Predominant Beta Subunit Isoform of the Na + , K + ‐ATPase in Photoreceptor Neurons Undergoes S‐Palmitoylation

Many proteins undergo post‐translational lipid modifications, which are often essential to aid in subcellular trafficking, protein‐protein interactions, and membrane association. S‐palmitoylation is a unique post‐translational lipid modification which involves reversible attachment of the 16‐ carbon fatty acid palmitate to cysteine residues on proteins. In the retina, photoreceptor neurons are responsible for capturing photons of light and converting them into electrical signals, which are subsequently sent to the brain for visual processing. Many crucial phototransduction proteins in photoreceptors, including rhodopsin, phosphodiesterase‐6, and the transducin complex, have been shown to require post‐translational lipidation for proper function. To better understand the importance of “palmitoyl” lipid modifications in retinal photoreceptor cells, our lab employed acyl‐resin assisted capture (acyl‐RAC) followed by mass‐spectrometry analysis to isolate and identify proteins in retinal tissue which undergo S‐palmitoylation. From the mass‐spectrometry data pool, our lab chose several candidate proteins to study in more detail, including the β2‐ subunit of the Na + , K + ‐ATPase (ATP1B2). ATP1B2 has been shown to be the predominant β‐subunit isoform of the Na + , K + ‐ATPase in photoreceptor cells and is thought to be involved in maturation and trafficking of its catalytic α‐subunit counterpart. Additionally, ATP1B2 has been shown to be required by the retinal‐specific protein retinoschisin‐1 (RS1) for its localization and association with photoreceptor and bipolar cell membranes. Although it is known that the Na + , K + ‐ATPase regulates the photocurrent in photoreceptors needed to send visual information to downstream neurons, the distinct role of ATP1B2 remains unknown. Acyl‐RAC followed by immunoblot served as a complementary approach to confirm that ATP1B2 undergoes palmitoylation, whereas its homolog, the β1 isoform, does not. After palmitoylation prediction software analysis, we utilized mammalian cell culture and in vivo subretinal injection/electroporation to demonstrate that ATP1B2 is palmitoylated on the 10 th amino acid. Additionally, click chemistry with the palmitate analog 17‐Octadecynoic acid, was used as an independent approach to validate ATP1B2 palmitoylation. Altogether, these data confirm that ATP1B2 is post‐translationally lipid modified by palmitoylation on the 10 th amino acid. However, the functional role of palmitoylation of ATP1B2 remains unknown and warrants further investigation. Support or Funding Information NIH 1RO1EY028959‐01, WVU bridge funding, and intramural RFDG funding award (SK) This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .