Determining The Role Of Melanopsin C-Tail In Deactivation And Trafficking

Melanopsin is a unique non-image forming visual pigment expressed in intrinsically photosensitive retinal ganglion cells in the vertebrate retina. These cells are involved in many non-image forming functions such as the photoentrainment of circadian rhythm and the pupillary light reflex. Melanopsin is deactivated through the phosphorylation of the C-tail followed by the binding of a β-arrestin molecule. β-arrestin contains a signal on its C-terminus that allows for internalization of G-protein coupled receptors (GPCRs) after inactivation. However, it is currently unknown whether melanopsin is internalized. Angiotensin II type 1A receptor (ATII1AR) and B2 adrenergic receptor (B2AR) are two GPCRs known to bind β-arrestin and undergo endocytosis. To study the role of the C-tail in melanopsin deactivation and trafficking, the C-tail of melanopsin is replaced with either ATII1AR or B2AR c-tail using cloning techniques. We then introduce our plasmids into Human Embryonic Kidney (HEK) cells to assess the localization and signaling of the constructs. Sequencing has confirmed that several chimeric constructs have successfully been made. Calcium imaging has confirmed that the Mel/B2AR chimeric constructs signals in a similar manner as melanopsin in the presence of light. These results will help determine the role of the melanopsin C-tail in its deactivation and trafficking.