Pannexin 2 protein expression is not restricted to the CNS.

Maxence Le Vasseur,John F Bechberger,Christian C Naus,Wun-Chey Sin,Jonathan Lelowski

doi:10.3389/fncel.2014.00392

Maxence Le Vasseur, John F Bechberger + Show 3 more

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https://doi.org/10.3389/fncel.2014.00392

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Abstract

Pannexins (Panx) are proteins homologous to the invertebrate gap junction proteins called innexins (Inx) and are traditionally described as transmembrane channels connecting the intracellular and extracellular compartments. Three distinct Panx paralogs (Panx1, Panx2 and Panx3) have been identified in vertebrates but previous reports on Panx expression and functionality focused primarily on Panx1 and Panx3 proteins. Several gene expression studies reported that Panx2 transcript is largely restricted to the central nervous system (CNS) hence suggesting that Panx2 might serve an important role in the CNS. However, the lack of suitable antibodies prevented the creation of a comprehensive map of Panx2 protein expression and Panx2 protein localization profile is currently mostly inferred from the distribution of its transcript. In this study, we characterized novel commercial monoclonal antibodies and surveyed Panx2 expression and distribution at the mRNA and protein level by real-time qPCR, Western blotting and immunofluorescence. Panx2 protein levels were readily detected in every tissue examined, even when transcriptional analysis predicted very low Panx2 protein expression. Furthermore, our results indicate that Panx2 transcriptional activity is a poor predictor of Panx2 protein abundance and does not correlate with Panx2 protein levels. Despite showing disproportionately high transcript levels, the CNS expressed less Panx2 protein than any other tissues analyzed. Additionally, we showed that Panx2 protein does not localize at the plasma membrane like other gap junction proteins but remains confined within cytoplasmic compartments. Overall, our results demonstrate that the endogenous expression of Panx2 protein is not restricted to the CNS and is more ubiquitous than initially predicted.

Highlights

Gap junction proteins are traditionally described as aqueous plasma membrane channels which allow rapid cell-to-cell communication by directly connecting the cytoplasm of adjacent cells
By performing real-time quantitative real-time PCR (qPCR) and www.frontiersin.org semi-quantitative Western blot analysis on a panel of mouse tissues, we showed that fluctuations in Panx2 mRNA abundance do not predict changes in Panx2 protein levels
We showed that Panx2 protein levels are surprisingly more abundant in non-neural tissues than in the central nervous system (CNS); an observation opposite to Panx2 transcriptional activity which is weak in non-neural tissues and largely predominant in the CNS

Summary

Introduction

Gap junction proteins are traditionally described as aqueous plasma membrane channels which allow rapid cell-to-cell communication by directly connecting the cytoplasm of adjacent cells. Connexins (Cxs) are the canonical gap junction proteins while gap junctions in invertebrates are formed exclusively by the evolutionarily unrelated innexin (Inx) family. In 2000, another small gene family named pannexin (Panx) was identified based on sequence homology with the Inx family and was found to be expressed alongside Cxs in chordates (Panchin et al, 2000). Sequence similarity between Inxs/Panxs and Cxs, both families share structural resemblance. Despite sharing structural resemblance with Cxs, the ability of Panx channels to form gap junctional coupling remains controversial. It is largely accepted that under physiological conditions, Panx channels primarily form non-junctional membrane channels controlling the exchange of ions and small molecules between the cytoplasm and extracellular space and do not significantly contribute to direct cell-to-cell gap junctional communication (Sosinsky et al, 2011)

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