Journal of Integrative Plant Biology 2008, 50 (7): 918–927 Novel Nuclear Protein ALC-INTERACTING PROTEIN1 is Expressed in Vascular and Mesocarp Cells in Arabidopsis Fang Wang 1,2 , Dong-Qiao Shi 1 , Jie Liu 1 and Wei-Cai Yang 1 * ( 1 Key Laboratory of Molecular and Developmental Biology, National Centre for Plant Gene Research, Institute of Genetics and Developmental Biology, the Chinese Academy of Sciences, Beijing 100101, China; 2 Graduate University, the Chinese Academy of Sciences, Beijing 100039, China) Abstract Pod shattering is an agronomical trait that is a result of the coordinated action of cell differentiation and separation. In Arabidopsis, pod shattering is controlled by a complex genetic network in which ALCATRAZ (ALC), a member of the basic helix-loop-helix family, is critical for cell separation during fruit dehiscence. Herein, we report the identification of ALC-INTERACTING PROTEIN1 (ACI1) via the yeast two-hybrid screen. ACI1 encodes a nuclear protein with a lysine-rich domain and a C-terminal serine-rich domain. ACI1 is mainly expressed in the vascular system throughout the plant and mesocarp of the valve in siliques. Our data showed that ACI1 interacts strongly with the N-terminal portion of ALC in yeast cells and in plant cells in the nucleus as demonstrated by bimolecular fluorescence complementation assay. Both ACI1 and ALC share an overlapping expression pattern, suggesting that they likely function together in planta. However, no detectable phenotype was found in plants with reduced ACI1 expression by RNA interference technology, suggesting that ACI1 may be redundant. Taken together, these data indicate that ALC may interact with ACI1 and its homologs to control cell separation during fruit dehiscence in Arabidopsi s. Key words: ALC-INTERACTING PROTEIN1; ALCATRAZ; Arabidopsis; basic helix-loop-helix; nuclear protein; transcription factor. Wang F, Shi DQ, Liu J, Yang WC (2008). Novel nuclear protein ALC-INTERACTING PROTEIN1 is expressed in vascular and mesocarp cells in Arabidopsis. J. Integr. Plant Biol. 50(7), 918–927. Available online at www.jipb.net Seed dispersal or pod shattering is a result of coordinated action of cell differentiation, cell separation and programmed cell death; therefore, it is an ideal system to address these basic biological questions (Patterson 2001; Ferr ´ andiz 2002; Dinneny and Yanofsky 2005). On the other hand, premature or pod shattering at harvest causes significant loss in agricul- ture. Recent genetic studies revealed that pod shattering in Arabidopsis is controlled by a complex genetic network (Gu et al. 1998; Ferr ´ andiz et al. 2000; Liljegren et al. 2000, 2004). Understanding of the pod shattering mechanism will provide us not only with understanding of the molecular mechanisms governing these cellular processes, but also a novel strategy to Received 27 Mar. 2008 Accepted 14 Apr. 2008 Supported by a Grant from the Ministry of Science and Technology of China to W.C. Yang (JY03-A-24). ∗ Author for correspondence. Tel(Fax): +86 10 6255 1272; E-mail: <wcyang@genetics.ac.cn>. C  2008 Institute of Botany, the Chinese Academy of Sciences doi: 10.1111/j.1744-7909.2008.00694.x prevent unwanted pod shattering in agriculture, as manifested by the production of pod shattering-resistant Brassica fruit with ectopic expression of the FRUITFULL (FUL) gene (Østergaad et al. 2006). In the model species Arabidopsis thaliana, pod shattering is controlled by upstream transcription regulators including AGAMOUS (AG), SHATTERPROOF1 (SHP1), SHP2, FUL, INDIHESCENT (IND) and ALCATRAZ (ALC), and downstream genes coding for cell wall hydrolytic enzymes (Gu et al. 1998; Ferr ´ andiz et al. 2000; Liljegren et al. 2000, 2004; Rajani and Sundaresan 2001). These transcription regulators belong to two families, MADS-box family (AG, SHP1, SHP2 and FUL) and the basic helix-loop-helix (bHLH) (IND and ALC) family. The bHLH transcription factors comprise a superfamily with more than 140 and 160 members defined by the bHLH signature domain in Arabidopsis and rice, respectively (Toledo-Ortiz et al. 2003; Li et al. 2006). They are involved in many developmental processes, such as phytochrome signal transduction (Fairchild et al. 2000; Huq and Quail 2002; Duek et al. 2004; Al-Sady et al. 2006), phytohormone signaling pathway (Friedrichsen et al. 2002; Abe et al. 2003; Boter et al. 2004; Kim and Kim 2006), response to the environmental factors (Chinnusamy et al. 2003),