Human tRNASec associates with HeLa membranes, cell lipid liposomes, and synthetic lipid bilayers

Abstract

We have shown previously that simple RNA structures bind pure phospholipid liposomes. However, binding of bona fide cellular RNAs under physiological ionic conditions is shown here for the first time. Human tRNASec contains a hydrophobic anticodon-loop modification: N6-isopentenyladenosine (i6A) adjacent to its anticodon. Using a highly specific double-probe hybridization assay, we show mature human tRNASec specifically retained in HeLa intermediate-density membranes. Further, isolated human tRNASec rebinds to liposomes from isolated HeLa membrane lipids, to a much greater extent than an unmodified tRNASec transcript. To better define this affinity, experiments with pure lipids show that liposomes forming rafts or including positively charged sphingosine, or particularly both together, exhibit increased tRNASec binding. Thus tRNASec residence on membranes is determined by several factors, such as hydrophobic modification (likely isopentenylation of tRNASec), lipid structure (particularly lipid rafts), or sphingosine at a physiological concentration in rafted membranes. From prior work, RNA structure and ionic conditions also appear important. tRNASec dissociation from HeLa liposomes implies a mean membrane residence of 7.6 min at 24°C (t1/2 = 5.3 min). Clearly RNA with a 5-carbon hydrophobic modification binds HeLa membranes, probably favoring raft domains containing specific lipids, for times sufficient to alter biological fates. Copyright © 2012 RNA Society.