Abstract
Purpose :
The signal recognition particle (SRP) is a ribonucleoprotein complex that recognizes the nascent peptide localization signals of ribosome-bound mRNAs targeted to the endoplasmic reticulum (ER). SRP are composed of 7 protein subunits and a structural long noncoding 7S RNA. We hypothesized that SRP and ER are highly abundant in photoreceptors (PR) and at PR inner segments (PIS), where integral membrane proteins associated with phototransduction are highly expressed to support constant production of segments.
Methods :
Deep RNA sequencing of total RNA from mouse retinas was used to evaluate 7S abundance and its distribution was visualized in mouse and rat retinal thin sections using in situ hybridization (ISH); ISH of rhodopsin (Rho) mRNA was used as a control. Immunofluorescence (IF) of retinal sections was used to localize SRP proteins (SRP14, SRP19 and SRP68). ER distribution was examined in retinal sections using a well-validated antibody to glucose-regulated protein 78 (GRP78). Retinal ganglion cells (RGC) were identified using an antibody to RNA binding protein with multiple splicing (RBPMS). RiboTag-isolation and sequencing of RGC-expressed mRNAs and bioinformatics were used to characterize genes highly expressed by mouse RGC.
Results :
7S was the most abundant RNA species in mouse retina, being 4.6-fold greater than Rho mRNA. As predicted, ISH demonstrated 7S RNA within PR soma and a very high density of 7S RNA at PIS, corresponded closely to RhomRNA. Surprisingly, 7S RNA was also abundant in the inner nuclear layer (INL) and ganglion cell layer (GCL). SRP protein content was relatively high at PIS, INL and GCL. Similarly, the ER marker GRP78 was relatively high at PIS, INL and GCL. GRP78 was highly co-localized with RBPMS. The mouse RGC transcriptome revealed that genes corresponding to integral-membrane and secreted proteins were highly expressed by RGC, and bioinformatic analyses of these provided novel insights into RGC function.
Conclusions :
Our recently published findings revealed that mouse RGC exhibit relatively high levels of general protein synthesis (Fort, PE, et al., 2022, JBC 298(6):101944). High levels of SRP and ER in RGC suggest that these cells are actively synthesizing relatively large amounts of ER-targeted integral-membrane and secreted proteins that contribute to their unique physiology.
This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.