Perturbation of GABA Biosynthesis Links Cell Cycle to ControlArabidopsis thalianaLeaf Development

AbstractTo investigate the molecular mechanism underlying increasing leaf area in γ-Aminobutyric acid (GABA) biosynthetic mutants, the first pair of true leaves of GABA biosynthetic mutants was measured. The results showed that the leaf blade area in GABA biosynthetic mutants was larger than that of the wild type to different extents, and the area of the leaf epidermal cells in mutants was larger than that of the wild type. DNA polyploid analysis showed that polyploid cells in GABA biosynthetic mutants were appearing earlier and more abundant than in the wild type. To check the correlation between cell size and endoreplication, the expression of factors involving endocycles, including D-type cyclin gene (CYCD3;1, CYCD3;2, CYCD3;3, andCYCD4;1) and kinaseCKDA;1, were analysed by qRT-PCR. The results showed thatCKDA;1in GABA biosynthetic mutants was downregulated, and four types ofCYCDsshowed different expression patterns in different GABA biosynthetic mutants. Inconsistent with this result, forCCS52A(CELL CYCLE SWITCH 52A) (controlling the endocycle entry) ingad2andgad1/gad2mutants, the expression ofCCS52A2was significantly higher than that in the wild type. The expression ofSIM(SIAMESE) andSMR(SIAMESE-RELATED), which inhibit kinase activity, were also upregulated compared with the control. To further study the possible potential relationship between GABA metabolism and endoreplication, we analysed the reactive oxygen species (ROS) levels in guard cells using ROS fluorescent probes. ROS levels were significantly higher in GABA biosynthetic mutants than the control. All results indicated that cyclin, the cyclin-dependent kinase, and its inhibitory protein were coordinated to participate in endoreplication control at the transcription level in the leaves of GABA biosynthetic mutantArabidopsis.Contribution to the field statementγ-Aminobutyric acid (GABA) metabolic pathway plays a dual role in plant development. This research investigated the perturbation of GABA biosynthesis onArabidopsisleave endoreplication for the first time. In the GABA biosynthetic mutants, many genes, participating in cell division regulation, are coordinately transcriptionally expressed to trigger the onset and maintenance of endoreplication, and this led to the cell expansion and the increase leaf blade area. However, this initiation of endoreplication links with the decrease of endogenous GABA level and the increase Reactive oxygen species (ROS). This may be a compensation mechanism to adapt to abnormal GABA level in plant leaf development. Present evidence provided hypothesized that the normal GABA level in plant leaf development plays a brake to inhibit the immature cell expansion and differentiation, and this negative regulation functions a guarantee mechanism to watchdog the normal leaf development. In all, this contribution provides an updated perspective on the role of GABA in plant development.