The establishment and maintenance of auxin maxima in vascular plants is regulated by auxin biosynthesis and polar intercellular auxin flow. The disruption of normal auxin biosynthesis in mouse-ear cress (Arabidopsis thaliana) leads to severe abnormalities, suggesting that spatiotemporal regulation of auxin biosynthesis is fundamental for normal growth and development. We have shown previously that the induction of the SHORT-INTERNODES/STYLISH (SHI/STY) family member STY1 results in increased transcript levels of the YUCCA (YUC) family member YUC4 and also higher auxin levels and auxin biosynthesis rates in Arabidopsis seedlings. We have also shown previously that SHI/STY family members redundantly affect development of flowers and leaves. Here, we further examine the function of STY1 by analyzing its DNA and protein binding properties. Our results suggest that STY1, and most likely other SHI/STY members, are DNA binding transcriptional activators that target genes encoding proteins mediating auxin biosynthesis. This suggests that the SHI/STY family members are essential regulators of auxin-mediated leaf and flower development. Furthermore, the lack of a shoot apical meristem in seedlings carrying a fusion construct between STY1 and a repressor domain, SRDX, suggests that STY1, and other SHI/STY members, has a role in the formation and/or maintenance of the shoot apical meristem, possibly by regulating auxin levels in the embryo.

SUMMARYThe plant hormone auxin plays fundamental roles in vascular plants. Although exogenous auxin also stimulates developmental transitions and growth in non-vascular plants, the effects of manipulating endogenous auxin levels have thus far not been reported. Here, we have altered the levels and sites of auxin production and accumulation in the moss Physcomitrella patens by changing the expression level of homologues of the Arabidopsis SHI/STY family proteins, which are positive regulators of auxin biosynthesis genes. Constitutive expression of PpSHI1 resulted in elevated auxin levels, increased and ectopic expression of the auxin response reporter GmGH3pro:GUS, and in an increased caulonema/chloronema ratio, an effect also induced by exogenous auxin application. In addition, we observed premature ageing and necrosis in cells ectopically expressing PpSHI1. Knockout of either of the two PpSHI genes resulted in reduced auxin levels and auxin biosynthesis rates in leafy shoots, reduced internode elongation, delayed ageing, a decreased caulonema/chloronema ratio and an increased number of axillary hairs, which constitute potential auxin biosynthesis sites. Some of the identified auxin functions appear to be analogous in vascular and non-vascular plants. Furthermore, the spatiotemporal expression of the PpSHI genes and GmGH3pro:GUS strongly overlap, suggesting that local auxin biosynthesis is important for the regulation of auxin peak formation in non-vascular plants.

Summary• Patterning of the Arabidopsis thaliana gynoecium is dependent on the localization and concentration of the plant hormone auxin and it has been previously reported that STYLISH1 (STY1) activates transcription of the auxin biosynthesis gene YUCCA4 (YUC4) and affects gynoecium development. Here, the relationship between auxin, STY1 and other regulators of gynoecium development was examined.• Exogenous auxin in droplets of lanolin paste were applied to young gynoecia; auxin biosynthesis rate was measured and STY1 overexpression or chemically mediated polar auxin transport (PAT) inhibition were induced in various mutants.• The style phenotype of sty1-1sty2-1 mutants was restored by exogenous application of auxin, and STY1 over-activation resulted in an elevated auxin biosynthesis rate. Both over-activation of STY1 and inhibition of PAT restored the stylar defects of several unrelated mutants, but with regard to gynoecium apical-basal patterning the mutants responded differently to inhibition of PAT.• These results suggest that reduced auxin concentrations cause the sty1-1 sty2-1 phenotype, that STY1 induces auxin biosynthesis, that elevated apical auxin concentrations can compensate for the loss of several style-promoting factors, and that auxin may act downstream of, or in parallel with these during style development but is dependent on their action in apical-basal patterning. IntroductionA characteristic feature of angiosperm plants is the fused carpels forming the gynoecium of the flower. In A. thaliana the gynoecium consists of two laterally placed carpels that fuse congenitally in the medial plane. The apical portions of the carpels fuse postgenitally to form the style and stigma. The stigma consists of a single layer of papillar cells mediating the adherence and germination of the pollen grains on the gynoecium. The style is a short solid structure connecting the stigma with the ovary. The ovary consists of the two fused carpels forming the lateral walls, or valves, and the medial replum, which internally forms the septum bisecting the ovary. Placentae with ovules form internally in association with the septal and valve margins.The plant hormone auxin has profound influence on the patterning of the gynoecium, as shown by the defects in plants with impaired auxin transport or signaling (Okada et al., 1991; Bennett et al., 1995;Sessions et al., 1997). The most apparent effect of chemical or genetic inhibition of PAT is the altered apical-basal patterning. When PAT is inhibited, the *These authors contributed equally to this work.

Auxin/indole-3-acetic acid (IAA) biosynthesis in Arabidopsis (Arabidopsis thaliana) plays a major role in growth responses to developmental and genetic signals as well as to environmental stimuli. Knowledge of its regulation, however, remains rudimentary, and few proteins acting as transcriptional modulators of auxin biosynthesis have been identified. We have previously shown that alteration in the expression level of the SHORT INTERNODES/STYLISH (SHI/STY) family member STY1 affects IAA biosynthesis rates and IAA levels and that STY1 acts as a transcriptional activator of genes encoding auxin biosynthesis enzymes. Here, we have analyzed the upstream regulation of SHI/STY family members to gain further insight into transcriptional regulation of auxin biosynthesis. We attempted to modulate the normal expression pattern of STY1 by mutating a putative regulatory element, a GCC box, located in the proximal promoter region and conserved in most SHI/STY genes in Arabidopsis. Mutations in the GCC box abolish expression in aerial organs of the adult plant. We also show that induction of the transcriptional activator DORNRÖ SCHEN-LIKE (DRNL) activates the transcription of STY1 and other SHI/STY family members and that this activation is dependent on a functional GCC box. Additionally, STY1 expression in the strong drnl-2 mutant or the drn drnl-1 puchi-1 triple mutant, carrying knockdown mutations in both DRNL and its close paralogue DRN as well as one of their closest homologs, PUCHI, was significantly reduced, suggesting that DRNL regulates STY1 during normal plant development and that several other genes might have redundant functions.