The plant hormone indoleacetic acid (IAA) transcriptionally activates early genes in plants. The Aux͞IAA family of early genes encodes proteins that are short-lived and nuclear-localized. They also contain a putative prokaryotic ␤␣␣ DNA binding motif whose formation requires protein dimerization. Here, we show that the pea PS-IAA4 and Arabidopsis IAA1 and IAA2 proteins perform homo-and heterotypic interactions in yeast using the two-hybrid system. Gel-filtration chromatography and chemical cross-linking experiments demonstrate that the PS-IAA4 and IAA1 proteins interact to form homodimers in vitro. Deletion analysis of PS-IAA4 indicates that the ␤␣␣ containing acidic C terminus of the protein is necessary for homotypic interactions in the yeast two-hybrid system. Screening an Arabidopsis -ACT cDNA library using IAA1 as a bait reveals heterotypic interactions of IAA1 with known and newly discovered members of the Arabidopsis Aux͞IAA gene family. The new member IAA24 has similarity to ARF1, a transcription factor that binds to an auxin response element. Combinatorial interactions among the various members of the Aux͞IAA gene family may regulate a variety of late genes as well as serve as autoregulators of early auxin-regulated gene expression. These interactions provide a molecular basis for the developmental and tissue-specific manner of auxin action.

Plants constantly monitor environmental fluctuations to optimize their growth and metabolism. One example is adaptive growth occurring in response to biotic and abiotic stresses. Here, we demonstrate that GH3-mediated auxin homeostasis is an essential constituent of the complex network of auxin actions that regulates stress adaptation responses in Arabidopsis. Endogenous auxin pool is regulated, at least in part, through negative feedback by a group of auxin-inducible GH3 genes encoding auxin-conjugating enzymes. An Arabidopsis mutant, wes1-D, in which a GH3 gene WES1 is activated by nearby insertion of the 35 S enhancer, exhibited auxin-deficient traits, including reduced growth and altered leaf shape. Interestingly, WES1 is also induced by various stress conditions as well as by salicylic acid and abscisic acid. Accordingly, wes1-D was resistant to both biotic and abiotic stresses, and stress-responsive genes, such as pathogenesis-related genes and CBF genes, were upregulated in this mutant. In contrast, a T-DNA insertional mutant showed reduced stress resistance. We therefore propose that GH3-mediated growth suppression directs reallocation of metabolic resources to resistance establishment and represents the fitness costs of induced resistance.

A multistep two-component signaling system is established as a key element of cytokinin signaling in Arabidopsis. Here, we provide evidence for a function of the two-component signaling system in cold stress response in Arabidopsis. Cold significantly induced the expression of a subset of A-type ARR genes and of GUS in Pro ARR7 :GUS transgenic Arabidopsis. AHK2 and AHK3 were found to be primarily involved in mediating cold to express A-type ARRs despite cytokinin deficiency. Cold neither significantly induced AHK2 and AHK3 expression nor altered the cytokinin contents of wild type within the 4 h during which the A-type ARR genes exhibited peak expression in response to cold, indicating that cold might induce ARR expression via the AHK2 and AHK3 proteins without alterations in cytokinin levels. The ahk2 ahk3 and ahk3 ahk4 mutants exhibited enhanced freezing tolerance compared with wild type. These ahk double mutants acclimated as efficiently to cold as did wild type. The overexpression of the cold-inducible ARR7 in Arabidopsis resulted in a hypersensitivity response to freezing temperatures under coldacclimated conditions. The expression of C-repeat/dehydration-responsive element target genes was not affected by ARR7 overexpression as well as in ahk double mutants. By contrast, the arr7 mutants showed increased freezing tolerance. The ahk2 ahk3 and arr7 mutants showed hypersensitive response to abscisic acid (ABA) for germination, whereas ARR7 overexpression lines exhibited insensitive response to ABA. These results suggest that AHK2 and AHK3 and the cold-inducible A-type ARRs play a negative regulatory role in cold stress signaling via inhibition of ABA response, occurring independently of the cold acclimation pathway.Cytokinins are plant hormones that regulate a variety of developmental and physiological processes, including cell division, cell proliferation, root and leaf differentiation, chloroplast biogenesis, and the inhibition of leaf senescence (1). Arabidopsis cytokinin signaling utilizes a multistep phospho-relay composed of a sensor kinase, a histidine phosphotransfer protein, and a response regulator similar to the TCS 2 of bacterial and yeast cells (2). A hybrid-type histidine kinase referred to as CYTOKININ INDEPENDENT1 (CKI1) is essential for megagametogenesis (3). CYTOKININ RESPONSE1 (CRE1)/ WOODEN LEG1 (WOL1)/ARABIDOPSIS HISTIDINE KINASE4 (AHK4) were shown to bind directly to a variety of natural and synthetic cytokinins in vitro with high specificity as well as in a yeast system and thus to be a primary receptor for cytokinins (4 -8). The experiments conducted using a heterologous phospho-relay system demonstrated that AHK2 and AHK3 are also cytokinin receptors. The primary functions of these Arabidopsis histidine kinase (AHK) genes involve the triggering of cell division and the maintenance of the meristematic competence of cells to prevent subsequent differentiation (9, 10). Partially redundant functions of cytokinin receptors have also been revealed in shoot growth, root development, leaf sen...

The LATERAL ORGAN BOUNDARIES DOMAIN/ASYMMETRIC LEAVES2-LIKE (LBD/ASL) genes encode proteins harboring a conserved amino acid domain, referred to as the LOB (for lateral organ boundaries) domain. While recent studies have revealed developmental functions of some LBD genes in Arabidopsis (Arabidopsis thaliana) and in crop plants, the biological functions of many other LBD genes remain to be determined. In this study, we have demonstrated that the lbd18 mutant evidenced a reduced number of lateral roots and that lbd16 lbd18 double mutants exhibited a dramatic reduction in the number of lateral roots compared with lbd16 or lbd18. Consistent with this observation, significant b-glucuronidase (GUS) expression in Pro LBD18 :GUS seedlings was detected in lateral root primordia as well as in the emerged lateral roots. Whereas the numbers of primordia of lbd16, lbd18, and lbd16 lbd18 mutants were similar to those observed in the wild type, the numbers of emerged lateral roots of lbd16 and lbd18 single mutants were reduced significantly. lbd16 lbd18 double mutants exhibited additively reduced numbers of emerged lateral roots compared with single mutants. This finding indicates that LBD16 and LBD18 may function in the initiation and emergence of lateral root formation via a different pathway. LBD18 was shown to be localized into the nucleus. We determined whether LBD18 functions in the nucleus using a steroid regulator-inducible system in which the nuclear translocation of LBD18 can be regulated by dexamethasone in the wild-type, lbd18, and lbd16 lbd18 backgrounds. Whereas LBD18 overexpression in the wild-type background induced lateral root formation to some degree, other lines manifested the growth-inhibition phenotype. However, LBD18 overexpression rescued lateral root formation in lbd18 and lbd16 lbd18 mutants without inducing any other phenotypes. Furthermore, we demonstrated that LBD18 overexpression can stimulate lateral root formation in auxin response factor7/19 (arf7 arf19) mutants with blocked lateral root formation. Taken together, our results suggest that LBD18 functions in the initiation and emergence of lateral roots, in conjunction with LBD16, downstream of ARF7 and ARF19.

SUMMARYCold signals interact with other environmental cues to modulate plant developmental processes. Recent studies have shown that many Pathogenesis-Related (PR) genes are induced and disease resistance is enhanced after exposure to low temperatures, linking cold signals with pathogenesis in plants. However, the underlying molecular mechanisms and signaling schemes are largely unknown. Here, we demonstrate that cold stimulates proteolytic activation of a plasma membrane-tethered NAC (NAM/ATAF1/2/CUC2) transcription factor NTL6. The transcriptionally active NTL6 protein enters the nucleus, where it induces a subset of PR genes by directly binding to a conserved sequence in the promoters of cold-responsive PR genes, such as PR1, PR2, and PR5. While transgenic plants overexpressing an active NTL6 form exhibited enhanced disease resistance, RNAi plants with reduced NTL6 activity were more susceptible to pathogen infection at low temperatures. Accordingly, cold induction of PR1 disappeared in the RNAi plants. Consistent with the close relationship between cold and pathogenesis, cold-acclimated plants showed enhanced resistance to pathogen infection. In this signaling cascade, controlled activation of the membrane-tethered, dormant NTL6 transcription factor serves as a molecular link that incorporates cold signals into pathogen resistance responses. However, the NTL6-mediated cold induction of the PR genes is independent of salicylic acid (SA). The PR genes were still induced by SA in the NTL6 RNAi plants. Cold regulation of the PR genes through the membrane-mediated transcriptional control is thought to be an adaptive process that ensures quick plant responses to incoming pathogens that frequently occur during cold seasons.