation by P in plant cells. Yet, they suggest that for promoters carrying high-affinity P-binding sites, there is no need to invoke a P partner.
Not all P-regulated genes have P-binding sites. In fact, I haven't been able to identify sequences to which P binds in vitro in the promoter of the Chi1 gene, which is dependent on the presence of P for its expression in the pericarp (Grotewold and Peterson, Mol. Gen. Genet. 242:1, 1994). This could possibly mean that P regulates this promoter through a different mechanism, which could be similar to the mechanism by which A1 is regulated in the absence of high-affinity P-binding sites (Grotewold et al., 1994; Tuerck and Fromm, 1994). These alternative ways of regulation by P could involve the existence of accessory factors that recruit P to promoter elements, or that change the DNA-binding specificity of P.
to the MNL 69 On-Line Index
Return to the Maize Newsletter Index
Return to the Maize Genome Database Page