Premium
Determining the role of melanopsin C‐tail in deactivation and trafficking using chimeric constructs (596.7)
Author(s) -
Ortiz Elelbin,
Cameron Evan,
Robinson Phyllis
Publication year - 2014
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.28.1_supplement.596.7
Subject(s) - melanopsin , microbiology and biotechnology , g protein coupled receptor , biology , hek 293 cells , intrinsically photosensitive retinal ganglion cells , receptor , chemistry , photopigment , retina , neuroscience , signal transduction , genetics , retinal ganglion cell
Melanopsin is a non‐image forming G protein‐coupled receptor expressed in intrisically photosensitive retinal ganglion cells in the vertebrate retina. These cells are involved in functions such as the photoentrainment of circadian rhythm and the pupillary light reflex. The deactivation of melanopsin occurs through the phosphorylation of the C‐tail followed by the binding of a β‐arrestin molecule. The binding of β‐arrestin allows for endocytosis of GPCRs after inactivation. However, preliminary data suggests that melanopsin is not internalized. We hypothesize that the C‐tail of melanopsin plays a key role in the prevention of internalization. Angiotensin II type 1A receptor (ATII1AR) and β‐2‐andrenergic receptor (B2AR) are GPCRs that bind β‐arrestin but differ in their trafficking via endocytic vesicles. In this study, the melanopsin C‐tail was replaced with either ATII1AR and B2AR C‐tail through cloning. The construction of the chimeras has been confirmed by DNA sequencing. Our next step is to introduce our plasmids into Human Embryonic Kidney (HEK) cells. These constructs will then be assayed for trafficking using immuno flow cytometry and immunohistochemistry. We hypothesize that the chimeric constructs will undergo endocytotic pathways similar to ATII1AR and B2AR. This would allow us to conclude that the melanopsin c‐tail is essential in maintaining melanopsin in the cell membrane. Grant Funding Source : Supported by HHMI, MARC U*STAR Program at UMBC, and a National Eye Institute grant to P.R.R.