Plant Molecular Biology 50: 237–248, 2002. © 2002 Kluwer Academic Publishers. Printed in the Netherlands. 237 Molecular characterization of AtNAM: a member of the Arabidopsis NAC domain superfamily Manuel Duval 1 , Tzung-Fu Hsieh 1 , Soo Young Kim 2 and Terry L. Thomas 1,∗ 1 Department of Biology, Texas A&M University, College Station, TX 77843, USA ( ∗ author for correspondence; e-mail tlthomas@tamu.edu); 2 Kumho Life and Environmental Science Laboratory, 572 Ssang-Am-Dong, Kwang- San-Gu, Kwangju 506-712, Korea Received 14 March 2001; accepted in revised form 1 February 2002 Key words: DNA-binding domain, shoot meristem, transcriptional activation Abstract The petunia NAM and Arabidopsis ATAF1 and CUC2 genes define the conserved NAC domain. In petunia, loss-of- function nam mutants result in embryos that fail to elaborate shoot apical meristems (SAM), and nam seedlings do not develop shoots and leaves. We have isolated a NAC domain gene, AtNAM, from an Arabidopsis developing seed cDNA library. Expression of AtNAM mRNA is restricted primarily to the region of the embryo including the SAM. The AtNAM gene contains three exons and is located on Chromosome 1. In vivo assays in yeast demonstrate that AtNAM encodes a transcription factor and that the NAC domain includes a specific DNA binding domain (DBD). The AtNAM DBD is contained within a 60 amino acid region which potentially folds into a helix-turn-helix motif that specifically binds to the CaMV 35S promoter. The putative transcriptional activation domain is located in the C-terminal region of the protein, a highly divergent region among NAC domain-containing genes. The Arabidopsis genome contains 90 predicted NAC domain genes; we refer to these collectively as the AtNAC superfamily. The first two exons of all members of this superfamily encode the NAC domain. Most AtNAC genes contain three exons with the last exon encoding an activation domain. A subfamily of AtNAC genes contains additional terminal exons coding for protein domains whose functions are unknown. Abbreviations: SAM, shoot apical meristem; NAC, NAM, ATAF1, and CUC2, DBD, DNA-binding domain Introduction Plant morphogenesis requires precise regulation of the rate of cell division, the orientation and position of the cell division plane and cell expansion. For example, the first cell division that follows egg fertilization in Arabidopsis is always asymmetric and occurs perpen- dicularly with respect to the longitudinal axis of the ovule. After the 2-cell stage, cell division conforms to a strict pattern and indeed determines the shape and the size of the embryo. Embryogenesis specifies the three polarity axes as well as the main organs and tissues of the plant. The nucleotide sequence reported will appear in the GenBank database under the accession number AF123311(AtNAM). Postembryonic development and growth is fueled by cell division occurring in specialized tissues re- ferred to as meristems. At the shoot apex lies the shoot apical meristem (SAM) which is specified as the em- bryo enters the early maturation stage. The SAM is a group of undifferentiated cells that generates the aerial organs of the plant. These cells keep their pluripotency and represent the reservoir of cells for the continu- ous growth of the plant during its life cycle. In the shoot apical meristem, the rate of cell division is bal- anced with the rate of cell differentiation to achieve a steady state with respect to the number of cells that constitutes the SAM. Homologues of the petunia NO APICAL MERIS- TEM gene have been shown to be involved in the development and maintenance of the shoot apical