Suppression of Thermotropic Lipid Clustering in Tetrahymena Nuclear Membranes upon Ca²⁺/Mg²⁺ Induced Membrane Contraction

Nuclear membranes surrounding the macronuclei isolated from Tetrahymena cells grown at either 28 C or 18 C are induced to expand or contract laterally by Ca 2+ /Mg 2+ (3/2) in final concentrations of 1 mM or 5 mM, respectively. Only expanded nuclear membranes are temperature‐responsive as revealed by freeze‐etch electron microscopy and electron spin resonance using 5‐doxylstearic acid as a spin label. In expanded nuclear membranes from growth at 28°C, electron spin resonance detects, upon lowering the temperature, a biphasic decrease of the lipid fluidity in terms of 2 T II with a discontinuity at ∼ 17°C. In this temperature range, freeze‐etch electron microscopy reveals an increase in the number of nuclear membrane fracture faces with smooth areas. We ascribe this response to a clustering of ordered lipid domains within a broad thermotropic fluid→ordered lipid phase separation [Wunderlich et al. (1978) Biochemistry, 17 , 2005–2010]. Expanded nuclear membranes from growth at 18°C shift their lipid clustering temperature down to ∼ 12°C. Their lipid fluidity (2 T II ) is increased in comparison with the nuclear membranes from growth at 28°C. This is probably due to an increased fatty acid unsaturation of the polar lipids. Thin‐layer‐chromatography reveals the following major polar lipids in the nuclei from growth at 18°C (28°C): 33.9%(31.4%) phosphatidylethanolamine, 14.2%(13.4%) phosphatidylethylamine, 22.5%(18.0%) unidentified phospholipid, 21.4%(19.1%) phosphatidylcholine, and 6.1%(10.4%) ceramide‐phosphonoethylamine. The major fatty acids are mainly even‐numbered ranging between C 12 and C 18 as analyzed by gas chromatography. The 18°‐nuclei (28°‐nuclei) contain 67.1% (60.4%), 74.5%(68.4%), and 32.3%(43.5%) unsaturated fatty acids in their total lipids, polar lipids, and neutral lipids, respectively. The individual phospholipids, except phosphatidylcholine and ceramide‐phosphonoethylamine, are more unsaturated in the nuclei from growth at 18°C. Contracted nuclear membranes from growth at 28°C and 18°C suppress thermotropic lipid clustering. They reveal an apparent higher lipid fluidity and percentage of fracture faces with smooth areas than the corresponding expanded nuclear membranes at 28°C and 18°C, respectively. The lipid clustering suppression is interpreted as being due to a change in lipid packing which in turn is induced by a Ca 2+ /Mg 2+ ‐induced contraction of the Ca 2+ /Mg 2+ ‐sensitive nucleoskeleton proteins associated with nuclear membranes.