Date Published: March 28, 2017
Publisher: Public Library of Science
Author(s): Ling Pan, David A. Pasternak, Jin Xu, Mingming Xu, Zhigang Lu, Gavril W. Pasternak, Ying-Xian Pan, Tuck Wah Soong.
The sigma1 receptor acts as a chaperone at the endoplasmic reticulum, associates with multiple proteins in various cellular systems, and involves in a number of diseases, such as addiction, pain, cancer and psychiatric disorders. The sigma1 receptor is encoded by the single copy SIGMAR1 gene. The current study identifies five alternatively spliced variants of the mouse sigma1 receptor gene using a polymerase chain reaction cloning approach. All the splice variants are generated by exon skipping or alternative 3’ or 5’ splicing, producing the truncated sigma1 receptor. Similar alternative splicing has been observed in the human SIGMAR1 gene based on the molecular cloning or genome sequence prediction, suggesting conservation of alternative splicing of SIGMAR1 gene. Using quantitative polymerase chain reactions, we demonstrate differential expression of several splice variants in mouse tissues and brain regions. When expressed in HEK293 cells, all the splice variants fail to bind sigma ligands, implicating that each truncated region in these splice variants is important for ligand binding. However, co-immunoprecipitation (Co-IP) study in HEK293 cells co-transfected with tagged constructs reveals that all the splice variants maintain their ability to physically associate with a mu opioid receptor (mMOR-1), providing useful information to correlate the motifs/sequences necessary for their physical association. Furthermore, a competition Co-IP study showed that all the variants can disrupt in a dose-dependent manner the dimerization of the original sigma1 receptor with mMOR-1, suggesting a potential dominant negative function and providing significant insights into their function.
Sigma receptors were initially proposed following the pharmacological studies with the benzomorphan opiate (±)SKF-10047 . Although initially thought to be related to opioid receptors, the sigma receptor is now recognized as a distinct protein unrelated to any traditional classes of receptor, including opioid receptors [2, 3]. Two subtypes of sigma receptors, sigma1 and sigma2 receptors, have been proposed based on their binding selectivity profiles . Sigma receptors are expressed in almost all tissues [5, 6] and in a wide range of tumors [7–9].
Alternative pre-mRNA splicing is one of the major mechanisms creating transcriptomic and proteomic diversity. Over 90% of human genes undergo alternative splicing [35, 36]. The current study further reveals extensive alternative splicing of the mouse Sigmar1 gene. Similar alternative splicing has been observed in the human SIGMAR1 gene based on the molecular cloning or genome sequence prediction (Fig 7). The human Sigma R1A  is homologous to mSIG-1A. A human isoform 6 has a similar splicing pattern as the mouse mSIG-1B and mSIG-1C. The hSIG-1SR (isoform 13) has identical splicing as the mouse mSIG-1SR . Several other human SIGMAR1 variants are also derived from alternative 3’ or 5’ splicing. All these variants create a series of truncated sigma1 receptors at protein level.