Phospholipid Acyl Chain and Phospholipase Dynamics during Cold Acclimation of Winter Wheat

on the phospholipid profiles. It has been suggested that alterations in concentration of specific phospholipids Phospholipid (PL) composition is known to change in plants explay a role in cold acclimation. Horvath et al. (1979) posed to cold temperature. The dynamics of PL acyl chain pairs and genes encoding phospholipase enzymes were studied in winter wheat observed an inverse relationship between survival and (Triticum aestivum L.) during cold acclimation. Mass spectrometry the loss of phosphatidylcholine by conversion to the corwas used to characterize PL dynamics, and quantitative real-time responding phosphatidic acid in field-grown wheat PCR was used to characterize phospholipase gene mRNA transcript plants. A similar conversion of phosphatidylcholine to dynamics during cold acclimation. The proportion of PLs with misphosphatidic acid was reported by De La Roche and matched acyl chains decreased concomitantly with an increase in total Andrews (1973) in plants grown at 2 C and morphologiPLs during the first week of cold exposure. Proportions of mismatched cally equivalent plants grown at 24 C, again suggesting acyl chains then increased, while total PLs varied little. Numbers of these phospholipid dynamics are a function of plant phemRNA transcripts of phospholipase (PL)D, PLC, and PLA2 increased nology rather than cold acclimation. in response to cold and remained at elevated levels throughout a 4-wk The proportion of unsaturated fatty acids, particularly period. Lysophosphatidylcholine (LPC) increased as much as 14-fold over the 5-wk period and increased significantly less in a less cold linolenic acid (18 carbons, three double bonds [18:3]), tolerant cultivar than more tolerant cultivars. It appeared that newly increased during cold acclimation at the expense of less synthesized PLs with equal-length acyl chains form a part of the initial saturated forms (De La Roche et al., 1972, 1975; Skocresponse to cold temperature; they are then modified to contain nearzowski et al., 1994; Sopin and Trunova, 1991; Willemot initial levels of mismatched acyl chains during acclimation. LPC is a and Pelletier, 1980; Willemot et al., 1977a, 1977b). Howhighly active signal molecule and PLA2, PLC, and PLD are involved ever, it has been demonstrated that the accumulation in generation of phospholipid-based signaling molecules; hence, it of linolenic acid per se is not required for the developappeared PL signaling is involved in initial and continuing responses ment of freezing tolerance (De La Roche, 1979). Szalai to cold temperature. et al. (2001) studied winter wheat, spring wheat, and chromosome substitution lines incorporating winter wheat chromosome 5A, 5D, or 7A into the spring wheat C in phospholipids in wheat plants respondbackground and concluded that the overall level of acyl ing to cold temperature have been investigated chain fatty acid unsaturation was not related to cold many times from a structural viewpoint. It repeatedly tolerance. However, they reported that the rate of loss of has been shown that the total phospholipid content intrans3–hexadecenoic acid, presumably because of its creased during cold acclimation of wheat plants (De La conversion to other fatty acids, was strongly correlated Roche et al., 1972; De Silva et al., 1975; Horvath et al., with cold tolerance. It also has been shown that the 1980; Izzo et al., 1984; Willemot and Pelletier, 1980) and phospholipid composition of chloroplast thylakoid memcultured cells (Sopin and Gavrilova, 1987). Horvath et al. branes changes during cold exposure, but the changes (1980) proposed a predictive regression equation relatwere not related to cold tolerance (Vigh et al., 1985). ing increased leaf phospholipid accumulation and acEach of these studies examined the dynamics of the quired cold tolerance. Phospholipid de novo synthesis phospholipids defined by the head group and/or the is not required for cold hardening (Willemot, 1975), and fatty acid composition of the acyl side chains. However, initial increases in phospholipid content were followed we are aware of only one recent study where the compoby decreases after 3 wk (spring wheat) or 5 wk (winter sition of the acyl side chains was considered as a unit. wheat) of cold exposure (De Silva et al., 1975). Very Using electrospray ionizing mass spectrometry to analittle difference was found in phospholipid concentralyze phospholipids extracted from Arabidopsis, Welti tions from plants grown at 2 C compared with plants et al. (2002) characterized the acyl side chains as a comgrown to a morphologically equivalent state, but over bined group. For example, a 36:2 value would represent a shorter time, at 24 C (De La Roche and Andrews, a particular pair of acyl chains containing a total of 36 1973). Cold temperature appeared to have little effect carbons with two double bonds, probably indicating a pair of 18:1 acyl chains. The composition of the acyl D.Z. Skinner, B.S. Bellinger, and K. Garland-Campbell, USDA-ARS, chain pair of a phospholipid may play a crucial role in the 209 Johnson Hall, Washington State Univ., Pullman, WA 99164; S. Halls and W.F. Siems, Dep. of Chemistry, Fulmer Hall, Washington behavior of the membrane or components embedded in State Univ., Pullman, WA 99164; K.-H. Baek, Dep. of Crop and Soil the membrane. For example, the temperature optimum Sciences, 201 Johnson Hall, Washington State Univ., Pullman, WA of membrane-bound ATPase varied significantly depend99164. Received 10 Dec. 2004. *Corresponding author ([email protected]). ing on the acyl chain composition of the surrounding Published in Crop Sci. 45:1858–1867 (2005). Crop Physiology & Metabolism doi:10.2135/cropsci2004.0721 Abbreviations: ESI-MS, electrospray ionizing mass spectrometry; LPC, lysophosphatidyl choline; PL, phospholipid; PLA, phospholi© Crop Science Society of America 677 S. Segoe Rd., Madison, WI 53711 USA pase A; PLC, phospholipase C; PLD, phospholipase D. 1858 Published online August 1, 2005