Abstract
Response Gene to Complement 32 (RGC-32) is an important mediator of the TGF-β signaling pathway, and an increasing amount of evidence implicates this protein in regulating astrocyte biology. We showed recently that spinal cord astrocytes in mice lacking RGC-32 display an immature phenotype reminiscent of progenitors and radial glia, with an overall elongated morphology, increased proliferative capacity, and increased expression of progenitor markers when compared to their wild-type (WT) counterparts that make them incapable of undergoing reactive changes during the acute phase of experimental autoimmune encephalomyelitis (EAE). Here, in order to decipher the molecular networks underlying RGC-32’s ability to regulate astrocytic maturation and reactivity, we performed next-generation sequencing of RNA from WT and RGC-32 knockout (KO) neonatal mouse brain astrocytes, either unstimulated or stimulated with the pleiotropic cytokine TGF-β. Pathway enrichment analysis showed that RGC-32 is critical for the TGF-β-induced up-regulation of transcripts encoding proteins involved in brain development and tissue remodeling, such as axonal guidance molecules, transcription factors, extracellular matrix (ECM)-related proteins, and proteoglycans. Our next-generation sequencing of RNA analysis also demonstrated that a lack of RGC-32 results in a significant induction of WD repeat and FYVE domain-containing protein 1 (Wdfy1) and stanniocalcin-1 (Stc1). Immunohistochemical analysis of spinal cords isolated from normal adult mice and mice with EAE at the peak of disease showed that RGC-32 is necessary for the in vivo expression of ephrin receptor type A7 in reactive astrocytes, and that the lack of RGC-32 results in a higher number of homeodomain-only protein homeobox (HOPX)+ and CD133+ radial glia cells. Collectively, these findings suggest that RGC-32 plays a major role in modulating the transcriptomic changes in astrocytes that ultimately lead to molecular programs involved in astrocytic differentiation and reactive changes during neuroinflammation.
Highlights
Astrocytes are important players in the cellular mechanisms of multiple sclerosis (MS) pathogenesis and its animal model, experimental autoimmune encephalomyelitis (EAE), both of which are chronic, demyelinating disorders of the central nervous system (CNS) [1]
We previously showed that Response Gene to Complement 32 (RGC-32) is an essential component of the signaling pathways downstream of TGF-b that are involved in the synthesis of extracellular matrix (ECM) constituents, such as fibronectin and collagens type I, IV, and V, because it physically interacts with SMAD3 and regulates its nuclear translocation [17]
We found that in RGC-32 KO astrocytes, the transcriptomic programs associated with brain development and neurogenesis are significantly affected, and the transcription of genes encoding axonal guidance molecules (AGM) are impaired, as are genes involved in astrocyte development
Summary
Astrocytes are important players in the cellular mechanisms of multiple sclerosis (MS) pathogenesis and its animal model, experimental autoimmune encephalomyelitis (EAE), both of which are chronic, demyelinating disorders of the central nervous system (CNS) [1]. Our results have shown that in the absence of RGC-32, spinal cord astrocytes are unable to acquire a reactive phenotype during acute EAE, remaining instead in a progenitor state reminiscent of radial glia These radial glia-like cells from RGC-32 knockout (KO) mice are characterized by an elongated bipolar shape and an increased expression of progenitor markers such as vimentin and fatty acidic binding protein (FABP7) [19]. Our experiments have demonstrated that RGC-32 regulates the nuclear translocation of signal transducer and activator of transcription 3 (STAT3), a transcription factor critical for astrogliogenesis and reactive astrocytosis [19] Based on these observations, we have concluded that RGC-32 is necessary for astrocytes to reach maturity and to undergo reactive changes at the peak of EAE, and a lack of RGC-32 confers a protective phenotype against EAE in part by suppressing the development of a pathogenic astrocytic phenotype. These results suggest that RGC32 is an important gene that orchestrates the transcriptomic changes undergone by astrocytes as they mature and adopt a reactive phenotype during EAE
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
