Deficits Associated With Loss of STIM1 in Purkinje Neurons Including Motor Coordination Can Be Rescued by Loss of Septin 7.

Sreeja Kumari Dhanya,Gaiti Hasan

doi:10.3389/fcell.2021.794807

Abstract

Septins are cytoskeletal proteins that can assemble to form heteromeric filamentous complexes and regulate a range of membrane-associated cellular functions. SEPT7, a member of the septin family, functions as a negative regulator of the plasma membrane–localized store-operated Ca2+ entry (SOCE) channel, Orai in Drosophila neurons, and in human neural progenitor cells. Knockdown of STIM, a Ca2+ sensor in the endoplasmic reticulum (ER) and an integral component of SOCE, leads to flight deficits in Drosophila that can be rescued by partial loss of SEPT7 in neurons. Here, we tested the effect of reducing and removing SEPT7 in mouse Purkinje neurons (PNs) with the loss of STIM1. Mice with the complete knockout of STIM1 in PNs exhibit several age-dependent changes. These include altered gene expression in PNs, which correlates with increased synapses between climbing fiber (CF) axons and Purkinje neuron (PN) dendrites and a reduced ability to learn a motor coordination task. Removal of either one or two copies of the SEPT7 gene in STIM1 KO PNs restored the expression of a subset of genes, including several in the category of neuron projection development. Importantly, the rescue of gene expression in these animals is accompanied by normal CF-PN innervation and an improved ability to learn a motor coordination task in aging mice. Thus, the loss of SEPT7 in PNs further modulates cerebellar circuit function in STIM1 KO animals. Our findings are relevant in the context of identifying SEPT7 as a putative therapeutic target for various neurodegenerative diseases caused by reduced intracellular Ca2+ signaling.

Highlights

To understand if negative regulation of store-operated Ca2+ entry (SOCE) by SEPT7 is conserved among murine models and to investigate if motor deficits observed in STIM1 knockout (STIM1PKO) mice (Hartmann et al, 2014; Dhanya and Hasan, 2020) could be rescued by altering SEPT7 levels, we generated Purkinje neuron (PN)–specific compound STIM1-SEPT7 double knockout mice strains using the conditional Cre-lox system (Figures 1A,B and methods)
STIM1flox/flox; SEPT7flox/flox and STIM1flox/flox; SEPT7flox/+ mouse strains in the absence of Purkinje cell protein-2 (PCP2) (L7)-Crecre/+ were used as controls for homozygous and heterozygous SEPT7 knockout strains, respectively
We show that partial or complete deletion of SEPT7 from adult murine Purkinje neurons reverses multiple phenotypes associated with PN-specific loss of the endoplasmic reticulum (ER)-Ca2+ sensor and SOCE protein STIM1

Summary

Introduction

Septins (SEPT) constitute a family of filament-forming GTPases that can assemble into heterooligomeric complexes in cells and regulate various cellular functions such as cytokinesis (Kinoshita et al, 1997; Gladfelter et al, 2001; Kinoshita and Noda, 2001), cell polarity determination and maintenance (Drees et al, 2001; Faty et al, 2002; Irazoqui and Lew, 2004), microtubule and actin dynamics (Kinoshita et al, 1997; Finger, 2002; Surka et al, 2002; Nagata et al, 2003), membrane associations, cell movement (Finger et al, 2003), vesicle trafficking (Hsu et al, 1998), and exocytosis (Beites et al, 1999). Regulation of cellular Ca2+ signaling by septins was first demonstrated in an siRNA screen in HeLa cells that identified mammalian septins of the SEPT2 class (SEPT2/4/5) as positive regulators of STIM/Orai-mediated store-operated Ca2+ entry (SOCE) (Sharma et al, 2013) and is further supported by recent studies (de Souza et al, 2021). Heterozygotes of the Drosophila SEPT7 mutant could rescue cellular and systemic phenotypes associated with neuronal knockdown of the SOCE molecules STIM/Orai in adult Drosophila. Subsequent genetic and cellular studies support the idea that SEPT7 function in human stem cell–derived neural precursors and differentiated neurons is similar to Drosophila. In both human neuronal cells and Drosophila neurons, SEPT7 orthologs prevent spontaneous Ca2+ entry through the SOCE channel, Orai (Deb et al, 2016; Deb et al, 2020)

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Results

Conclusion