A knotted1-like homeobox gene in Arabidopsis is expressed in the vegetative meristem and dramatically alters leaf morphology when overexpressed in transgenic plants.

Abstract

The homeobox gene knottedl (knl) was first isolated by transposon tagging a dominant leaf mutant in maize. Related maize genes, isolated by virtue of sequence conservation within the homeobox, fall into two classes based on sequence similarity and expression patterns. Here, we report the characterization of two genes, KNATl and KNAT2 (forbotted-like from Arabidopsis thaliana) that were cloned from Arabidopsis using the k n l homeobox as a heterologous probe. The homeodomains of KNATl and KNAT2 are very similar to the homeodomains of proteins encoded by ClaSS 1 maize genes, ranging from 78 to 95% amino acid identity. Overall, the deduced KNATl and KNAT2 proteins sham dmino acid identities of 53 and 4096, respectively, with the KN1 protein. lntron positions are also fairly well conserved among KNAT7, KNAT2, and knl. Based on in situ hybridization analysis, the expression pattern of KNATl during vegetative development is similar to that of class 1 maize genes. In the shoot apex, KNAT7 transcript is localized primarily to the shoot apical meristem; down-regulation of expression occurs as leaf primordia are initiated. In contrast to the expression of class 1 maize genes in floral and inflorescence meristems, the expression of KNATl in the shoot meristem decreases during the floral transition and is restricted to the cortex of the inflorescence stem. Transgenic Arabidopsis plants carrying the KNAT7 cDNA and the kn7 cDNA fused to the cauliflower mosaic virus 35s promoter were generated. Misexpression of KNATl and k n l resulted in highly abnormal leaf morphology that included severely lobed leaves. The expression pattern of KNATl in the shoot meristem combined with the results of transgenic overexpression experiments supports the hypothesis that class 1 knl-like genes play a role in morphogenesis.

Isolation of an Arabidopsis homologue of the maize homeobox Knotted-1 gene

“…In addition, our data show that Merihbl/Stm has the most homology to KN1 of any Arabidopsis gene isolated. Although several other Arabidopsis homeobox genes have been isolated, their expression patterns, where known, are distinct from that of Knl [8]. The similarity in expression patterns, the high degree of similarity in predicted protein sequence, and the appearance of 'opposite' mutant phenotypes described above are consistent with Merihbl as the Knotted-1 homologue in Arabidopsis.…”

“…mRNAs suggest that they have distinct functions [8,4] (Long et al, in press). At least one plant homeobox protein, HAT3.1, has been shown to bind D N A fragments [ 16].…”

“…2. Alignment of the MERIHB1 homeodomain with the homeodomains of SBH1 [10], RS1 (personal communication), KNAT1 [8], KNAT2 [8], ATHB1 [ 14], ATHB2 [3], HAT4 [ 15], and HAT3.1 [ 16]. Amino acids identical to those of MERIHB1 are indicated by dashes.…”

The conserved ELK-homeodomain of KNOTTED-1 contains two regions that signal nuclear localization

“…Knl, originally isolated from maize by transposon tagging, is expressed primarily in meristematic cells of vegetative tissues and is normally switched off in mature leaves [41,51 ]. Ectopic expression of Knl and its related genes in maize, tobacco and Arabidopsis have shown dramatic leaf phenotypes [24,28,39]. These results have led to the hypothesis that the Kn 1 homeodomain proteins may play pivotal roles in maintaining leaf cells in an indeterminate state.…”

“…Knl is the founding member of a family of homeodomain proteins conserved in higher plants [20,52]. cDNAs related to Knl have been isolated from maize, rice, soybean, Arabidopsis thaliana and Brassica napus [2,20,24,26,28,52]. This class of proteins is characterized by a conservation of amino acid residues in the recognition helix and N-terminus of the homeodomain.…”