DEVELOPMENT 3099 RESEARCH ARTICLE INTRODUCTION The final size and shape of plant organs are determined by developmental programs that coordinate cell proliferation, cell expansion and cell-type differentiation (Mizukami, 2001; Potter and Xu, 2001). The Arabidopsis ovule has a relatively simple structure and a specific, well-defined differentiation pattern, making it a useful model for understanding the regulation of growth and organogenesis. The ovule originates from the placenta as an elongate protrusion with a defined proximal/distal axis that can be separated into three regions. The distal portion, called the nucellus, is the site of megasporogenesis and embryo sac development. The proximal region differentiates into the funiculus or stalk, which attaches the ovule to the carpel wall. Two integuments initiate at the central chalazal region and eventually envelop the nucellus and form the seed coat. Similar to the process of leaf laminar expansion, initiation and expansion of the integument requires the juxtaposition of abaxial and adaxial factors such as those encoded by PHABULOSA (PHB), KANADI1 (KAN1), KAN2 and KAN3, ABERRANT TESTA SHAPE (also known as KAN4 – TAIR), and the YABBY-family member INNER NO OUTER (Eshed et al., 2001; Eshed et al., 2004; McAbee et al., 2006; Sieber et al., 2004; Villanueva et al., 1999). In addition, transcription factors such as AINTEGUMENTA and NOZZLE (also known as SPOROCYTELESS), and the mitochondria ribosomal protein HUELLENLOS, have been identified as necessary for integument initiation (Elliott et al., 1996; Schneitz et al., 1998; Skinner et al., 2004; Villanueva et al., 1999). A second group of loci affect the progression of integument growth after initiation. Several genes, including PRETTY FEW SEEDS 2 (PFS2), SHORT INTEGUMENTS 1 and 2 (SIN1 and 2) and TSO1, affect cell proliferation or expansion of the integuments, and their loss-of-function mutations result in reduction or loss of fertility (Hauser et al., 2000; Park et al., 2004; Park et al., 2005; Schneitz et al., 1997). Embryo sac failure is a secondary consequence of the absence of integuments; the gametophyte fails to develop in cases where the nucellus is not enclosed (Gasser et al., 1998). The cause of gametophyte abortion is not clear, but is likely to lie in the requirement for tight coordination of cell division and expansion within the ovule and communication between gametophytic and sporophytic tissue (Gasser et al., 1998). Relatively little is known about the genes involved in the coordination of growth or cell-cell communication within the ovule. This might be due to redundancy among the genes involved in this process, or to pleiotropic effects resulting from the loss of these genes. For instance, TSO1, TOUSLED (TSL) and SIN1 have multiple, sometimes detrimental, effects on vegetative and floral development in addition to ovule defects (Ehsan et al., 2004; Lang et al., 1994). ERECTA (ER) and its two paralogs, ERECTA-LIKE 1 (ERL1) and ERL2, regulate organ shape and inflorescence architecture and are members of the leucine-rich repeat receptor-like kinase (LRR-RLK) gene family in Arabidopsis (Shiu and Bleecker, 2001; Torii, 2004; Torii et al., 1996). Based on phylogeny, it was suggested that ERL1 and ERL2 evolved by recent gene duplication and are functionally related to ER, while maintaining overlapping but unique transcript expression patterns (Shpak et al., 2004). We report here that in the absence of functional ER and ERL1, Arabidopsis plants heterozygous at the ERL2 locus exhibit specific defects in integument development. Based on data from genetic and gene expression analysis, the ER family plays a key role in ovule development and fertility by regulating cell proliferation in the integuments. Our study highlights the unequal redundancy and dosage compensation among ER-family genes, and further reveals Haploinsufficiency after successive loss of signaling reveals a role for ERECTA-family genes in Arabidopsis ovule development Lynn Jo Pillitteri, Shannon M. Bemis, Elena D. Shpak* and Keiko U. Torii † The Arabidopsis genome contains three ERECTA-family genes, ERECTA (ER), ERECTA-LIKE 1 (ERL1) and ERL2 that encode leucine- rich repeat receptor-like kinases. This gene family acts synergistically to coordinate cell proliferation and growth during above- ground organogenesis with the major player, ER, masking the loss-of-function phenotypes of the other two members. To uncover the specific developmental consequence and minimum threshold requirement for signaling, ER-family gene function was successively eliminated. We report here that ERL2 is haploinsufficient for maintaining female fertility in the absence of ER and ERL1. Ovules of the haploinsufficient er-105 erl1-2 erl2-1/+ mutant exhibit abnormal development with reduced cell proliferation in the integuments and gametophyte abortion. Our analysis indicates that progression of integument growth requires ER-family signaling in a dosage-dependent manner and that transcriptional compensation among ER-family members occurs to maintain the required signaling threshold. The specific misregulation of cyclin A genes in the er-105 erl1-2 erl2-1/+ mutant suggests that downstream targets of the ER-signaling pathway might include these core cell-cycle regulators. Finally, genetic interaction of the ER family and the WOX-family gene, PFS2, reveals their contribution to integument development through interrelated mechanisms. KEY WORDS: Arabidopsis, Integument growth, Ovule, Receptor-like kinase, Haploinsufficient, Cell proliferation Development 134, 3099-3109 (2007) doi:10.1242/dev.004788 Department of Biology, University of Washington, Seattle WA 98195 USA. *Present address: Department of Biochemistry and Cellular and Molecular Biology, University of Tennessee, Knoxville, TN 37996, USA † Author for correspondence (e-mail: ktorii@u.washington.edu) Accepted 20 June 2007