glycyrrhizin To date little is known
To date, little is known about the regulation of PCD through phytocystatin inhibitory activity. Inhibitors that may regulate leaf senescence (Huang et al., 2001, Tajima et al., 2011) or may inhibit PCD induced by abiotic factors, such as mechanical tissue damage or oxidative stress caused by cold (Belenghi et al., 2003, Solomon et al., 1999) have been found in dicotyledons. However, only one phytocystatin, tobacco NtCYS1, which inhibits PCD in the suspensor during the early stages of embryogenesis, is known to participate in inhibiting PCD during seed development. The role of the suspensor is to transport nutrients and hormones to the embryo and when it is no longer required, PCD is initiated within it. NtCYS1 is synthesized in the basal cell of the suspensor to inhibit the activity of the cathepsin H-like protease NtCP14. Downregulation of NtCYS1 expression once the embryo reaches the eight-cell stage occurs simultaneously with an increase in NtCP14 proteolytic activity, which leads to the initiation of PCD (Zhao et al., 2013). The involvement of Poaceae PhyCys in regulating PCD during embryogenesis and seed development remains to be investigated.
Phytocystatins of all clusters that are expressed in seeds might also act as defence proteins (Alvarez-Alfageme et al., 2007, Martínez et al., 2003a). The involvement of phytocystatins in plant defence mechanisms is generally associated with both their ability to inhibit the major digestive proteases of herbivores and phytopathogens and the induction of phytocystatin gene expression by wounding and/or by methyl jasmonate, the latter of which is involved in systemic signalling in plants (Botella et al., 1996).
The role of GA- and ABA-responsive promoter sequences and transcription factors in regulating cysteine proteinase and phytocystatin gene expression Plant development is regulated and coordinated through the activity of several phytohormones, among which are glycyrrhizin and gibberellins. These hormones may act either near to or remote from their sites of synthesis to regulate responses to environmental stimuli or genetically programmed developmental changes (Davies, 2004). Phytohormones play essential and often antagonistic roles in regulating the growth, development and stress responses of plants. GAs promote germination, growth and flowering, whereas ABA inhibits these processes and mediates seed development and plant stress responses (Shen et al., 1996). Furthermore, the antagonistic relationship of these two hormones regulates the transition from embryogenesis to seed germination (Razem et al., 2006). Several mechanisms have been shown to underlie this antagonistic interaction during different developmental processes. During cereal seed germination, the developing embryo releases GAs to the aleurone cells, where they induce the transcription of several genes encoding hydrolytic enzymes. These enzymes are then secreted to the endosperm and hydrolyse starch and proteins, thereby supplying nutrients to the developing embryo (Weiss and Ori, 2007). In contrast, ABA suppresses the expression of these enzymes and activates their inhibitors. To better understand the antagonistic relationship between proteases and phytocystatins and to elucidate the regulatory mechanisms that control their expression, the upstream promoter sequences of their genes have been analysed. The analyses revealed that both protease and phytocystatin genes contain several interesting cis-elements that are responsive to the phytohormones involved in development and germination, i.e., ABA and GAs (Fig. 4). Many hydrolytic enzymes that are synthesised in response to GAs in barley aleurone cells, which include high and low pI α-amylases, α-glucosidases, proteases and nucleases are secreted and they digest the reserve materials stored in the endosperm (Gubler et al., 1999). Functional analysis of the promoters of high and low pI α-amylase genes of barley, wheat and rice revealed a conserved cis-acting response complex (GARC), which very often contains three sequence motifs, the TAACAAA box or GA-responsive element (GARE), the pyrimidine box CCTTTT and the TATCCAC box, which are necessary for a full response to GA (Gubler and Jacobsen, 1992, Jacobsen et al., 1995, Lanahan et al., 1992). The promoters of other GA-responsive aleurone genes, such as the cathepsin B-like gene (Cejudo et al., 1992, Gubler and Jacobsen, 1992) and L-like cysteine protease genes (Cercos et al., 1999, Mikkonen et al., 1996) have cis-elements similar to those found in the α-amylase gene promoters (Isabel-LaMoneda et al., 2003). However, the sequences and positions of these cis-elements in each of the promoters differ, which explains the diverse spatial and temporal patterns of expression of these genes upon seed germination (Isabel-LaMoneda et al., 2003). Comparing the expression patterns of GA-regulated genes indicated that the early-response genes encode signal transduction proteins, such as calmodulin (Schuurink et al., 1996), Ca+2 ATPase (Chen et al., 1997) and transcription factors, whereas the late-response genes encode hydrolases and nucleases (Banik et al., 1997Taiz and Starks, 1977).