Exploring the role of the ATRX chromatin remodeling protein in mouse astrocytes

Miguel A. Pena-Ortiz, Western University

Abstract

Mutations in the ATRX gene cause forms of intellectual disability. As with other neurodevelopmental disorder genes, ATRX has largely been studied with a focus on neurons. Astrocytes, the most abundant glial cells, are now recognized as major players in synaptic transmission and cognition. However, the role of ATRX in astrocytes has not yet been investigated. We initially optimized a fast and reliable method to obtain fluorescent-tagged glial nuclei suitable for RNA sequencing and chromatin related techniques. In the second portion of the study, mice with inducible Atrx deletion in astrocytes (Atrx^f/y;GlastCre^ERor “ATRX aiKO”) were generated to understand the contribution of astrocytic ATRX in cognition. Analysis of the hippocampal transcriptome of control and mutant mice revealed potential disruption in astrocytic cytoskeletal dynamics regulation and suggested a mild inflammatory response. Moreover, genes involved in neuronal function were also altered, suggesting that loss of ATRX in astrocytes influences neighbouring neurons at the transcriptional level. Evaluation of the astrocytic transcriptome confirmed changes in mRNA levels of genes involved in cytoskeletal regulation, immune activation, and metabolic function. Genome-wide chromatin accessibility profiles of ATRX-null astrocytes revealed a generally more open chromatin state. Chromatin accessibility was increased in genic regulatory regions like promoters and in binding sites for CTCF (a regulator of chromatin topology) and transcription factors implicated in interferon signaling. At the electrophysiological level, we showed that loss of ATRX in astrocytes leads to hyperexcitability and decreased capacitance of hippocampal CA1 neurons but no changes in dendrite number, length, and branching and only minor alterations in spine shape. Finally, behaviour testing of mutant mice demonstrated that loss of ATRX in astrocytes does not affect learning in the animals and disrupts specific types of long-term spatial and recognition memory. Together this study identifies important neuronal and cognitive dysfunction caused upon disruption of ATRX-mediated chromatin topology in astrocytes. This work also emphasizes the contributions of glial cells and particularly astrocytes to disorders like ATR-X syndrome.