Plants grown under dense canopies perceive through the phytochrome system a reduction in the ratio of red to far-red light as a warning of competition, and this triggers a series of morphological changes to avoid shade. Several phytochrome signaling intermediates acting as positive regulators of accelerated elongation growth and induction of flowering in shade avoidance have been identified. Here we report that a negative regulatory mechanism ensures that in the presence of far-red-rich light an exaggerated plant response does not occur. Strikingly, this unpredicted negative regulatory mechanism is centrally involved in the attenuation of virtually all plant responses to canopy shade.Supplemental material is available at http://www.genesdev.org.

Homeodomain-leucine zipper (HD-Zip) proteins are putative transcription factors identified only in plants. Related Arabidopsis homeobox genes, isolated by virtue of sequence conservation within the helix-3 region of the homeodomain, fall into four families based on sequence similarity. This paper reports the characterization of Athb-10, a 747 amino acid protein belonging to the fourth HD-ZIP family. The studies indicate that, although less conserved, the leucine zipper of Athb-10 can functionally replace that of Athb-2 in an in vitro DNA-binding assay. Gene mapping experiments and sequence comparison analysis revealed that Athb-10 corresponds to GLABRA2, a homeodomain protein involved in trichome development. The mRNA expression analysis revealed that Athb-10/GLABRA2 is expressed not only in trichome-bearing organs, but also in the root. The analysis of wild-type and mutant plants showed that the Athb-10/GLABRA2 gene expression in the aerial part of the plant and in the root is affected by mutations at the TTG locus. Morphological analysis of the g/2-1 mutant revealed that the gene is necessary not only for local outgrowth of the trichome, but also for the regulation of root hair development in a subset of epidermal cells. Interestingly, the development of root hair cells in a position normally occupied by non-hair cells is dependent upon the ethylene regime in which the gl2-1 plants are grown. Sequence analysis of the gl2-1 allele revealed that the mutant gene encodes a truncated protein that might still retain a partial activity responsible for the formation of aborted trichomes and for the ethylene-dependent regulation of root hair formation.

The Arabidopsis Athb‐1 and −2 proteins are characterized by the presence of a homeodomain (HD) with a closely linked leucine zipper motif (Zip). We have suggested that the HD‐Zip motif could, via dimerization of the leucine zippers, recognize dyad‐symmetric DNA sequences. Here we report an analysis of the DNA binding properties of the Athb‐1 homeodomain‐leucine zipper (HD‐Zip‐1) domain in vitro. DNA binding analysis performed using random‐sequence DNA templates showed that the HD‐Zip‐1 domain, but not the Athb‐1 HD alone, binds to DNA. The HD‐Zip‐1 domain recognizes a 9 bp dyad‐symmetric sequence [CAAT(A/T)ATTG], as determined by selecting high‐affinity binding sites from random‐sequence DNA. Gel retardation assays demonstrated that the HD‐Zip‐1 domain binds to DNA as a dimer. Moreover, the analysis of the DNA binding activity of Athb‐1 derivatives indicated that a correct spatial relationship between the HD and the Zip is essential for DNA binding. Finally, we determined that the Athb‐2 HD‐Zip domain recognizes a distinct 9 bp dyad‐symmetric sequence [CAAT(G/C)ATTG]. A model of DNA binding by the HD‐Zip proteins is proposed.

The Arabidopsis genome contains 10 genes belonging to the HD-Zip II family including ATHB2 and HAT2. Previous work has shown that ATHB2 is rapidly and strongly induced by light quality changes that provoke the shade avoidance response whereas HAT2 expression responds to auxin. Here, we present a genome-wide analysis of the HD-Zip II family. Phylogeny reconstruction revealed that almost all of the HD-Zip II genes can be subdivided into 4 clades (alpha-delta), each clade comprising 2-3 paralogs. Gene expression studies demonstrated that all the gamma and delta genes are regulated by light quality changes. Kinetics of induction, low R/FR/high R/FR reversibility and auxin response analyses strongly suggested that HAT1, HAT3 and ATHB4, as ATHB2, are under the control of the phytochrome system whereas HAT2 is up-regulated by low R/FR as a consequence of the induction of the auxin signaling pathway provoked by FR-rich light. Root and shoot digital in situ revealed that gamma and delta genes are also tightly regulated during plant development with both distinct and overlapping patterns. Phenotypes of gain of function and dominant negative lines demonstrated that one or more of the HD-Zip II gamma genes negatively regulate cell proliferation during leaf development in a high R/FR light environment. Finally, target gene analysis using a chimeric transcription factor (HD-Zip2-V-G), known to activate ATHB2 target genes in a glucocorticoid-dependent manner, revealed that all the 10 HD-Zip II genes can be recognized by the HD-Zip 2 domain in vivo, implying an intricate negative feedback network.