Chilling Tolerance in Arabidopsis Involves ALA1, a Member of a New Family of Putative Aminophospholipid Translocases

Abstract

The lipid composition of membranes is a key determinant for cold tolerance, and enzymes that modify membrane structure seem to be important for low-temperature acclimation. We have characterized ALA1 (for aminophospholipid ATPase1 ), a novel P-type ATPase in Arabidopsis that belongs to the gene family ALA1 to ALA11 . The deduced amino acid sequence of ALA1 is homologous with those of yeast DRS2 and bovine ATPase II, both of which are putative aminophospholipid translocases. ALA1 complements the deficiency in phosphatidylserine internalization into intact cells that is exhibited by the drs2 yeast mutant, and expression of ALA1 results in increased translocation of aminophospholipids in reconstituted yeast membrane vesicles. These lines of evidence suggest that ALA1 is involved in generating membrane lipid asymmetry and probably encodes an aminophospholipid translocase. ALA1 complements the cold sensitivity of the drs2 yeast mutant. Downregulation of ALA1 in Arabidopsis results in cold-affected plants that are much smaller than those of the wild type. These data suggest a link between regulation of transmembrane bilayer lipid asymmetry and the adaptation of plants to cold. INTRODUCTIONLow temperature is one of the major environmental factors limiting plant growth. Cold-sensitive plants are injured and stunted in growth when temperatures are well below those for normal growth but still above the freezing point (reviewed in Nishida and Murata, 1996;Pearce, 1999;Thomashow, 1999). Many tropical or subtropical plant species, including a large number of crops such as tomato, rice, cotton, cucumber, and maize, are susceptible to chilling injury, and substantial losses in productivity result from the inability of such crops to withstand cold stress. Normal functioning of integral membrane proteins such as transporters and receptor proteins depends on the fluidity of the membrane, which is strongly influenced at a given temperature by its lipid composition (Squier et al., 1988;Gasser et al., 1990; reviewed in Hazel, 1995).A major factor determining the fluidity of lipid membranes is the degree of unsaturation of membrane lipids. Thus, membranes with unsaturated acyl chains in phospholipids remain fluid at lower temperatures than do membranes with saturated lipids (Kates et al., 1984;Cevc, 1991;Cossins, 1994). One of the best-documented responses of plants to chilling stress is the increase in polyunsaturated acyl chains of membrane phospholipids, which allows membrane fluidity to be maintained (Hugly and Somerville, 1992;Nishida and Murata, 1996). This response indicates that after transfer to the cold, plants may increase the amount of unsaturated lipids by upregulating the activity of desaturase enzymes. Accordingly, expression in tobacco of a desaturase gene from a cyanobacterium results in increasing membrane lipid unsaturation in most membrane lipids concomitant with an increase in chilling tolerance (IshizakiNishizawa et al., 1996).The proportion of unsaturated fatty acids in the lipid acyl chains is particularly hig...

Glycerolipid transfer for the building of membranes in plant cells

“…In Arabidopsis, eleven genes belong to this family (ALA1 to ALA11) ( Table 2). ALA1 exhibits the strongest homology to DRS2 and its expression in drs2 yeast mutant restores cold resistance and internalisation of the plasma membrane PS [29]. However, ALA1 localization and function in planta are not characterized.…”

A phospholipid uptake system in the model plant Arabidopsis thaliana

“…For plants, [10][11][12][13][14][15] NBD-lipid-labelled seedlings were ground, resuspended in 100 ml ½MS media and extracted twice as above. The lipid containing chloroform/methanol phase was collected, dried and resuspended in a small volume of chloroform before separation by thin-layer chromatography using chloroform/ethanol/water/triethylamine (30/35/7/35, v/v/v/v; ref.…”

“…The catalytic a-subunit family consists of 12 members in A. thaliana, AminophosphoLipid ATPase1-12 (ALA1-12) 13,14 . A smaller family of b-subunits, ALA Interacting Subunits1-5 (ALIS1-5) 15 , assists in the maturation of a-subunits and their exit from the ER 16 .…”

“…Some of these plant ALA proteins have been characterized. ALA1, the first member of this family to be identified 14 , localizes to the plasma membrane; however, its biological function and the identity of its transported substrate remain unknown 17 . ALA2 flips phosphatidylserine (PS) across the lipid bilayer in the endosomal system 16 and ALA3 is a broad-specificity phospholipid flippase expressed in the Golgi apparatus, where it contributes to the formation of secretory vesicles 15 .…”

Assessment of Chilling Injury and Molecular Marker Analysis in Cucumber Cultivars (Cucumis sativus L.)

“…Low temperatures limit plant growth (Gomes et al, 2000) and can cause significant economic losses in vegetable crop species worldwide (Synder and Abreu, 2005). Temperatures near freezing disrupt plant growth in cold-sensitive plants causing stunting and injury (Thomashow, 1999).…”