Summary KNUCKLES ( KNU ) encodes a C 2 H 2 zinc-finger protein that regulates development of basal pattern elements of the Arabidopsis gynoecium

The primordia of plant lateral organs arise from the flanks of pluripotent meristems. After specification, organogenesis proceeds from the coordination of two complementary processes: cellular proliferation and differentiation. How these processes are coordinated, and how positional information is generated and regional identity maintained over the course of organ development, are fundamental questions which genetic studies of flower and fruit development have begun to address. Arabidopsis thalianafloral meristems each give rise to four concentric whorls of determinate lateral organs: four sepals, four petals, six stamens, and two fused carpels (the gynoecium). Thirdand fourth-whorl floral organs contain the reproductive male (micro-) and female (mega-) gametophytes, respectively. Each megagametophyte or embryo sac is enclosed by a specialized sporophytic structure, the ovule, which may give rise to a seed if fertilized. After anthesis (pollen shedding) and fertilization, the gynoecium lengthens considerably and develops into a seedcontaining silique, the Arabidopsisfruit (Fig. 1A). Elongation of the silique is accomplished primarily by longitudinal expansion of cells of the exocarp, schlerenchyma, and endocarp layers, whereas cell division accounts for most growth within the mesocarp (Vivian-Smith and Koltunow, 1999). A dehiscence zone allows for separation of the valves from the replum and seed dispersal (Ferrandiz et al., 1999). In the absence of fertilization, the gynoecium may undergo restricted post-anthesis elongation, but it remains a determinate organ that eventually abscises. Parthenocarpic Arabidopsis mutants, in which the gynoecium undergoes post-anthesis fruit development without fertilization, have been isolated (Ito and Meyerowitz, 2000; Vivian-Smith et al., 2001). The application of exogenous plant growth regulators to emasculated flowers induces fertilization-independent fruit development in Arabidopsis (Vivian-Smith and Koltunow, 1999), and transgenic Solanaceae producing elevated levels of auxin within placental tissue and ovules are also parthenocarpic (Rotino et al., 1997; Ficcadenti et al., 1999). It is likely that parthenocarpic mutants either produce abnormal levels of growth regulators or respond inappropriately to these compounds. Regulation of floral organ specification can be explained in terms of the ABC model (Bowman et al., 1991; Coen and Meyerowitz, 1991). The model posits three functional classes of regulators (A, B, and C) expressed in partially overlapping domains. Where expression of the B-function MADS box genes APETALA3(AP3) (Jack et al., 1992) and PISTILLATA (PI) (Goto and Meyerowitz, 1994) overlaps that of the Cfunction MADS box gene AGAMOUS(AG) (Yanofsky et al., 1990), stamens form in the third whorl. AG alone specifies Flowers of the parthenocarpic knuckles mutant are conditionally male sterile and contain ectopic stamens and carpels that originate from placental tissue within developing gynoecia. The mutation was mapped to a 123 Kb interval on chromosome 5 using molecular markers. All aspects of the knucklesphenotype could be complemented by a genomic fragment from the region which contained the annotated MAC12.2 gene. A guanine to adenine transition within a predicted C2H2 zinc finger-encoding region of MAC12.2causes the second of two critical zincbinding cysteine residues to be replaced by a tyrosine. Transgenic plants in which translational fusions of the GUS reporter to KNUCKLES were driven by the presumptive KNUCKLES promoter indicate that the gene is expressed first in developing carpel primordia, and later in stamens and ovules of flower buds. In situ hybridization experiments showed a broader pattern of transcript localization, suggesting that post-transcriptional regulatory mechanisms may limit KNUCKLES protein accumulation and localization. Based on genetic evidence and the presence of a carboxy-terminal motif demonstrated by others to function as an active repression domain, we propose that KNUCKLES might function as a transcriptional repressor of cellular proliferation that regulates floral determinacy and relative size of basal pattern elements along the proximo-distal axis of the developing Arabidopsisgynoecium.