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The plant SnRK1-type complex is orthologous to the yeast SNF1 and the mam- malian AMP-activated protein kinase (AMPK) complexes (Hardie, 2007; Polge and Thomas, 2007). They are metabolic sensors, activated primarily in response to low energy levels. These complexes alter expression of genes and also regulate activity of many proteins post-transcriptionally. The central importance of these complexes is highlighted by the fact that 400 of 1500 genes with altered expres- sion patterns inSaccharomyces cerevisiaein response to glucose starvation and the change from fermentative to oxidative metabolism are believed to be controlled by the SNF1 complex (DeRisi et al., 1997; Younget al., 2003); this is impressive given that there is believed to be approximately 5300-5400 genes in this organism (Mackiewiczet al., 2002).

Regulation of SnRK1-type kinases is by both upstream regulatory kinases and allosterically by altered energy indicators. Activity of SNF1/AMPK/SnRKs is in- hibited by high ATP/low AMP/ADP, glucose and phosphorylated hexokinases, T6P and, in some cases, some amino acids (see Crozet et al., 2014, for review). It has been reported, however, that SnRK1 activity is induced in response to su- crose or a high sucrose:glucose ratio (Purcellet al., 1998; Halford and Hey, 2009).

Upstream regulatory kinases phosphorylate a threonine residue in the catalytic subunit of the complex, a requirement for the highest level of activity. This in turn is related to energy levels; in yeast, while glucose levels are sufficient the threonine is predominantly non-phosphorylated, but when glucose levels are re- duced the upstream kinases phosphorylate the SNF1 complex (McCartney and Schmidt, 2001).

SnRK1-type complexes are heterotrimeric in nature; they contain a catalytic unit (often referred to as theαsubunit), a scaffold section (βsubunit) and a regulatory unit (γ subunit) (Polge and Thomas, 2007). The catalytic unit contains the thre- onine target of phosphorylation for complete activation of the complex, whereas the regulatory unit contains the allosteric inhibition sites. Allosteric interactions with the regulatory unit alter its association with the catalytic unit. The scaffold section, other than holding the complex together, is believed to play a role in sub- strate or target selection, containing a carbohydrate binding domain that closely resembles isoamylase domains and binding to such things as branched glycogen or starch. AMPK has been shown to exhibit inhibition not only due to AMP/ATP ratio and carbohydrates, but also specifically α(1-6) branched oligosaccharides which theβsubunit binds to (McBrideet al., 2009).

The SnRK1 complex alters transcription of genes via control of various tran- scription factors from a broad range of processes, including protein, amino acid, lipid, starch and sucrose degradation, photosynthesis, gluconeogenesis, stress responses, hormones, protein synthesis and histone/chromosome modification (Baena-Gonzalez et al., 2007). Such targets are sucrose synthase and α-amylase. Additionally, direct post-translational targets are nitrate reductase, HMG-CoA reductase, 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase, sucrose phos- phate synthase and trehalose-phosphate synthase, along with activation of ADP- glucose pyrophosphorylase, which is controlled by SnRK1 by modulating its re- dox state and gene expression (Polge and Thomas, 2007; Coelloet al., 2011).

Repression of SnRK1 in pea seeds using antisense RNA expression altered seed development, causing maturation defects such as growth retardation and abor- tion, altered cotyledon shape and symmetry, and precocious germination prior to desiccation, at which time germination was often not possible (Radchuket al., 2006). They also exhibited a reduced production of starch during development, increased GA biosynthetic gene expression and reduction of the ABA signalling gene abscisic acid insensitive 3 (ABI3); the phenotypes described are very sim-

ilar to those of ABA insensitivity. As mentioned in Chapters 3 and 5, many of the sugar-sensitive or insensitive mutants of plants are allelic to ABA related genes.

Cytokinin and ABA levels were reduced in SnRK1 repressed seeds compared to wild-type, and embryo growth was reduced in the transgenics (Radchuk et al., 2010). SnRK1 catalytic subunit expression was hormone and carbohydrate re- sponsive, with increased expression seen with addition of 2,4-dichlorophenoxyacetic acid (a synthetic auxin) and ABA or with reduced sucrose in experiments using isolated protoplasts. This is supported by work in tomatoes and wheat (Brad- ford et al., 2003; Coello et al., 2012). Several of the proteins responsive to ABA are known to be post-translationally modified in response to ABA presence, in- cluding phosphorylation; several of these have phosphorylated serine residues within motifs which match the known targets of SnRK1 (Bradford et al., 2003; Halfordet al., 2003).

Work in potatoes using both trehalose 6-phosphate synthase (TPS, which pro- duces T6P) and trehalose 6-phosphate phosphatase (TPP, which degrades T6P) overexpression in the tubers demonstrated the effect of under- and over-activating SnRK1 (Debast et al., 2011). Those with decreased SnRK1 activity (TPS lines) showed a reduced starch content, decreased ATP levels and increased metabolic activity. Conversely, increased SnRK1 activity (TPP) led to increased soluble car- bohydrates, hexose phosphates and ATP, unchanged starch levels but a reduced yield per plant, due to decreased tuber size; tuber numbers per plant were, how- ever, increased in these lines.

The TPP lines also sprouted earlier than the control lines, and TPS lines were sig- nificantly delayed in their sprouting rates (Debastet al., 2011). TPS lines showed a greatly reduced sprouting rate compared to wild-type with the addition of ei- ther GA or cytokinin, yet TPP lines sprouted far faster than controls with the same treatments; this indicates that increased SnRK1 activity leads to GA and cytokinin oversensitivity, yet reduced activity leads to insensitivity. ABA levels of the lines were also altered, with TPP increased SnRK1 activity lines showing a large reduction of ABA content and increased ABA 8’OH (which degrades ABA) expression. These results show that increased SnRK1 activity decreases poten- tial dormancy and induces precocious sprouting in potato. This is supported by similar work in rice and Arabidopsis (Luet al., 2007; Jossieret al., 2009).