Capítulo III De los Extranjeros
Artículo 34. Son ciudadanos de la República los varones y mujeres que, teniendo la calidad de
All enzymes involved in the AAA pathway have been biochemically characterized, and the corresponding genes have been identified in both microbes and plants (Maeda and Dudareva 2012). Microbial enzymes of the pathway have been extensively studied with genetic analyses that have clarified AAA pathway regulation. In contrast, only limited genetic studies have been performed in plant enzymes of the AAA pathway, and thus, the regulation of the AAA pathway remains poorly understood in plants (Maeda and Dudareva 2012). Although the regulation of the AAA pathway is not completely clear, recent advances have elucidated some possible transcriptional and post- transcriptional regulations.
Transcriptional regulation
It has been reported that the presence of many transcription factors co-regulate gene expression of the AAA pathway. This regulation is based in several transcription factors: ODORANT1 (ODO1), EMISSION OF BENZENOIDS II (EOBII) (Verdonk 2005) and EPF1, a C2H2-type zinc finger DNA-binding protein (Takatsuji et al. 1992) (Fig. GI.6).
ODO1 and EOBII are closely related; in fact, it has been proposed that EOBII activates ODO1 in Petunia hybrida (Van Moerkercke et al. 2011). The suppression of both transcription factors decreased the expression of CM, and EOBII suppression resulted in partial ODO1 downregulation (Spitzer-Rimon et al. 2010) (Fig. GI.6). In contrast, the overexpression of EOBII did not significantly alter the expression of ODO1 or CM (Spitzer-Rimon et al. 2010); therefore, additional factors are likely involved (Maeda and Dudareva 2012). ODO1 affected the expression of several enzymes in the AAA pathway; indeed,
DAHPS and EPSPS expression were affected by this transcription factor
(Verdonk 2005) (Fig. GI.6).
It was demonstrated that in Petunia hybrida, EPF1 exhibited a very similar activation pattern in expression compared with the EPSPS activation pattern (Takatsuji et al. 1992). This common pattern was proposed as evidence of specific regulation, in which EPF1 is a positive transcription factor of EPSPS (Takatsuji et al. 1992) (Fig. GI.6).
It was reported in Arabidopsis thaliana that other transcription factor members of the MYB family (MYB 51 and MYB 34) activated genes encoding
DAHPS (Bender and Fink 1998) and AS (Gigolashvili et al. 2007) and that AS
had complementary regulation with ORCA3 (another MYB) in Catharanthus
roseus (Van Der Fits and Memelink 2000) (Fig. GI.6).
Other transcription factors, such as NST1, NST3, and MYB 8, had a general effect on the transcription of all the enzymes in the pre-chorismate part of the AAA pathway (Maeda and Dudareva 2012).
Post‐transcriptional regulation
It has been proposed that a general feedback regulation is carried out by the final products of the AAA pathway, with Trp regulating DAHPS and AS, and Tyr and Phe regulating CM (Tzin and Galili 2010b; Galili et al. 2016), although in the case of DAHPS inhibition by Trp, feedback inhibition remains controversial. In fact, other studies have reported the stimulation of this enzyme when Trp levels increase (Fig. GI.6) (Pinto et al. 1988; Maeda and Dudareva 2012). In addition, it has been proposed that Trp stimulates CM enzyme activity (Maeda and Dudareva 2012; Galili et al. 2016), in which case this enzyme acts in the opposite direction as the other two aromatic amino acids (Fig. GI.6).
In the case of Tyr and Phe, it has been proposed that there is not only allosteric inhibition of CM but also inhibition of the final enzymes whose products are the other two AAAs – arogenate dehydrogenase (ADT) in the case of Phe, and arogenate dehydratase (ArDH) in the case of Tyr (Maeda and Dudareva 2012; Galili et al. 2016) (Fig. GI.6).
Currently, it has not been confirmed whether or not there is any kind of post-transcriptional regulation over EPSPS, although an additional control of transcriptional regulation has been proposed (Mazzucotelli et al. 2008; Délye 2012).
A general coordination of transcriptional and post-transcriptional regulation in some important amino acid pathways, including the AAA pathway, has been proposed (Less and Galili 2008). These authors proposed that the stimulation or repression produced by stress causes a reduction or increase in the level of amino acids. This provokes a concomitant elevation or reduction in the allosteric feedback inhibition (red and green arrows in Fig. GI.6), resulting
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in an acceleration or slowdown of the carbon flux into the pathway. Only when the levels of the biosynthetic enzymes are not sufficient to maintain flux through the pathway in response to a stress (e.g., glyphosate) would the expression of the genes also be stimulated (or repressed) (Less and Galili 2008).
Figure GI.6 Regulation of the AAA pathway with their main enzymes: D-arabino-heptulosonate 7-
phosphate synthase (DAHPS), dehydroquinate synthase (DHQS), 3-dehydroquinate dehydratase/shikimate dehydrogenase (DQSD), shikimate kinase (SK), 5-enolpyruvylshikimate 3- phosphate synthase (EPSPS), chorismate synthase (CS), anthranilate synthase (AS), chorismate mutase (CM) and two post-prephenate enzymes with a regulatory role:arogenate dehydratase ADT and arogenate dehydrogenase (ArDH). Arrows in purple: transcriptional regulation produced by transcriptional factors. Arrows in red, green and brown: Post-transcriptional regulation, being red for inhibition loop, green for enhanced loop, and brown for controversial behavior. In purple circles transcriptional factors: myelobastosis family (MYBs), octadecanoid-derivative responsive Catharanthus AP2-domain 3 (ORCA3), ODORANT1 (ODO1), EPIDERMAL PATTERNING FACTOR 1 (EPF1) and EMISSION OF BENZENOIDS II (EOBII) and in blue boxes enzymes of the pathway. (Based on Maeda and Dudareva 2012).
The stimulation of the genes is probably mediated by transcription factors (Maeda and Dudareva 2012) (purple arrows in Fig. GI.6). It has been proposed a general feedback regulation produced by the final products of the AAA pathway, the three aromatic amino acids (Trp over DAHPS and AS; and
Tyr and Phe over CM) (Tzin and Galili 2010b; Galili et al. 2016) although in the case of DAHPS inhibition by Trp, feedback inhibition is controversial. In fact, other studies reported the stimulation of this enzyme when Trp levels increase (Fig. GI.6) (Pinto et al. 1988; Maeda and Dudareva 2012). In addition, it has been proposed a stimulation of CM enzyme activity by Trp (Maeda and Dudareva 2012; Galili et al. 2016), that acts over this enzyme in the opposite direction comparing to the other two aromatic amino acids (Fig. GI.6).