Unbiased Expression Mapping Identifies a Link between the Complement and Cholinergic Systems in the Rat Central Nervous System

Abstract

The complement system is activated in a wide spectrum of CNS diseases and is suggested to play a role in degenerative phenomena such as elimination of synaptic terminals. Still, little is known of mechanisms regulating complement activation in the CNS. Loss of synaptic terminals in the spinal cord after an experimental nerve injury is increased in the inbred DA strain compared with the PVG strain and is associated with expression of the upstream complement components C1q and C3, in the absence of membrane attack complex activation and neutrophil infiltration. To further dissect pathways regulating complement expression, we performed genome-wide expression profiling and linkage analysis in a large F2(DA × PVG) intercross, which identified quantitative trait loci regulating expression of C1qa, C1qb, C3, and C9. Unlike C1qa, C1qb, and C9, which all displayed distinct coregulation with different cis-regulated C-type lectins, C3 was regulated in a coexpression network immediately downstream of butyrylcholinesterase. Butyrylcholinesterase hydrolyses acetylcholine, which exerts immunoregulatory effects partly through TNF-α pathways. Accordingly, increased C3, but not C1q, expression was demonstrated in rat and mouse glia following TNF-α stimulation, which was abrogated in a dose-dependent manner by acetylcholine. These findings demonstrate new pathways regulating CNS complement expression using unbiased mapping in an experimental in vivo system. A direct link between cholinergic activity and complement activation is supported by in vitro experiments. The identification of distinct pathways subjected to regulation by naturally occurring genetic variability is of relevance for the understanding of disease mechanisms in neurologic conditions characterized by neuronal injury and complement activation.

Complement Component C3 and Butyrylcholinesterase Activity Are Associated with Neurodegeneration and Clinical Disability in Multiple Sclerosis

“…The genetic link between local expression of BuChE and C3 after experimental nerve injury provided a rationale to determine BuChE activity in the clinical samples [ 19 , 34 ]. In addition, activity of AChE, the other main ACh hydrolyzing enzyme [ 35 ], was determined.…”

“…We have previously found that genetically determined susceptibility to neurodegeneration after a standardized nerve injury among inbred rat strains is associated with a more prominent local inflammatory response with elevated expression of the upstream complement proteins C1q and C3 [ 18 ]. By performing genome-wide expression profiling and linkage analysis in a large inbred rat strain intercross we found co-regulation of complement C3 immediately downstream of butyrylcholinesterase (BuChE), an enzyme that hydrolyses acetylcholine (ACh) [ 19 ]. BuChE has been proposed as a risk gene for sporadic Alzheimer´s disease (AD) [ 20 ] and we recently showed that such patients carrying a BuChE allele coding for a protein with reduced activity displays lower levels of glial activation markers [ 21 ].…”

From integrative genomics to systems genetics in the rat to link genotypes to phenotypes

“…Studies linking mRNA levels to the genome in a single tissue include Monti et al (2008), Petretto et al (2008), Pravenec et al (2008), Jirout et al (2010), Low et al (2013) and Thessen Hedreul et al (2013), and across different tissues include Hubner et al (2005) and Langley et al (2013). Gene regulatory networks can also be linked to the genome (exemplified by Lindblom et al, 2014; Saba et al, 2015; Wang et al, 2015). (B) Metabolite levels, or the metabolome, can also be linked to the genome (exemplified by Dumas et al, 2007).…”

“…For instance, Lindblom et al published a study in 2014 (Table 1) in which they performed eQTL mapping in data collected from spinal cord tissue of an F 2 rat intercross (Lindblom et al, 2014). The aim of this work was to better understand the mechanisms regulating complement activation in the central nervous system.…”

“…This uncovered gene co-expression networks and functional processes involved in complement activation. Furthermore, by carrying out enrichments of transcription factor binding sites within the genes forming part of the trans -regulated networks, they identified a previously unknown FOX family transcription factor as a potential candidate for the regulation of a trans -eQTL cluster that includes the complement protein C3 gene (Lindblom et al, 2014). An elegant study by Saba et al (Table 1) used the B×H/H×B RI strains panel to build co-expression networks in the brain and associate these to predisposition towards alcohol consumption (Saba et al, 2015).…”