Stage-Specific Regulation ofSolanum lycopersicumLeaf Maturation by Class 1 KNOTTED1-LIKE HOMEOBOX Proteins  

Abstract

Class 1 KNOTTED1-LIKE HOMEOBOX (KNOXI) genes encode transcription factors that are expressed in the shoot apical meristem (SAM) and are essential for SAM maintenance. In some species with compound leaves, including tomato (Solanum lycopersicum), KNOXI genes are also expressed during leaf development and affect leaf morphology. To dissect the role of KNOXI proteins in leaf patterning, we expressed in tomato leaves a fusion of the tomato KNOXI gene Tkn2 with a sequence encoding a repressor domain, expected to repress common targets of tomato KNOXI proteins. This resulted in the formation of small, narrow, and simple leaves due to accelerated differentiation. Overexpression of the wild-type form of Tkn1 or Tkn2 in young leaves also resulted in narrow and simple leaves, but in this case, leaf development was blocked at the initiation stage. Expression of Tkn1 or Tkn2 during a series of spatial and temporal windows in leaf development identified leaf initiation and primary morphogenesis as specific developmental contexts at which the tomato leaf is responsive to KNOXI activity. Arabidopsis thaliana leaves responded to overexpression of Arabidopsis or tomato KNOXI genes during the morphogenetic stage but were largely insensitive to their overexpression during leaf initiation. These results imply that KNOXI proteins act at specific stages within the compound-leaf development program to delay maturation and enable leaflet formation, rather than set the compound leaf route.

KNOX genes: versatile regulators of plant development and diversity

“…Ten years after KNOX overexpression was first shown to increase tomato leaf complexity, genetic evidence was provided that KNOX activity is necessary and sufficient for leaflet formation in C. hirsuta (Hay and Tsiantis, 2006). More recently, activity of the tomato Class I KNOX protein Tomato knotted 2 (Tkn2) was perturbed by means of translational fusion to a 12 amino acid EAR repressor motif of the Arabidopsis SUPERMAN gene termed SDRX, which confirmed a role for Tkn2 in tomato leaflet initiation (Shani et al, 2009).…”

“…Expression of the KNOX genes BP, KNAT2 and KNAT6 is reactivated in leaf primordia of Arabidopsis as1 mutants, producing asymmetric leaves with lobed margins and occasional shoot meristems (Byrne et al, 2000;Ori et al, 2000). Transgenic expression of each of these KNOX genes is sufficient to elicit similar alterations in Arabidopsis leaf patterning (Shani et al, 2009), and loss of all Box 3. Novel KNOX proteins lack a homeobox A new class of mini Knotted1-like homeobox (KNOX) proteins that lack a homeodomain were recently discovered in Arabidopsis and tomato, and are conserved in eudicots but not in monocots (Kimura et al, 2008;Magnani and Hake, 2008).…”

“…This study also defined a window of competence during Arabidopsis leaf development in which only proliferating, but not differentiating, tissues could respond to KN1 activity. A recent study elegantly demonstrated the importance of developmental timing for the phenotypic output of KNOX overexpression in Arabidopsis and tomato leaves, which we further discuss in the next section (Shani et al, 2009). This work raises the issue of what the molecular basis of a differential competence to respond to KNOX expression could be.…”

“…Strikingly, this correlation between the expression of KNOX genes in the leaf and a more complex leaf shape was shown to hold for tomato and many other plant species with naturally compound leaves (also known as dissected leaves; see Glossary, Box 1) (Bharathan et al, 2002;Hareven et al, 1996). It is now clear that KNOX gene expression is reactivated following leaf initiation in order to facilitate leaflet formation in both tomato and the Arabidopsis relative Cardamine hirsuta (Hay and Tsiantis, 2006;Shani et al, 2009). Gain of KNOX function in these species where KNOX genes are part of a compound leaf development programme can produce a striking reiteration of leaflets upon leaflets (see Fig.…”

“…For example, transgenic expression of any Class I KNOX gene ectopically in the leaves of Arabidopsis can dramatically alter the simple leaf margin to produce a highly lobed shape (Fig. 3E,F) (Shani et al, 2009 …”