School of Bio Science and Technology, Vellore Institute of Technology, Vellore, 632014
Leaves are the lateral vegetative organs on a plant stem that carry out vital functions such as photosynthesis and respiration. They are initiated at the flanks of the shoot apical meristem (SAM) as small bulges called primordia, which later develop into mature leaves. Development of a mature leaf involves two main cellular processes, namely proliferation and differentiation. During proliferation, cells divide rapidly with mitosis-associated cell growth, while in the differentiation phase the cells undergo post-mitotic expansion. Several genes are known to be involved in the regulation of these two processes. An important group of leaf morphogenesis regulators is the TCP family of proteins. They are plant-specific transcription factors that have a conserved TCP domain involved in DNA binding. Out of 24 TCP proteins encoded by the Arabidopsis genome, 13 are classified into class I and the rest fall under class II. This classification is based on features within the TCP domain and their binding specificities. Functionally, most class I TCP proteins act as positive regulators of growth by inducing proliferation, even though exceptions exist. On the other hand, the class II TCP proteins act as negative regulators of growth by suppressing proliferation and inducing differentiation. Five class II TCPs - namely TCP 2, 3, 4, 10 and 24 - are regulated by the micro RNA miR319. The main aim of this project was to perform a detailed study on the role of these miR319-targeted TCP proteins in leaf morphogenesis. The approaches include biochemical characterization of these proteins and establishing gain-of-function lines of two family members - TCP3 and TCP4. Through the biochemical approach, it was earlier shown that all the five miR319-targeted TCPs bind to class II binding sequence, while TCP3 binds to the class I sequence as well. In this study, a His-tagged form of TCP3 protein has been purified using Ni-NTA beads. In the genetic approach, we have selected transgenic lines (pTCP3::mTCP4:GR and 35S::mTCP3:GR) in the T2 generation that show a strong phenotype. They will be further carried forward to establish homozygosity.
Keywords or phrases: Shoot Apical Meristem, TCP proteins, DNA-binding transcription factors, Growth Regulators, Leaf Morphogenesis.