Enhanced Cell Expansion in a KRP2 Overexpressor is Mediated by Increased V-ATPase Activity

Abstract

Decreased cell numbers during leaf development often trigger increased cell size, a phenomenon called compensation. In compensation-exhibiting mutants, the unusually high cell expansion activity occurs through two different mechanisms during the post-mitotic stage of leaf development, except in the KIP-RELATED PROTEIN 2-overexpressing line (KRP2 o/e), whose cell sizes are 2-fold greater during proliferative growth. However, the molecular basis of compensated cell expansion (CCE) has not been characterized. The det3-1 mutant has a mutation in the C-subunit of the vacuolar-type H(+)-ATPase (V-ATPase) complex that causes a 50% decrease in its activity and cell size. To determine the contribution of V-ATPase activity to CCE, the cellular phenotypes of double mutants between det3-1 and compensation-exhibiting fugu5-1, an3-4, fas1-5 and KRP2 o/e were analyzed in detail. Interestingly, while decreased V-ATPase activity caused by det3-1 did not suppress CCE in fugu5-1, fas1-5 and an3-4, CCE in KRP2 o/e was totally suppressed. Furthermore, measurements revealed that the activity and quantity of the A-subunit of the V-ATPase complex were significantly increased in the shoots of KRP2 o/e plants. Importantly, the unusually increased size of actively dividing KRP2 o/e cells was restored to normal in the det3-1 background. Taken together, our data strongly suggest that CCE in KRP2 o/e, but not in other compensation-exhibiting mutants, occurs exclusively through the increase of V-ATPase activity.

Class III compensation, represented byKRP2overexpression, depends on V-ATPase activity in proliferative cells

“…11 Although the molecular nature of the first 2 classes of CCEs remains unclear, the third class, represented by KIP-RELATED PROTEIN 2 (KRP2) overexpressors (o/e), was described in our recent study. 12 We demonstrated that the unusually large cell size in KRP2-overexpressing plants relies on increased vacuolar type H + -ATPase (V-ATPase) activity. This finding increased our understanding of the molecular nature of one major pathway mediating CCE in plant leaves.…”

“…This finding increased our understanding of the molecular nature of one major pathway mediating CCE in plant leaves. 12 The Arabidopsis genome is known to possess 7 KRP genes with low sequence similarities and distinct expression patterns. 13 11,13,14,15 We analyzed the cellular dynamism (i.e., cell number and size) in leaf primordia from 4 d after sowing (DAS) to leaf maturity (30 DAS).…”

“…11 Interestingly, in all compensation mutants analyzed to date, the actively dividing cells were similar in size to those in wild type, excluding class III observed in KRP2 o/e, in which the size of the dividing cells was 2-fold larger than in wild type. [11][12][13] Based on the central and often redundant functions of plant KRPs in cell cycle regulation, similarities between plants overproducing KRP1, KRP2, and KRP3 were expected. However, the reason that a gain of function triggers an increase in cell size during the cell cycle is not understood.…”

Roles of the vacuolar H⁺-PPase in seed storage oil mobilization and plant development

“…Such compensatory coordination between cell cycling and expansion is known to occur in the leaves of many mutants and transgenic lines, and has been the topic of several investigations. For example, by considering the manner through which a cell reaches its ultimate size and performing kinematic analyses of cell size dynamism, three classes of compensation have been identified: Class I, an enhanced postmitotic cell expansion rate (seen in angustifolia [an]3-4, fugu2-1/fasciata [fas]1-5, and erecta [er]-102); Class II, an extended post-mitotic cell expansion period (seen in fugu5-1); and Class III, increase in the size of dividing cells (seen in KIP-RELATED PROTEIN 2 [KRP2] over-expressing cells) (Ferjani et al 2007(Ferjani et al , 2008(Ferjani et al , 2010(Ferjani et al , 2013a(Ferjani et al , 2013b(Ferjani et al , 2014. Furthermore, during compensation, cell-autonomous and non-cell-autonomous pathways have been identified (Kawade et al 2010, 2013, Ferjani et al 2013a.…”

“…For example, by considering the manner through which a cell reaches its ultimate size and performing kinematic analyses of cell size dynamism, three classes of compensation have been identified: Class I, an enhanced postmitotic cell expansion rate (seen in angustifolia [an]3-4, fugu2-1/fasciata [fas]1-5, and erecta [er]-102); Class II, an extended post-mitotic cell expansion period (seen in fugu5-1); and Class III, increase in the size of dividing cells (seen in KIP-RELATED PROTEIN 2 [KRP2] over-expressing cells) (Ferjani et al 2007(Ferjani et al , 2008(Ferjani et al , 2010(Ferjani et al , 2013a(Ferjani et al , 2013b(Ferjani et al , 2014. Furthermore, during compensation, cell-autonomous and non-cell-autonomous pathways have been identified (Kawade et al 2010, 2013, Ferjani et al 2013a. Finally, the fact that compensation occurs in a wide range of plant species other than Arabidopsis, including tobacco, rice, and snapdragon, suggests that common developmental mechanisms that trigger compensation are widely conserved in angiosperms (Hemerly et al 1995, Barrôco et al 2006, Delgado-Benarroch et al 2009.…”