ANGUSTIFOLIA, a plant homolog of CtBP/BARS, functions outside the nucleus

SUMMARYCtBP/BARS is a unique protein family in having quite diversified cellular functions, intercellular localizations,and developmental roles. ANGUSTIFOLIA (AN) is the sole homolog of CtBP/BARS from Arabidopsis thaliana,although it has plant AN-specific motifs and a long C-terminus. Previous studies suggested that AN wouldfunction in the nucleus as a transcriptional co-repressor, as CtBPs function in animals; however, preciseverification has been lacking. In this paper, we isolated a homologous gene (MAN)ofAN from liverwort,Marchantia polymorpha. Transformation of the Arabidopsis an-1 mutant with 35S-driven MAN completelycomplemented the an-1 phenotype, although it lacks the putative nuclear localization signal (NLS) that existsinANproteinsisolatedfromother plantspecies.WeconstructedseveralplasmidsforexpressingmodifiedANswith amino acid substitutions in known motifs. The results clearly indicated that modified AN with mutationsin the putative NLS-like domain could complement the an-1 phenotype. Therefore, we re-examinedlocalization of AN using several techniques. Our results demonstrated that AN localizes on punctuatestructures around the Golgi, partially overlapping with a trans-Golgi network resident, which highlighted anunexpected link between leaf development and membrane trafficking. We should reconsider the roles andevolutionary traits of AN based on these findings.Keywords: ANGUSTIFOLIA, CtBP/BARS, Golgi, Marchantia polymorpha, TGN.INTRODUCTIONLeaf development is divided into two processes, cell prolif-eration and cell differentiation. In the cell differentiationprocess, genetic pathways controlling leaf cell shape havebeen revealed by the characterization of two mutants ofArabidopsis thaliana (hereafter Arabidopsis): angustifolia(an) and rotundifolia3 (rot3). The an mutant was originallyisolated by Re´dei (1962) as a mutant having a narrow leafand twisted fruit. Later studies reported that an has tric-homes with two branches (Hu¨lskampet al., 1994) and its leafcells are narrower and thicker than wild-type cells (Tsukayaet al., 1994; Tsuge et al., 1996). In contrast, rot3 has short,rounded leaves resulting from reduced cell expansion alongthe leaf length direction (Tsuge et al., 1996). These axis-specific defects in cell expansion demonstrate two inde-pendent polarity-dependent cell expansion systems (Tsugeet al., 1996; Tsukaya, 2002).