Abstract
The sigma-1 receptor (Sig1R) is an endoplasmic reticulum chaperonin that is attracting tremendous interest as a potential anti-neurodegenerative target. While this membrane protein is known to reside in the inner nuclear envelope (NE) and influences transcription, apparent Sig1R presence in the nucleoplasm is often observed, seemingly contradicting its NE localization. We addressed this confounding issue by applying an antibody-free approach of electron microscopy (EM) to define Sig1R nuclear localization. We expressed APEX2 peroxidase fused to Sig1R-GFP in a Sig1R-null NSC34 neuronal cell line generated with CRISPR-Cas9. APEX2-catalyzed gold/silver precipitation markedly improved EM clarity and confirmed an apparent intra-nuclear presence of Sig1R. However, serial sectioning combined with APEX2-enhanced EM revealed that Sig1R actually resided in the nucleoplasmic reticulum (NR), a specialized nuclear compartment formed via NE invagination into the nucleoplasm. NR cross-sections also indicated Sig1R in ring-shaped NR membranes. Thus, this study distinguishes Sig1R in the NR which could otherwise appear localized in the nucleoplasm if detected with low-resolution methods. Our finding is important for uncovering potential Sig1R regulations in the nucleus.
Highlights
The sigma-1 receptor (Sig1R) had long been deemed an orphan receptor or mistaken as an opioid receptor subtype [1]
Serial sectioning combined with APEX2enhanced electron microscopy (EM) revealed that Sig1R resided in the nucleoplasmic reticulum (NR), a specialized nuclear compartment formed via nuclear envelope (NE) invagination into the nucleoplasm
APEX is a peroxidase originally engineered for use in high resolution electron microscopy (EM) [28]
Summary
The sigma-1 receptor (Sig1R) had long been deemed an orphan receptor or mistaken as an opioid receptor subtype [1]. Later studies revealed Sig1R is unique: 1) It has no homolog in mammalian genomes [2]; 2) its structure is like no other [3]; 3) it represents a new class of signaling modulator [4, 5] whose functions may not be apparent until challenged with stress [1, 6,7,8]. Sig1R mutations are linked to familial amyotrophic lateral sclerosis (ALS) [9, 10]. Likely due to the lack of homologs, the molecular mechanisms underlying Sig1R functions are poorly understood, impeding Sig1R-targeted therapeutic development
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