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NMD Takes the Immune Road to NDD
Neuron
Previews NMD Takes the Immune Road to NDD Eunha Kim1 and Jun R. Huh1,2,* 1Department
of Immunology, Blavatnik Institute, Harvard Medical School, Boston, MA 02115, USA Center for Immunologic Diseases, Harvard Medical School and Brigham and Women’s Hospital, Boston, MA 02115, USA *Correspondence: [email protected] https://doi.org/10.1016/j.neuron.2019.10.042 2Evergrande
Proper mRNA quality control prevents immune activation; when it goes awry, mice and flies develop abnormal behavioral phenotypes. In this issue of Neuron, Johnson et al. (2019) report that inhibiting nonsense-mediated mRNA decay (NMD) contributes to the pathogenesis of neurodevelopmental disorder (NDD) phenotypes by triggering aberrant immune activation. In the business sector, quality control (QC) is an essential step to ensure successful business outcomes. Similar to the business world, proper regulation of mRNA transcription and translation is regarded as an essential attribute in supporting wellness in living organisms (Frischmeyer and Dietz, 1999). To avoid producing C-terminally truncated proteins that can exert detrimental, dominant-negative functions to many cellular processes, those mRNAs with premature termination codons (PTCs) are recognized and subsequently degraded through the nonsense-mediated mRNA decay (NMD) pathway (Behm-Ansmant and Izaurralde, 2006). What happens if this mRNA QC pathway ceases to function during animal development? Clinical cases of both copy number variants (CNVs) and single-nucleotide variants, linked to the NMD pathway components, have been reported in human patients with neurodevelopmental disorder (NDD) symptoms including speech and language impairment, learning and memory deficits, and autism spectrum disorders (ASDs) (Laumonnier et al., 2010; Nguyen et al., 2014; Tarpey et al., 2007). To interrogate the critical function of the NMD pathway in developing brains, Johnson et al. (2019) generated a mouse line lacking expression of a key component of the NMD pathway, Upf2, in the mouse forebrain regions by crossing Upf2loxP/loxP with Camk2a-Cre mice (Upf2 fb-KO mice) (Figure 1). As expected, in the Upf2 fb-KO mice, the levels of NMD mRNA substrates such as Atf4 (activating transcription factor 4), Gadd45b (growth arrest and DNA damage 45b), Pdrg1 (P53 and DNA damage regulated 1), and Rassf (ras association
domain family member 1) were significantly increased in the hippocampus and the cortex, compared to control animals, but not in the cerebellum. Because humans carrying pathogenic variants of Upf2 exhibit speech and language impairment, the authors first studied the role of Upf2 in regulating vocal communication. Consistent with observations in mouse models of ASD (Wo¨hr and Scattoni, 2013), Upf2 fb-KO mice emitted fewer ultrasonic vocalization (USV) calls and their USVs had abnormal patterns. Additionally, Upf2 fb-KO mice exhibit impaired long-term memory (LTM), assessed by both the contextual fear conditioning and the Morris water maze tests. Impairment of LTM in Upf2 fb-KO mice was accompanied by significantly impaired long-term potentiation (LTP), measured in hippocampal slices of control versus Upf2 fb-KO mice. Johnson et al. (2019) also found that Upf2 fb-KO mice manifested deficits in social approach behavior as well as in reciprocal social interaction. Therefore, Upf2 fb-KO mice produced an array of behavioral symptoms, reminiscent of those observed in human NDD patients with CNVs or single-nucleotide pathogenic variants in UPF2 or other NMD components (Figure 1). Importantly, the authors also confirmed the evolutionarily conserved role of Upf2 in LTM formation using the Drosophila system. For example, classical olfactory conditioning experiments suggested that reduced expression of Upf2 in fly neurons (using Elav-Gal4) led to deficits in LTM responses in Drosophila. The authors next examined how impaired NMD responses lead to abnormal behavioral phenotypes in
mice. To address this question, Johnson et al. (2019) performed gene expression analyses with cells isolated from the forebrain regions of control versus Upf2 fb-KO mice. Intriguingly, differentially expressed genes between these two groups include those involved in immune-related pathways, such as the innate and adaptive immune response, antigen presentation and processing, and the complement cascade pathways. Exposure to heightened immune responses during development is a known risk factor for neurodevelopmental disorders and associated behavioral abnormalities (Choi et al., 2016; Smith et al., 2007). Given the significant increase in the immune activationrelated features in the brains of Upf2 fb-KO mice, the authors further analyzed the immune cell makeup in the brains. Compared to those of control mice, increased numbers of CD3+, CD11b+, B220+, and Ly6G+ immune cells were found in the brains but not in the spleens of Upf2 fb-KO mice (Figure 1). Furthermore, inflammatory cytokines such as RANTES (regulated upon activation, normal T cell expressed and secreted) and T cell chemo-attractants such as IP10 (interferon-g-induced protein 10) and MIG (monokine induced by gamma interferon) were highly expressed in the forebrain regions of Upf2 fb-KO mice. Consistent with these, the authors also observed both microgliosis and astrogliosis in the hippocampus as well as the neocortex of Upf2 fb-KO mice. Johnson et al. (2019) next tested if a causal relationship exists between hyper-activated immune phenotypes and the behavioral abnormalities manifested in Upf2 fb-KO mice. To do so, they treated Upf2 fb-KO mice with a
Neuron 104, November 20, 2019 ª 2019 Elsevier Inc. 625
Neuron
Previews P
Normal behavioral phenotypes (learning and memory, social novelty, behavioral exibility vocal communication)
SMG1
UPF1 UPF2 UPF3
mRNA decay
EJC
PTC
PTC-containing mRNA accumulation
SMG1 UPF1 UPF2
Truncated proteins
UPF3 EJC
PTC
REFERENCES Neurodevelopmental disorders/ behavioral de cits (Impaired learning and memory, social de cits, behavioral in exibility impaired vocal communication)
Immune activation
Rescued from behavioral de cits
SMG1 UPF1 UPF2 UPF3 PTC
EJC
Immunosuppresant
Figure 1. Impaired NMD Results in Neuroinflammation and Promotes Behavioral Deficits Mice with impaired Upf2-mediated NMD pathway display a wide range of behavioral deficits. They are also found with increased numbers of immune cell infiltrates and enhanced inflammatory cytokine production in the brains. Importantly, administration of immunosuppressive reagents to these affected mice ameliorates behavioral symptoms. The illustration was created with BioRender.com.
low concentration of cyclophosphamide (CyP), an immunosuppressive drug, starting at 1 month of age. These treatments reduced the inflammatory response in the hippocampal regions of Upf2 fb-KO mice, leading to decreased immune cell infiltration. Furthermore, the CyP treatments rescued mice from developing neurodevelopmental disorder-like phenotypes, including reduced USV calls, deficits in LTM, abnormal social phenotypes, and behavioral inflexibility. Additionally, the authors used minocycline (Mino), a tetracycline antibiotic known to have an immunosuppressive effect; Mino treatment of Upf2 fb-KO mice resulted in behavioral rescue (Figure 1). Taken together, the data from Johnson et al. (2019) identify a novel connection between Upf2-mediated NMD and suppression of the immune system in the forebrain regions, failure of which causes abnormal behavioral phenotypes with impaired learning and memory, social
626 Neuron 104, November 20, 2019
modulating immune cell activities may serve as a novel therapeutic option to treat patients with various NDD and neuropathological diseases in the future.
deficits, behavioral inflexibility, and deficits in vocal communication. The authors went on to show that such NDD phenotypes can be reversed with the help of reagents alleviating inflammation. Like many other exciting findings, the current study also generates many future directions to explore. What are the mechanisms by which impaired NMD increases neuroinflammation? How/ whether do components of the innate immune pathway sense increased mRNA species produced by deficits in the NMD pathway? What types of immune cells and immune-related molecules play critical roles in promoting abnormal behavioral phenotypes? And how do immune cells and/or enhanced inflammation affect brain development and synaptic plasticity? The current study beautifully illustrates how a mutation in a fundamental biological pathway such as NMD leads to aberrant immune activation that then promotes NDD phenotypes. Altogether,
Behm-Ansmant, I., and Izaurralde, E. (2006). Quality control of gene expression: a stepwise assembly pathway for the surveillance complex that triggers nonsense-mediated mRNA decay. Genes Dev. 20, 391–398. Choi, G.B., Yim, Y.S., Wong, H., Kim, S., Kim, H., Kim, S.V., Hoeffer, C.A., Littman, D.R., and Huh, J.R. (2016). The maternal interleukin-17a pathway in mice promotes autism-like phenotypes in offspring. Science 351, 933–939. Frischmeyer, P.A., and Dietz, H.C. (1999). Nonsense-mediated mRNA decay in health and disease. Hum. Mol. Genet. 8, 1893–1900. Johnson, J.L., Stoica, L., Liu, Y., Zhu, P.J., Bhattacharya, A., Buffington, S.A., Huq, R., Eissa, N.T., Larsson, O., Porse, B.T., et al. (2019). Inhibition of Upf2-Dependent NonsenseMediated Decay Leads to Behavioral and Neurophysiological Abnormalities by Activating the Immune Response. Neuron 104, this issue, 665–679. Laumonnier, F., Shoubridge, C., Antar, C., Nguyen, L.S., Van Esch, H., Kleefstra, T., Briault, S., Fryns, J.P., Hamel, B., Chelly, J., et al. (2010). Mutations of the UPF3B gene, which encodes a protein widely expressed in neurons, are associated with nonspecific mental retardation with or without autism. Mol. Psychiatry 15, 767–776. Nguyen, L.S., Wilkinson, M.F., and Gecz, J. (2014). Nonsense-mediated mRNA decay: inter-individual variability and human disease. Neurosci. Biobehav. Rev. 46, 175–186. Smith, S.E.P., Li, J., Garbett, K., Mirnics, K., and Patterson, P.H. (2007). Maternal immune activation alters fetal brain development through interleukin6. J. Neurosci. 27, 10695–10702. Tarpey, P.S., Raymond, F.L., Nguyen, L.S., Rodriguez, J., Hackett, A., Vandeleur, L., Smith, R., Shoubridge, C., Edkins, S., Stevens, C., et al. (2007). Mutations in UPF3B, a member of the nonsense-mediated mRNA decay complex, cause syndromic and nonsyndromic mental retardation. Nat. Genet. 39, 1127–1133. Wo¨hr, M., and Scattoni, M.L. (2013). Behavioural methods used in rodent models of autism spectrum disorders: current standards and new developments. Behav. Brain Res. 251, 5–17.
Previews NMD Takes the Immune Road to NDD Eunha Kim1 and Jun R. Huh1,2,* 1Department
of Immunology, Blavatnik Institute, Harvard Medical School, Boston, MA 02115, USA Center for Immunologic Diseases, Harvard Medical School and Brigham and Women’s Hospital, Boston, MA 02115, USA *Correspondence: [email protected] https://doi.org/10.1016/j.neuron.2019.10.042 2Evergrande
Proper mRNA quality control prevents immune activation; when it goes awry, mice and flies develop abnormal behavioral phenotypes. In this issue of Neuron, Johnson et al. (2019) report that inhibiting nonsense-mediated mRNA decay (NMD) contributes to the pathogenesis of neurodevelopmental disorder (NDD) phenotypes by triggering aberrant immune activation. In the business sector, quality control (QC) is an essential step to ensure successful business outcomes. Similar to the business world, proper regulation of mRNA transcription and translation is regarded as an essential attribute in supporting wellness in living organisms (Frischmeyer and Dietz, 1999). To avoid producing C-terminally truncated proteins that can exert detrimental, dominant-negative functions to many cellular processes, those mRNAs with premature termination codons (PTCs) are recognized and subsequently degraded through the nonsense-mediated mRNA decay (NMD) pathway (Behm-Ansmant and Izaurralde, 2006). What happens if this mRNA QC pathway ceases to function during animal development? Clinical cases of both copy number variants (CNVs) and single-nucleotide variants, linked to the NMD pathway components, have been reported in human patients with neurodevelopmental disorder (NDD) symptoms including speech and language impairment, learning and memory deficits, and autism spectrum disorders (ASDs) (Laumonnier et al., 2010; Nguyen et al., 2014; Tarpey et al., 2007). To interrogate the critical function of the NMD pathway in developing brains, Johnson et al. (2019) generated a mouse line lacking expression of a key component of the NMD pathway, Upf2, in the mouse forebrain regions by crossing Upf2loxP/loxP with Camk2a-Cre mice (Upf2 fb-KO mice) (Figure 1). As expected, in the Upf2 fb-KO mice, the levels of NMD mRNA substrates such as Atf4 (activating transcription factor 4), Gadd45b (growth arrest and DNA damage 45b), Pdrg1 (P53 and DNA damage regulated 1), and Rassf (ras association
domain family member 1) were significantly increased in the hippocampus and the cortex, compared to control animals, but not in the cerebellum. Because humans carrying pathogenic variants of Upf2 exhibit speech and language impairment, the authors first studied the role of Upf2 in regulating vocal communication. Consistent with observations in mouse models of ASD (Wo¨hr and Scattoni, 2013), Upf2 fb-KO mice emitted fewer ultrasonic vocalization (USV) calls and their USVs had abnormal patterns. Additionally, Upf2 fb-KO mice exhibit impaired long-term memory (LTM), assessed by both the contextual fear conditioning and the Morris water maze tests. Impairment of LTM in Upf2 fb-KO mice was accompanied by significantly impaired long-term potentiation (LTP), measured in hippocampal slices of control versus Upf2 fb-KO mice. Johnson et al. (2019) also found that Upf2 fb-KO mice manifested deficits in social approach behavior as well as in reciprocal social interaction. Therefore, Upf2 fb-KO mice produced an array of behavioral symptoms, reminiscent of those observed in human NDD patients with CNVs or single-nucleotide pathogenic variants in UPF2 or other NMD components (Figure 1). Importantly, the authors also confirmed the evolutionarily conserved role of Upf2 in LTM formation using the Drosophila system. For example, classical olfactory conditioning experiments suggested that reduced expression of Upf2 in fly neurons (using Elav-Gal4) led to deficits in LTM responses in Drosophila. The authors next examined how impaired NMD responses lead to abnormal behavioral phenotypes in
mice. To address this question, Johnson et al. (2019) performed gene expression analyses with cells isolated from the forebrain regions of control versus Upf2 fb-KO mice. Intriguingly, differentially expressed genes between these two groups include those involved in immune-related pathways, such as the innate and adaptive immune response, antigen presentation and processing, and the complement cascade pathways. Exposure to heightened immune responses during development is a known risk factor for neurodevelopmental disorders and associated behavioral abnormalities (Choi et al., 2016; Smith et al., 2007). Given the significant increase in the immune activationrelated features in the brains of Upf2 fb-KO mice, the authors further analyzed the immune cell makeup in the brains. Compared to those of control mice, increased numbers of CD3+, CD11b+, B220+, and Ly6G+ immune cells were found in the brains but not in the spleens of Upf2 fb-KO mice (Figure 1). Furthermore, inflammatory cytokines such as RANTES (regulated upon activation, normal T cell expressed and secreted) and T cell chemo-attractants such as IP10 (interferon-g-induced protein 10) and MIG (monokine induced by gamma interferon) were highly expressed in the forebrain regions of Upf2 fb-KO mice. Consistent with these, the authors also observed both microgliosis and astrogliosis in the hippocampus as well as the neocortex of Upf2 fb-KO mice. Johnson et al. (2019) next tested if a causal relationship exists between hyper-activated immune phenotypes and the behavioral abnormalities manifested in Upf2 fb-KO mice. To do so, they treated Upf2 fb-KO mice with a
Neuron 104, November 20, 2019 ª 2019 Elsevier Inc. 625
Neuron
Previews P
Normal behavioral phenotypes (learning and memory, social novelty, behavioral exibility vocal communication)
SMG1
UPF1 UPF2 UPF3
mRNA decay
EJC
PTC
PTC-containing mRNA accumulation
SMG1 UPF1 UPF2
Truncated proteins
UPF3 EJC
PTC
REFERENCES Neurodevelopmental disorders/ behavioral de cits (Impaired learning and memory, social de cits, behavioral in exibility impaired vocal communication)
Immune activation
Rescued from behavioral de cits
SMG1 UPF1 UPF2 UPF3 PTC
EJC
Immunosuppresant
Figure 1. Impaired NMD Results in Neuroinflammation and Promotes Behavioral Deficits Mice with impaired Upf2-mediated NMD pathway display a wide range of behavioral deficits. They are also found with increased numbers of immune cell infiltrates and enhanced inflammatory cytokine production in the brains. Importantly, administration of immunosuppressive reagents to these affected mice ameliorates behavioral symptoms. The illustration was created with BioRender.com.
low concentration of cyclophosphamide (CyP), an immunosuppressive drug, starting at 1 month of age. These treatments reduced the inflammatory response in the hippocampal regions of Upf2 fb-KO mice, leading to decreased immune cell infiltration. Furthermore, the CyP treatments rescued mice from developing neurodevelopmental disorder-like phenotypes, including reduced USV calls, deficits in LTM, abnormal social phenotypes, and behavioral inflexibility. Additionally, the authors used minocycline (Mino), a tetracycline antibiotic known to have an immunosuppressive effect; Mino treatment of Upf2 fb-KO mice resulted in behavioral rescue (Figure 1). Taken together, the data from Johnson et al. (2019) identify a novel connection between Upf2-mediated NMD and suppression of the immune system in the forebrain regions, failure of which causes abnormal behavioral phenotypes with impaired learning and memory, social
626 Neuron 104, November 20, 2019
modulating immune cell activities may serve as a novel therapeutic option to treat patients with various NDD and neuropathological diseases in the future.
deficits, behavioral inflexibility, and deficits in vocal communication. The authors went on to show that such NDD phenotypes can be reversed with the help of reagents alleviating inflammation. Like many other exciting findings, the current study also generates many future directions to explore. What are the mechanisms by which impaired NMD increases neuroinflammation? How/ whether do components of the innate immune pathway sense increased mRNA species produced by deficits in the NMD pathway? What types of immune cells and immune-related molecules play critical roles in promoting abnormal behavioral phenotypes? And how do immune cells and/or enhanced inflammation affect brain development and synaptic plasticity? The current study beautifully illustrates how a mutation in a fundamental biological pathway such as NMD leads to aberrant immune activation that then promotes NDD phenotypes. Altogether,
Behm-Ansmant, I., and Izaurralde, E. (2006). Quality control of gene expression: a stepwise assembly pathway for the surveillance complex that triggers nonsense-mediated mRNA decay. Genes Dev. 20, 391–398. Choi, G.B., Yim, Y.S., Wong, H., Kim, S., Kim, H., Kim, S.V., Hoeffer, C.A., Littman, D.R., and Huh, J.R. (2016). The maternal interleukin-17a pathway in mice promotes autism-like phenotypes in offspring. Science 351, 933–939. Frischmeyer, P.A., and Dietz, H.C. (1999). Nonsense-mediated mRNA decay in health and disease. Hum. Mol. Genet. 8, 1893–1900. Johnson, J.L., Stoica, L., Liu, Y., Zhu, P.J., Bhattacharya, A., Buffington, S.A., Huq, R., Eissa, N.T., Larsson, O., Porse, B.T., et al. (2019). Inhibition of Upf2-Dependent NonsenseMediated Decay Leads to Behavioral and Neurophysiological Abnormalities by Activating the Immune Response. Neuron 104, this issue, 665–679. Laumonnier, F., Shoubridge, C., Antar, C., Nguyen, L.S., Van Esch, H., Kleefstra, T., Briault, S., Fryns, J.P., Hamel, B., Chelly, J., et al. (2010). Mutations of the UPF3B gene, which encodes a protein widely expressed in neurons, are associated with nonspecific mental retardation with or without autism. Mol. Psychiatry 15, 767–776. Nguyen, L.S., Wilkinson, M.F., and Gecz, J. (2014). Nonsense-mediated mRNA decay: inter-individual variability and human disease. Neurosci. Biobehav. Rev. 46, 175–186. Smith, S.E.P., Li, J., Garbett, K., Mirnics, K., and Patterson, P.H. (2007). Maternal immune activation alters fetal brain development through interleukin6. J. Neurosci. 27, 10695–10702. Tarpey, P.S., Raymond, F.L., Nguyen, L.S., Rodriguez, J., Hackett, A., Vandeleur, L., Smith, R., Shoubridge, C., Edkins, S., Stevens, C., et al. (2007). Mutations in UPF3B, a member of the nonsense-mediated mRNA decay complex, cause syndromic and nonsyndromic mental retardation. Nat. Genet. 39, 1127–1133. Wo¨hr, M., and Scattoni, M.L. (2013). Behavioural methods used in rodent models of autism spectrum disorders: current standards and new developments. Behav. Brain Res. 251, 5–17.