HORT640 - Metabolic Plant Physiology
GABA accumulation under anaerobic stress
The accumulation of the amino acid 4-aminobutyrate (gamma-aminobutyrate) (GABA) is markedly stimulated under anaerobic conditions (Streeter and Thompson, 1972; Stewart and Larher, 1980; Aurisano et al, 1995ab; Ratcliffe, 1995).
In part this may be a consequence of cytoplasmic acidification, stimulating the activity of glutamate decarboxylase, which has an acid pH optimum (~ 5.8) (Patterson and Graham, 1987):
Glutamate decarboxylase [EC 126.96.36.199]
glutamate + H+ ---> GABA + CO2
Because this reaction is proton consuming, it could represent an adaptive response contributing to regulation of cytoplasmic pH (Patterson and Graham, 1987; Crawford et al, 1994; Ratcliffe, 1995).
Crawford et al (1994) have shown that weak acids causing cytoplasmic acidification also induce GABA accumulation, and they conclude that this response is consistent with a role for GABA synthesis in active pH regulation.
GABA can be transaminated with pyruvate to yield alanine (or with 2-oxoglutarate to yield glutamate), generating succinic semialdehyde, which can then be metabolized to succinate via the action of succinate semialdehyde dehydrogenase (Vandewalle and Olson, 1983; Patterson and Graham, 1987; Shelp et al, 1995).
The latter enzymes are mitochondrial (Hearl and Churchich, 1984; Breitkreuz and Shelp, 1995), and have alkaline pH optima (Patterson and Graham, 1987). GABA accumulation promoted by cytosolic acidification may result in part from inhibition of GABA-transaminases.
The conversion of glutamate to succinate via the action of glutamate decarboxylase, GABA transaminase [EC 188.8.131.52], and succinic semialdehyde dehydrogenase [EC 184.108.40.206 or 220.127.116.11] is known as the "GABA shunt" (Vandewalle and Olsson, 1983; Patterson and Graham, 1987; Breitkreuz and Shelp, 1995; Shelp et al, 1995) affording an alternative pathway for glutamate entry into the TCA (Krebs) cycle.
Although the glutamate decarboxylase pathway of GABA synthesis is thought to be the predominant pathway of GABA synthesis in plants, it should be noted that GABA can also be derived from putrescine (via gamma-aminobutyraldehyde) through the reactions catalyzed by diamine oxidase [EC 18.104.22.168] and gamma-aminobutyraldehyde dehydrogenase [EC 22.214.171.124] (Flores et al, 1989) (see Polyamines). The latter enzyme may be the same as betaine aldehyde dehydrogenase (BADH) [EC 126.96.36.199] involved in glycinebetaine synthesis (Trossat et al, 1997) (see Quaternary ammonium and tertiary sulfonium compounds).
GABA accumulation is induced in response to a sudden decrease in temperature (Wallace et al, 1984; Patterson and Graham, 1985), in response to heat shock (Mayer et al, 1990), mechanical manipulation (Wallace et al, 1984), and water stress (Rhodes et al, 1986). Heat shock is known to induce rapid, transient changes in cytoplasmic calcium (Gong et al, 1998).
Rapid GABA accumulation in response to wounding may play a role in plant defense against insects (Ramputh and Brown, 1996).
Glutamate decarboxylase is a cytosol localized enzyme (Breitkreuz and Shelp, 1995) and has recently been shown to be a calmodulin-binding protein that is Ca2+/calmodulin activated (Ling et al, 1994; Baum et al, 1996; Arazi et al, 1995; Snedden et al, 1995). For a recent review of glutamate decarboxylase regulation see: Ireland RJ, Lea PJ 1999 The enzymes of glutamine, glutamate, asparagine, and aspartate metabolism. In (BK Singh ed) "Plant Amino Acids: Biochemistry and Biotechnology", Marcel Dekker, NY, pp. 49-109.
Crawford et al (1994) note that reduced cytosolic pH values increase Ca2+ levels, and rapid and transient increases in Ca2+ levels are known to occur in response to mechanical stress and cold stress; conditions which elicit rapid GABA accumulation (Wallace et al, 1984). Mitochondria contribute to the anoxic Ca2+ signal in maize suspension-cultured cells (Subbaiah et al, 1998).
The Ca2+/calmodulin activation of glutamate decarboxylase provides a link between intermediary amino acid metabolism and perturbations of cytosolic Ca2+ (Ling et al, 1994; Baum et al, 1996; Arazi et al, 1995; Snedden et al, 1995) known to regulate a host of other metabolic activities (Allan and Trewavas, 1985; Bush, 1995; Sanders et al, 1999). For a recent review of GABA metabolism and functions see: Bown and Shelp (1997).
It has recently been shown that GABA stimulates ethylene production in sunflower, apparently by causing increases in ACC synthase [EC 188.8.131.52], mRNA accumulation, ACC levels, ACC oxidase mRNA levels and ACC oxidase activity, suggesting that GABA may play a role in signaling (Kathiresan et al, 1997).
The only other enzyme of glutamate metabolism known to be stimulated by Ca2+ in plants is glutamate dehydrogenase (GDH), a mitochondrial enzyme (Turano et al, 1997).
Glutamate dehydrogenase [EC 184.108.40.206]
NH3 + 2-oxoglutarate + NADH <---> glutamate + NAD+
(see also Ammonia assimilation and recycling)
In both maize and Arabidopsis, GDH is a hexameric enzyme whose subunits are encoded by two separate genes, gdh1 and gdh2 (Pryor, 1990; Magalhaes et al, 1990; Turano et al, 1997). In wildtype plants this results in 7 isoenzymes whose relative abundance is largely determined by the relative abundance of mRNA transcript abundance of the two genes (Turano et al, 1997). In gdh1 null mutants of maize a single GDH isozyme is detected, corresponding to the hexamer of the gdh2 gene product (Magalhaes et al, 1990).
A calcium binding domain has been identified in the B-subunit of GDH encoded by GDH2, but not in the A-subunit of GDH encoded by GDH1 in Arabidopsis, suggesting that the different isoforms of GDH composed of different combinations of subunits, may be differentially regulated by Ca2+ (Turano et al, 1997).
The Ca2+ stimulation of the aminating activity of the B-subunit of GDH encoded by GDH2 could possibly serve to provide the glutamate substrate required for GABA synthesis and accumulation in response to environmental stress-induced perturbations of intracellular Ca2+?
Allan EF, Trewavas AJ 1987 The role of calcium in metabolic control. In "The Biochemistry of Plants" (DD Davies ed), Vol. 12, Academic Press, New York, pp. 117-149.
Arazi T, Baum G, Sneddon WA, Shelp BJ, Fromm H 1995 Molecular and biochemical analysis of calmodulin interactions with the calmodulin-binding domain of plant glutamate decarboxylase. Plant Physiol. 108: 551-561.
Aurisano N, Bertani A, Reggiani R 1995a Anaerobic accumulation of 4-aminobutyrate in rice seedlings; causes and significance. Phytochem. 38: 1147-1150.
Aurisano N, Bertani A, Reggiani R 1995b Involvement of calcium and calmodulin in protein and amino acid metabolism in rice roots under anoxia. Plant & Cell Physiol. 36: 1525-1529.
Baum G, Lev-Yadun S, Fridmann Y, Arazi T, Katsnelson H, Zik M, Fromm H 1996 Calmodulin binding to glutamate decarboxylase is required for regulation of glutamate and GABA metabolism and normal development in plants. EMBO J. 15: 2988-2996.
Bown AW, Shelp BJ 1997 The metabolism and functions of gamma-aminobutyric acid. Plant Physiol. 115: 1-5.
Breitkreuz KE, Shelp BJ 1995 Subcellular compartmentation of the 4-aminobutyrate shunt in protoplasts from developing soybean cotyledons. Plant Physiol. 108: 99-103.
Bush DS 1995 Calcium regulation in plant cells and its role in signalling. Annu. Rev. Plant Physiol. Plant Mol. Biol. 46: 95-122.
Crawford LA, Bown AW, Breitkreuz KE, Guinel FC 1994 The synthesis of gamma-aminobutyric acid in response to treatments reducing cytosolic pH. Plant Physiol. 104: 865-871.
Flores HE, Protacio CM, Signs MW 1989 Primary and secondary metabolism of polyamines in plants. In "Plant Nitrogen Metabolism" (JE Poulton, JT Romeo, EE Conn eds), Rec. Adv. Phytochem., Vol 23, Plenum Press, New York, pp. 329-393.
Gong M, van der Luit AH, Knight MR, Trewavas AJ 1998 Heat-shock-induced changes in intracellular Ca2+ level in tobacco seedlings in relation to thermotolerance. Plant Physiol. 116: 429-437.
Hearl WG, Churchich JE 1984 Interactions between 4-aminobutyrate aminotransferase and succinic semialdehyde dehydrogenase, two mitochondrial enzymes. J. Biol. Chem. 259: 11459-11463.
Ireland RJ, Lea PJ 1999 The enzymes of glutamine, glutamate, asparagine, and aspartate metabolism. In (BK Singh ed) "Plant Amino Acids: Biochemistry and Biotechnology", Marcel Dekker, NY, pp. 49-109.
Kathiresan A, Tung P, Chinnappa CC, Reid DM 1997 Gamma-aminobutyric acid stimulates ethylene biosynthesis in sunflower. Plant Physiol. 115: 129-135.
Ling V, Snedden WA, Shelp BJ, Assman SM 1994 Analysis of a soluble calmodulin binding protein from fava bean roots: identification of glutamate decarboxylase as a calmodulin activated enzyme. Plant Cell 6: 1135-1143.
Magalhaes JR, Ju GC, Rich PJ, Rhodes D 1990 Kinetics of 15NH4+ assimilation in Zea mays: Preliminary studies with a glutamate dehydrogenase (GDH1) null mutant. Plant Physiol. 94: 647-656.
Mayer RR, Cherry JH, Rhodes D 1990 Effects of heat shock on amino acid metabolism of cowpea cells. Plant Physiol. 94: 796-810.
Patterson BD, Graham D 1987 Temperature and metabolism. In (DD Davies ed) "The Biochemistry of Plants", Vol 12, Academic Press, New York, pp. 153-199.
Pryor A 1990 A maize glutamate dehydrogenase null mutant is cold temperature sensitive. Maydica 35: 367-372.
Ratcliffe RG 1995 Metabolic aspects of the anoxic response in plant tissue. In "Environment and Plant Metabolism: Flexibility and Acclimation" (N Smirnoff ed), Bios Scientific, Oxford, pp. 111-127.
Ramputh A-I, Bown AW 1996 Rapid gama-aminobutyric acid synthesis and the inhibition of the growth and development of oblique-banded leaf-roller larvae. Plant Physiol. 111: 1349-1352.
Rhodes D, Handa S, Bressan RA 1986 Metabolic changes associated with adaptation of plant cells to water stress. Plant Physiol. 82: 890-903.
Sanders D, Brownlee C, Harper JF 1999 Communicating with calcium. Plant Cell 11: 691-706.
Shelp BJ, Walton CS, Snedden WA, Tuin LG, Oresnik IJ, Layzell DB 1995 Gaba shunt in developing soybean seed is associated with hypoxia. Plant Physiol. 94: 219-218.
Snedden WA, Arazi T, Fromm H, Shelp BJ 1995 Calcium/calmodulin activation of soybean glutamate decarboxylase. Plant Physiol. 108: 543-549.
Stewart GR, Larher F 1980 Accumulation of amino acids and related compounds in relation to environmental stress. In "The Biochemistry of Plants" (BJ Miflin ed), Vol. 5, Academic Press, New York, pp. 609-635.
Streeter JG, Thompson JF 1972 Anaerobic accumulation of gamma-aminobutyric acid and alanine in radish leaves (Raphanus sativus L.). Plant Physiol. 49: 572-578.
Subbaiah CC, Bush DS, Sachs MM 1998 Mitochondrial contribution to the anoxic Ca2+ signal in maize suspension-cultured cells. Plant Physiol. 118: 759-771.
Trossat C, Rathinasabapathi B, Hanson AD 1997 Transgenically expressed betaine aldehyde dehydrogenase efficiently catalyzes oxidation of dimethylsulfoniopropionaldehyde and omega-aminoaldehydes. Plant Physiol. 113: 1457-1461.
Turano FJ, Thakkar SS, Fang T, Weisemann JM 1997 Characterization and expression of NAD(H)-dependent glutamate dehydrogenase genes in Arabidopsis. Plant Physiol. 113: 1329-1341.
Vandewalle I, Olsson R 1983 The gamma-aminobutyric acid shunt in germinating Sinapsis alba seeds. Plant Sci. Lett. 31: 269-273.
Wallace W, Secor J, Schrader LE 1984 Rapid accumulation of gamma-aminobutyric acid and alanine in soybean leaves in response to an abrupt transfer to lower temperature, darkness, or mechanical manipulation. Plant Physiol. 75: 170-175.
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