The essentiality of long chain n-3 fatty acids in relation to development and function of the brain and retina.

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Analysis of human genetic linkage

Lipid–protein interactions in biological membranes: a structural perspective

The Na(+)-K(+)-ATPase, or sodium pump, is the membrane-bound enzyme that maintains the Na(+) and K(+) gradients across the plasma membrane of animal cells. Because of its importance in many basic and specialized cellular functions, this enzyme must be able to adapt to changing cellular and physiological stimuli. This review presents an overview of the many mechanisms in place to regulate sodium pump activity in a tissue-specific manner. These mechanisms include regulation by substrates, membrane-associated components such as cytoskeletal elements and the gamma-subunit, and circulating endogenous inhibitors as well as a variety of hormones, including corticosteroids, peptide hormones, and catecholamines. In addition, the review considers the effects of a range of specific intracellular signaling pathways involved in the regulation of pump activity and subcellular distribution, with particular consideration given to the effects of protein kinases and phosphatases.

Mechanisms of sodium pump regulation

Disparate biological processes involve fusion of two membranes into one and fission of one membrane into two. To formulate the possible job description for the proteins that mediate remodeling of biological membranes, we analyze the energy price of disruption and bending of membrane lipid bilayers at the different stages of bilayer fusion. The phenomenology and the pathways of the well-characterized reactions of biological remodeling, such as fusion mediated by influenza hemagglutinin, are compared with those studied for protein-free bilayers. We briefly consider some proteins involved in fusion and fission, and the dependence of remodeling on the lipid composition of the membranes. The specific hypothetical mechanisms by which the proteins can lower the energy price of the bilayer rearrangement are discussed in light of the experimental data and the requirements imposed by the elastic properties of the bilayer.

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Protein-lipid interplay in fusion and fission of biological membranes.

The (Na+,K+)-ATPase ATP hydrolyzing activity from rabbit kidney medulla basolateral membrane vesicles was studied as a function of the cholesterol content of the basolateral membranes The cholesterol content of the membranes was modified by incubation with phospholipid vesicles When the cholesterol content was increased above that found in the native membrane, the (Na+,K+)-ATPase ATP hydrolyzing activity was inhibited When the cholesterol content was decreased from that found in the native membranes, the (Na+,K+)-ATPase ATP hydrolyzing activity was inhibited Analogous effects were found with the K+-activated phosphatase activity of the same membrane vesicles Therefore, at low cholesterol contents, cholesterol was stimulatory, and at high cholesterol contents, cholesterol was inhibitory The structural specificity of this effect was tested by introducing lanosterol and ergosterol as 50% of the membrane sterol Ergosterol was the least effective at supporting (Na+,K+)-ATPase ATP hydrolyzing activity, while lanosterol was more effective, but still not as effective as cholesterol

Effects of cholesterol on sodium-potassium ATPase ATP hydrolyzing activity in bovine kidney

PURPOSE Gene therapy for retinal degeneration requires well-defined promoters that drive expression in rod and cone photoreceptors This study was undertaken to develop short, active derivatives of the human rhodopsin kinase (RK) gene promoter for targeting transgene expression in rods and cones RK, also known as G protein-coupled receptor kinase 1 (GRK1), is a component of the light adaptation pathway expressed in rods and conesMETHODS Human RK (hRK) promoter and its concatemers or derivatives extending into the conserved 5' untranslated region (5'-UTR) were assayed for promoter activity in WERI retinoblastoma or Crx/Sp1-supplemented HEK-293 cells The derivative displaying the highest activity was linked to a GFP reporter and packaged in a pseudotyped adenoassociated viral vector (AAV2/5) The AAV vector was tested in vivo by sub-retinal injections in wild-type mice, in the all-cone Nrl(-/-) mice, and in the cone-rich diurnal Nile grass rat (Arvicanthis niloticus) Control eyes received a similar AAV2/5 vector carrying a mouse rod opsin (mOps) promoter-controlled GFP reporterRESULTS The hRK promoter with the full 5' untranslated sequence (-112 to +180) was the most active in cell culture Delivered by the AAV2/5 vector, RK promoter drove GFP expression specifically in photoreceptors In rods, hRK promoter-mediated expression was as efficient as, but appeared more uniform than, mOps promoter-mediated expression In cones, the hRK promoter drove expression, whereas the mOps promoter did notCONCLUSIONS The hRK promoter is active and specific for rod and cone photoreceptors Because of its small size and proven activity in cones, it is a promoter of choice for somatic gene transfer and gene therapy targeting rods and cones

AAV-mediated expression targeting of rod and cone photoreceptors with a human rhodopsin kinase promoter

Rhodopsin-cholesterol interactions in bovine rod outer segment disk membranes

The ability of lysophosphatidylcholine to inhibit membrane fusion at subsolubilizing concentrations (between 1 and 9 mol % with respect to the membrane lipids) was examined. Fusion between N-methyldioleoylphosphatidylethanolamine (DOPE) large unilamellar vesicles (LUV) and fusion between Sendai virus and N-methyl-DOPE LUV were measured. A contents mixing fusion assay was used for LUV fusion (ANTS/DPX), and a lipid mixing assay (octadecylrhodamine B) was used for the virus fusion experiments. Lysophosphatidylcholine was effective at inhibiting both LUV fusion and Sendai virus/LUV fusion. Lysophosphatidylcholine also inhibited leakage from N-methyl-DOPE LUV, 31P nuclear magnetic resonance data were obtained of N-methyl-DOPE in the presence of lysophosphatidylcholine. Lysophosphatidylcholine stabilized the lamellar phase and reduced the incidence of nonlamellar structures at all temperatures. The destabilization of nonlamellar structures with a negative radius of curvature may be a mechanism for inhibition of fusion by lysophosphatidylcholine in these systems.

Joyce E. Young

Papers

Effects of cholesterol on sodium-potassium ATPase ATP hydrolyzing activity in bovine kidney

AAV-mediated expression targeting of rod and cone photoreceptors with a human rhodopsin kinase promoter

Rhodopsin-cholesterol interactions in bovine rod outer segment disk membranes

Inhibition of membrane fusion by lysophosphatidylcholine.

Cholesterol dynamics in membranes.