BIOLOGIA PLANTARUM 55 (4): 641-646, 2011 641 Early stages of leaf development in has mutant of Arabidopsis thaliana D. JANOŠEVIĆ 1 *, B. UZELAC 2 , D. STOJIČIĆ 3 , Lj. TUBIĆ 2 and S. BUDIMIR 2 Institute of Botany and Botanical Garden "Jevremovac", Faculty of Biology, University of Belgrade, Takovska 43, 11000 Belgrade, Serbia 1 Institute for Biological Research "Siniša Stanković", University of Belgrade, Bulevar despota Stefana 142, 11060 Belgrade, Serbia 2 Faculty of Science, University of Niš, Višegradska 33, 18000 Niš, Serbia 3 Abstract The elucidation of molecular mechanisms underlying the leaf development can be facilitated by the detailed anatomical study of leaf development mutants. We present an analysis of leaf anatomy and morphogenesis during early developmental stages in has mutant of Arabidopsis thaliana. The recessive has mutation affects a number of aspects in plant development, including the shape and size of both cotyledons and leaves. The earliest developmental observations suggest almost synchronous growth of the first two leaf primordia of has mutant. No significant disruption of the cell division pattern in the internal tissue is observed at the earliest stages of development, with the major anatomical difference compared to wild type primordia being the untimely maturation of mesophyll tissue cells in has mutant. At the stage of leaf blade formation, structure disruption becomes clearly evident, by irregular arrangement of the cell layers and the lack of polarity in juvenile has leaves. One distinguishing feature of the mutant leaf anatomy is the absence of mesophyll tissue differentiation. Altered has mutant leaf morphology could be at least partially accounted for by the ectopic STM activity that was found at the base of leaf primordia during early stages of leaf development in has plants. Additional key words: in situ hybridization, insertional mutagenesis, leaf morphogenesis. Introduction Leaves are determinate, bilaterally symmetrical, lateral organs which are produced from the cells recruited from the shoot apical meristem (SAM). Leaf primordium initiation involves the formation of an outgrowth on the surface of the meristem by periclinal cell divisions in the inner layers of peripheral zone of SAM. A number of genes have been associated with processes necessary for organ formation (Clark 1997, Barton 2001, Traas and Doonan 2001). A very early event in organ initiation is the inactivation of SHOOT MERISTEMLESS (STM) gene, a member of KNOX family of homeobox genes in Arabidopsis (Jackson et. al. 1994, Long et al. 1996, Reiser et al. 2000, Barton 2001). These meristematic genes are necessary for maintaining the pool of meristematic cells at the shoot apex and are expressed in the SAM but not in the presumptive leaf primordium, the initiating leaf primordium and developing leaf (Kerstetter et al. 1994, Long et al. 1996). Downregulation of KNOX genes in lateral organ primordia is a critical event in organ patterning, as the ectopic expression of these genes disrupts normal leaf development (Byrne et al. 2000, Chuck et al. 1996, Reiser et al. 2000). Set of genes other than these are also associated with the first steps in organ initiation, which include the attribution of cells to primordia, organ outgrowth, organ separation, and determination of organ identity (Traas and Doonan 2001). In the course of development, the three-dimensional form of leaf is specified. Leaf develops and differentiates along proximodistal (base-tip), dorsoventral (top-bottom) and mediolateral (middle-to-margin) planes (Steeves and Sussex 1989, Kim and Cho 2006). After the leaf polarity ⎯⎯⎯⎯ Received 19 April 2010, accepted 21 June 2010. Abbreviations: MS - Murashige and Skoog; SAM - shoot apical meristem; SEM - scanning electron microscopy; STM - SHOOT MERISTEMLESS gene. Acknowledgements: The Ministry of Science and Technological Development of Serbia, Grant No. 173015, supported this research. * Corresponding author, fax: (+381) 11 2761 433, e-mail: dusicaj@ibiss.bg.ac.rs