Mutational alteration of membrane phospholipid composition and voltage-sensitive ion channel function in paramecium.

Abstract

A behavioral mutant of Paramecium tetraurelia (baA) has been isolated that has an abnormal response when placed in solutions containing Ba2+. This mutant is shown here to have a dramatic alteration of the sphingolipid and phosphonolipid composition ofits ciliary membrane. This biochemical defect is present in independently isolated alleles at baA locus and segregates in crosses with the behavioral phenotype. Electrophysiologically, the mutation reduces significantly conductance of both voltage-sensitive Ca2+ channels and voltage-sensitive K+ channels. When the mutant is grown in sterol-supplemented medium, its behavior, electrophysiological properties, and lipid composition are hardly distinguishable from wild type grown under similar conditions. This mutant then, provides strong evidence that membrane lipids significantly influence the function of the membrane molecules responsible for the generation of action potentials.Many membrane enzymatic and transport functions have been shown to be modulated by the membrane lipid composition (1-3). The membrane proteins responsible for the generation of action potentials are similar to those with enzymatic activity in that their function is influenced by membrane lipids as inferred from indirect studies on the effects of temperature (4), enzyme digestions (5-9), and the application of agents (e.g., local anesthetics or pressure), which are expected to alter the physical properties of membrane lipid (10-13).The ciliated protozoan Paramecium tetraurelia is well suited to the study ofthe molecular basis ofmembrane excitability. The cilia of Paramecium are covered by a membrane that is continuous with the body surface membrane (14). Membrane depolarization causes these channels to open allowing an influx of Ca2+, which increases transiently the intracellular Ca2+ concentration, reversing the direction ofthe ciliary beat and swimming direction (15,16 (33), and hydrostatic pressure (34) have suggested that lipids also affect ion channel activity in Paramecium. In this report we now show that the functions ofvoltage-sensitive ion channels in the excitable membrane of Paramecium appear to be affected by a mutationally induced change in the phospholipid composition of the ciliary membrane. This mutant provides strong evidence that membrane lipids significantly influence the function of membrane proteins responsible for the generation of action potentials. METHODSCulture Conditions and Stocks. Cells were grown by routine methods (21, 35). H332PO4 was added to 0.5 /Ci/ml (1 Ci = 3.7 x 1010 becquerels) prior to addition of bacteria to the medium. In some cases stigmasterol (5 mg/liter) was also added to the medium prior to bacteria addition. The strains were: wild type, 51s; baA-i, d4-592; baA-2, d4-593; baA-3, d4-594; fna, d4-91; pwA, d4-94; baB, d5-599 (27).Cilia Isolation and Lipid Extraction. Cells in late logarithmic growth were harvested, washed extensively, and deciliated; the cilia were separated from bodies as described by Adoutte et al (25). Lipids were extracted...

Evolutionary change during experimental ocean acidification

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Ciliopathy proteins establish a bipartite signaling compartment in aC. elegansthermosensory neuron

“…This high-fluidity lipid composition, however, might be incompatible with CNG channel function. The functions of ciliary channels are known to require a membrane rich in sphingolipids, which are saturated fatty acids (Forte et al, 1981), and lipid bilayer composition is known to substantially affect the functions of various ion channels (Schmidt et al, 2006;daCosta and Baenziger, 2009). Therefore, one function of ciliary subcompartments might be to provide appropriate lipid environment for different signaling proteins.…”

Phosphoinositides Regulate Ciliary Protein Trafficking to Modulate Hedgehog Signaling

“…In Paramecia, the ciliary membrane is enriched in sphingolipids and a mutation that alters ciliary lipid composition affects ciliary channel activity, suggesting that the ciliary lipid composition is critical for its function (Kaneshiro et al 1984; Andrews and Nelson 1979; Forte et al 1981). Similarly, in Tetrahymena and Chlamydomonas certain lipids are enriched in their cilia or flagella (Kennedy and Thompson Jr 1970; Jonah and Erwin 1971; Smith et al 1970; Gealt et al 1981; Bloodgood et al 1985).…”