White Lupin (Lupinus albus) response to phosphorus stress: Evidence for complex regulation of LaSAP1

Proteoid roots are a unique adaptation that allow white lupin (Lupinus albus L. var Ultra) to survive under extreme phosphorus (P) deficient conditions. The cascade of events that signals Pdeficiency induced gene expression in proteoid roots remains unknown. Through promoter::GUS analysis we showed that expression of acid phosphatase (LaSAP1) in P-deficient proteoid roots depends on DNA located from -465 bp to -345 bp 5′ of the ATG start codon and that the P1BS (PHR1 Binding Site) element, located at -160 bp, also contributes regulatory control. DNA located within the -414 bp to -250 bp region of the LaSAP1 promoter was bound by nuclear proteins isolated from P-sufficient normal roots in electrophoretic mobility shift assays (EMSA), suggesting negative regulation. Competition experiments were performed with unlabeled oligonucleotides to further delineate the region of the LaSAP1 promoter bound by P-sufficient normal root nuclear proteins to a motif spanning -361 bp to -346 bp. The promoter motif characterized through EMSA spanning -361 bp to -345 bp was used as "bait" in a yeast onehybrid (Y1H) experiment and 31 putative DNA binding proteins were isolated. Taken together, our results increase understanding of P-deficiency signaling by identifying regulatory regions and putative regulatory proteins for LaSAP1 expression.