. C9orf72 is required for proper macrophage and microglial function in mice. Science. 2016 Mar 18;351(6279):1324-9. PubMed.


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  1. This study from the Baloh group describes the generation and characterization of mice deficient in the C9orf72 ortholog 3110043O21Rik. O’Rourke and colleagues generated heterozygous and homozygous C9orf72-null mice by two independent methods. Dose-dependent reduction in levels of the C9orf72 ortholog is demonstrated in heterozygous and homozygous mice obtained from both strategies. None of the mice developed any neuromuscular dysfunction, but both lines displayed splenomegaly and lymphadenopathy, which the authors hypothesize was related to dysfunction of myeloid cells, including peripheral macrophages and CNS microglia. In isolated myeloid cells they demonstrate accumulation of Lamp1-positive material, suggesting a deficiency in autophagy pathways; this is accompanied by a pro-inflammatory cytokine response and is rescued by the expression of human C9orf72.

    Autophagy dysfunction is consistent with what is already known about C9orf72 function: Analysis of C9orf72 protein domains led to the hypothesis that it is a DENN-domain protein with a role in vesicle trafficking (Levine et al., 2013). This was supported by the observed sign epistasis with TMEM106B (Gallagher et al., 2014, reviewed in Cooper-Knock et al., 2015). At the ALS Symposium in December 2015, Clotilde Lagier-Tourenne reported early findings from another loss-of-function C9orf72 mouse model that had been interpreted as signs of lymphoma. While we await the final paper, many of the changes she described are similar to those outlined in this report, suggesting the two studies might be consistent.

    Atanasio and colleagues report a similar phenotype with splenomegaly, lymphadenopathy, and proliferation of myeloid cells. In fact, this mouse model has perhaps a more severe phenotype, because homozygous C9orf72 nulls developed an autoimmune syndrome with similarities to systemic lupus erythematosus, including prominent glomerulonephropathy and even motor weakness, but not a classical ALS-like phenotype.  

    The strength of this report from O'Rourke et al. lies in the consistency of the findings from two mouse lines. The suggestion that all of the inflammatory changes seen are secondary to the dysfunction of the myeloid cells is interesting but not proven beyond all doubt in this study; that would require demonstration that the myeloid cell abnormalities are necessary and sufficient to produce the entire phenotype.

    O'Rourke et al. relate their findings to human C9orf72 disease by demonstrating accumulation of Lamp1-positive material in microglia of C9orf72-ALS patients and observing similar transcriptome changes (particularly with respect to genes involved in inflammation) in the mice to those seen in a study of C9orf72-ALS CNS tissue (Prudencio et al., 2015). This leads to an attractive hypothesis: Loss of C9orf72 function contributes to disease pathogenesis via impaired clearance of protein aggregates by microglia, but the aggregates might develop in response to other stimuli. Therefore C9orf72 loss of function would not be expected to produce neuronal toxicity in isolation, but might by working synergistically with other mechanisms. In the context of C9orf72 disease, a number of toxic gain-of-function pathways have been proposed based on RNA foci and dipeptide repeat proteins derived from the C9orf72 GGGGCC-repeat expansion. The authors point to studies suggesting a loss of C9orf72 function in the human C9orf72 disease and studies linking ALS and microglial dysfunction as support for their hypothesis. However, without a neurodegenerative phenotype in the C9orf72-null mice, this remains speculation to some extent. Indeed, Atanasio et al. come to the opposite conclusion despite very similar observations, though it should be noted that Atanasio and colleagues did not specifically compare their findings to the human disease or assess autophagy in C9orf72-null myeloid cells.

    It will be fascinating to see what happens when these C9orf72-null mice are crossed with the C9BAC mice recently reported by the same authors (O’Rourke et al., 2015). The C9BAC mice express the GGGGCC-expansion in the context of the full length human C9orf72 gene via a bacterial artificial chromosome (BAC); they develop RNA foci and dipeptide repeat protein inclusions but not other important features of C9orf72 disease, including neurodegeneration and, notably, neuroinflammation. If the hypothesis proposed by O’Rourke and colleagues is correct, then combining the two models might finally precipitate neuronal toxicity. 


    . The product of C9orf72, a gene strongly implicated in neurodegeneration, is structurally related to DENN Rab-GEFs. Bioinformatics. 2013 Feb 15;29(4):499-503. Epub 2013 Jan 16 PubMed.

    . TMEM106B is a genetic modifier of frontotemporal lobar degeneration with C9orf72 hexanucleotide repeat expansions. Acta Neuropathol. 2014 Mar;127(3):407-18. Epub 2014 Jan 19 PubMed.

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    . C9orf72 BAC Transgenic Mice Display Typical Pathologic Features of ALS/FTD. Neuron. 2015 Dec 2;88(5):892-901. PubMed.

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