Differential distribution of proteins and lipids in detergent-resistant and detergent-soluble domains in rod outer segment plasma membranes and disks.
Membrane heterogeneity plays a significant role in regulating signal transduction and other cellular activities. We examined the protein and lipid components associated with the detergent-resistant membrane (DRM) fractions from retinal rod outer segment (ROS) disk and plasma membrane-enriched preparations. Proteomics and correlative western blot analysis revealed the presence of alpha and beta subunits of the rod cGMP-gated ion channel and glucose transporter type 1, among other proteins. The glucose transporter was present exclusively in ROS plasma membrane (not disks) and was highly enriched in DRMs, as was the cGMP-gated channel beta-subunit. In contrast, the majority of rod opsin and ATP-binding cassette transporter A4 was localized to detergent-soluble domains in disks. As expected, the cholesterol : fatty acid mole ratio was higher in DRMs than in the corresponding parent membranes (disk and plasma membranes, respectively) and was also higher in disks compared to plasma membranes. Furthermore, the ratio of saturated : polyunsaturated fatty acids was also higher in DRMs compared to their respective parent membranes (disk and plasma membranes). These results confirm that DRMs prepared from both disks and plasma membranes are enriched in cholesterol and in saturated fatty acids compared to their parent membranes. The dominant fatty acids in DRMs were 16 : 0 and 18 : 0; 22 : 6n3 and 18 : 1 levels were threefold higher and twofold lower, respectively, in disk-derived DRMs compared to plasma membrane-derived DRMs. We estimate, based on fatty acid recovery that DRMs account for only approximately 8% of disks and approximately 12% of ROS plasma membrane.
Elliott
Michael H
MH
Department of Ophthalmology, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma 73104, USA.
Nash
Zack A
ZA
Takemori
Nobuaki
N
Fliesler
Steven J
SJ
McClellan
Mark E
ME
Naash
Muna I
MI
D000818:Animals; D002417:Cattle; D002462:Cell Membrane / Q000737:chemistry* / Q000187:drug effects*; D003902:Detergents / Q000494:pharmacology*; D008055:Lipids; D016253:Microscopy, Immunoelectron / Q000379:methods; D011506:Proteins / Q000187:drug effects / Q000378:metabolism*; D040901:Proteomics / Q000379:methods; D012160:Retina / Q000166:cytology; D012374:Rod Cell Outer Segment / Q000648:ultrastructure*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods
D003902:Detergents; D008055:Lipids; D011506:Proteins