Abstract
Purpose :
Protein kinase C-alpha (PKCα) is a potent regulator of cell signaling cascades that plays a role in the modulation of the retinal rod bipolar cell (RBC) light response. Trophoblast glycoprotein (TPBG) is a leucine-rich repeat protein that is localized to the dendrites and synaptic terminals RBCs, but its function in the retina has not yet been examined. Using a phosphoproteomics approach, we identified TPBG as a PKCα-dependent phosphoprotein in RBCs. TPBG is phosphorylated on two serine residues within a PDZ-interacting motif in the C-terminal, cytoplasmic domain. We sought to identify a visual phenotype of antibody-mediated block of the TPBG C-terminal and N-terminal domains in vivo.
Methods :
WT and PKCα-KO mouse eyes were injected intravitreally with TPBG antibodies coupled to the cell penetrating peptide, pep-1. The contralateral eyes were injected with either pep-1 alone, or pep-1 coupled to heat inactivated antibody as intra-animal controls. The mice were allowed to recover overnight in the dark, and scotopic and photopic electroretinograms (ERGs) were recorded simultaneously from both eyes.
Results :
An antibody against the PDZ-interacting motif on the C-terminus of TPBG significantly attenuated the scotopic ERG b-wave in both WT and PKCα-KO mice when compared to controls. The effect was greater in PKCα-KO than WT mice for dim intensities.The N-terminal antibody had no effect on the scotopic b-wave in either group, and there were no changes under photopic conditions for either antibody. Following the ERGs, some retinas were removed and processed for immunfluorescence microscopy, which confirmed the accumulation of the TPBG antibody in RBCs.
Conclusions :
Our results suggest that interactions between TPBG and PDZ domain-containing proteins may be involved in modulation of the RBC light response. Evidence from other tissues suggests that TPBG may be involved in actin polymerization, filopodia formation, dendritic arborization, and synaptogenesis. Thus, TPBG may interact with proteins in RBC dendrites and synaptic terminals in a PKCα-dependent manner to shape RBC dendritic and synaptic morphology and to modulate the RBC light response.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.