ORIGINAL ARTICLE Yongming Zhou ® Hong Wang ® Susan Gilmer Steve Whitwill ® Wilf Keller ® Larry C. Fowke Control of petal and pollen development by the plant cyclin-dependent kinase inhibitor ICK1 in transgenic Brassica plants Received: 21 September 2001 /Accepted: 21 January 2002 / Published online: 26 March 2002 Ó Springer-Verlag 2002 Abstract The cyclin-dependent protein kinases (CDKs) have a central role in cell cycle regulation and can be inhibited by the binding of small protein CDK inhibi- tors. The first plant CDK inhibitor gene ICK1 was previously identified in Arabidopsis thaliana. In com- parison to known animal CDK inhibitors, ICK1 protein exhibits unique structural and functional properties. The expression of ICK1 directed by the constitutive CaMV 35S promoter was shown to inhibit cell division and plant growth. The aim of this study was to determine the effects of ICK1 overexpression on particular organs and cells. ICK1 was expressed in specific tissues or cells of Brassica napus L. plants using two tissue-specific pro- moters, Arabidopsis AP3 and Brassica Bgp1. Transgenic AP3-ICK1 plants were morphologically normal except for some modified flowers either without petals or with petals of reduced size. Surprisingly, petals of novel shapes such as tubular petals were also observed, indi- cating a profound effect of cell division inhibition on morphogenesis. The cell size in the smaller modified petals was similar to that in control petals, suggesting that the reduction of petal size is mainly due to the re- duction of cell numbers and that the inhibition of cell division does not necessarily lead to an increase in cell size. Transgenic Bgp1-ICK1 plants were normal mor- phologically; however, dramatic decreases in seed pro- duction were observed in some plants. In those plants, the ability of pollen to germinate and pollen nuclear number were affected. These results are discussed in re- lation to the cell cycle and plant development. Keywords Arabidopsis ® Brassica (cell cycle) ® CDK inhibitor ICK1 ® Cell cycle ® Petal ® Pollen Abbreviations CaMV: cauliflower mosaic virus ® CDK: cyclin-dependent kinase ® DAPI: 4¢,6-diamidino-2- phenylindole ® GUS: b-glucuronidase Introduction The cyclin-dependent kinase (CDK) plays a central role in regulating the cell cycle in eucaryotic organisms. The activity of CDKs can be regulated by other proteins through phosphorylation/dephosphorylation, direct binding or proteolysis (Morgan 1995; King et al. 1996). Basedonsequencesimilarityinthecyclin-bindingregion, plant CDKs can be classified into two major groups, the typical PSTAIRE group and the atypical non-PSTAIRE group (Doonan and Fobert 1997; Huntley and Murray 1999; Mironov et al. 1999). More-specific classification has recently been proposed based on a comprehensive sequence analysis of known plant CDKs (Joubes et al. 2000). In Arabidopsis, cdc2a (Arath;CDKA;1) could ge- netically complement the yeast cdc2 mutants while cdc2b (Arath;CDKB;1) could not (Ferreira et al. 1991; Hiray- ama et al. 1991). A third CDK gene cdc2c (Arath;CDKC;1) expressed only in flowers has similarity tohumanCHEDproteinkinase(Lessardetal.1999)and its functional properties remain uncharacterized. In ad- dition to CDKs, several major classes of plant cell cycle regulators such as cyclins, CDK inhibitors, Rb, E2F and Cks1 have been identified (see reviews by Huntley and Murray 1999; Mironov et al. 1999). Planta (2002) 215: 248–257 DOI 10.1007/s00425-002-0752-2 Y. Zhou ® H. Wang (&) 1 ® S. Whitwill Saskatoon Research Centre, 107 Science Place, Saskatoon, SK S7N 0X2, Canada Y. Zhou ® S. Gilmer ® L.C. Fowke Department of Biology, University of Saskatchewan, Saskatoon, SK S7N 5E2, Canada Y. Zhou National Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University, Wuhan 430070, China W. Keller Plant Biotechnology Institute, 110 Gymnasium Place, Saskatoon, SK S7N 0W9, Canada Present address: 1 Department of Biochemistry, University of Saskatchewan, Saskatoon, SK S7N 5E5, Canada e-mail: hong.wang@usask.ca Fax: +1-306-9664390