Deletion of endoplasmic reticulum (ER) Ca²⁺ efflux channel inositol-1,4,5-trisphosphate receptor type 1 or ryanodine receptor 2 promotes ER retrotranslocation/proteostasis in cyclic nucleotide-gated channel-deficient mice

Abstract

Purpose : Previous studies showed that cone death in cyclic nucleotide-gated (CNG) channel-deficient mice involves endoplasmic reticulum (ER) stress-associated apoptosis and dysregulation of ER Ca²⁺ homeostasis. We also demonstrated that expression/activity of ER Ca²⁺ efflux channels inositol 1,4,5-trisphosphate receptor 1 (IP₃R1) and ryanodine receptor 2 (RyR2) were increased, and ER stress/cone death was reduced after deletion of the two channels. This work investigates whether deletion of ER Ca²⁺ efflux channels promotes ER retrotranslocation/proteostasis in CNG channel-deficient retinas.

Methods : Cnga3^-/-/Nrl^-/- and Cnga3^-/-/Nrl^-/- mice with cone-specific deletions of Itpr1 (encoding IP₃R1, Cnga3^-/-/Nrl^-/-/Itpr1^fl/fl/Hrgp^cre) or Ryr2 (encoding RyR2, Cnga3^-/-/Nrl^-/-/Ryr2^fl/fl/Hrgp^cre) mice were used. ER retrotranslocation/proteostasis were evaluated by examining ER retrotranslocation machinery proteins syvn 1 (E3 ubiquitin-protein ligase synoviolin 1), Sel1L (ERAD E3 ligase adaptor subunit), Herpud1 (homocysteine inducible ER protein with ubiquitin like domain 1), Derl-1 (degradation in ER protein 1), and proteasome subunits PA28α and PSMD11 using immunoblotting. Chemical chaperone tauroursodeoxycholic acid (TUDCA, 500 mg/kg, body weight) or vehicle was given to Cnga3^-/-/Nrl^-/- mice by subcutaneous injection every 3 days for 12 days, starting at P5. Retinas were collected at the end of the treatment for immunoblotting.

Results : Expression levels of syvn 1, Sel1L, and Herpud1 were unchanged in Cnga3^-/-/Nrl^-/- mice, compared with Nrl^-/- controls. However, deletion of IP₃R1 or RyR2 significantly increased their expression levels. Derl-1 expression level was increased in Cnga3^-/-/Nrl^-/- retinas, and deletion of IP₃R1 completely abolished the upregulation. Expression level of PA28α was not different between Cnga3^-/-/Nrl^-/- and Nrl^-/- retinas. Deletion of IP₃R1 or RyR2 significantly reduced its level. Similar findings were observed after TUDCA treatment.

Conclusions : Our results showed that ER retrotranslocation/proteostasis was increased/improved after deletion of ER Ca²⁺ efflux channels or after treatment with TUDCA. These findings suggest that promoting ER retrotranslocation/proteostasis may represent a promising strategy to reduce ER stress/cone death in CNG channel deficiency.

This is a 2021 ARVO Annual Meeting abstract.