Ennerfelt H, Frost EL, Shapiro DA, Holliday C, Zengeler KE, Voithofer G, Bolte AC, Lammert CR, Kulas JA, Ulland TK, Lukens JR. SYK coordinates neuroprotective microglial responses in neurodegenerative disease. Cell. 2022 Oct 27;185(22):4135-4152.e22. Epub 2022 Oct 17 PubMed.

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  1. In this new manuscript by Ennerfelt et al., the authors mechanistically assess the contributions of microglial spleen tyrosine kinase (SYK) across multiple mouse models that recapitulate aspects of drivers associated with neurodegeneration. Multiple complementary approaches were taken to examine known pathobiological responses in light of a loss-of-function conditional removal of Syk from microglia to ultimately determine whether this kinase is a critical regulatory checkpoint of multiple cell-surface receptors (i.e., Clec7a, CD33, and TREM2), which have been previously implicated in neurodegenerative conditions. Excitingly, the authors demonstrate a consistently worsened outcome in both the 5xFAD and cuprizone/experimental autoimmune encephalomyelitis models if they lack microglial Syk. Further, removal of Syk from microglia reduced the proportion of disease-associated microglia (DAM) transcriptional phenotypes in favor of both homeostatic and a newly described high Cd36-expressing population. These findings add further support to the notion that the maintenance of certain microglial subsets (i.e., DAM) is neuroprotective in the chronic stages of neurodegenerative disease models.

    Critically, the authors demonstrate that SYK is an indispensable regulatory molecule for maintaining these subsets in the degenerative milieu. Given the signal transduction role of SYK relative to TREM2, these findings are in line with several prior works demonstrating that removal of TREM2 exacerbates neuropathological sequelae in similar animal models (Piccio et al., 2007; Wang et al., 2015). Excitingly, the findings by Ennerfelt et al. suggest that new therapies with agonistic targeting of SYK may offer beneficial outcomes for neurodegenerative disease, similar to work targeting TREM2 directly (Zhao et al., 2022). 

    Complementary to these findings, Wang et al. demonstrate the TREM2-SYK signaling axis as a primary effector driving microglial responses to amyloid-induced neuropathology, implicating TREM2-SYK signaling as a key regulatory checkpoint for microglial transcriptional phenotype progression into the “DAM” staging. What is interesting in this study is that the authors demonstrate potential alternative mechanisms to engage SYK functionality if/when TREM2 signaling may be defective. They demonstrate that engaging the cell-surface receptor DECTIN1 (CLEC7A) via monoclonal antibody treatment was able to alter some pathological burden.

    Collectively, both of these manuscripts highlight potential therapeutic targets associated with driving increased SYK signaling as a means to ramp up microglial function in the face of amyloid-induced neuropathology.

    References:

    Piccio L, Buonsanti C, Mariani M, Cella M, Gilfillan S, Cross AH, Colonna M, Panina-Bordignon P. Blockade of TREM-2 exacerbates experimental autoimmune encephalomyelitis. Eur J Immunol. 2007 May;37(5):1290-301. PubMed.

    Wang Y, Cella M, Mallinson K, Ulrich JD, Young KL, Robinette ML, Gilfillan S, Krishnan GM, Sudhakar S, Zinselmeyer BH, Holtzman DM, Cirrito JR, Colonna M. TREM2 lipid sensing sustains the microglial response in an Alzheimer's disease model. Cell. 2015 Mar 12;160(6):1061-71. Epub 2015 Feb 26 PubMed.

    Zhao P, Xu Y, Fan X, Li L, Li X, Arase H, Tong Q, Zhang N, An Z. Discovery and engineering of an anti-TREM2 antibody to promote amyloid plaque clearance by microglia in 5XFAD mice. MAbs. 2022 Jan-Dec;14(1):2107971. PubMed.

    View all comments by Josh Morganti

  2. Wang et al. elegantly described new mechanistic insights into how TREM2 induces microglial cell responses to Aβ in mouse models of Alzheimer’s disease (AD) through SYK-dependent and SYK-independent pathways.

    Recent studies have shown characteristic expression changes in microglia around Aβ plaques, labeling them as disease-associated microglia (Keren-Shaul et al., 2017). DAM microglia have been characterized by TREM2-dependent upregulation of phagocytic and lipid metabolism genes (Keren-Shaul et al., 2017; Krasermann et al., 2017). Several innate immune receptors that are upregulated in DAM signal via SYK, including TREM2 and CLEC7A. Here, Wang et al. showed that Syk-deficient microglia impaired the formation of a protective microglial barrier around Aβ plaques and enhanced Aβ accumulation and behavioral deficits in in the 5xFAD mouse model of AD. Targeted deletion of SYK in microglia disrupted the PI3K-AKT-GSK3β-mTOR signaling pathway, thereby leading to metabolic defects. In line with the new and interesting study by Ennerfelt and colleagues, showing SYK modulates microglial activation and that SYK loss of function exacerbates Alzheimer’s disease neuropathology, Wang et al. likewise found that SYK was required for microglial clustering around Aβ plaques during both disease onset and AD progression in 5xFAD mice.

    While the articles by Ennerfelt et al. and Wang et al. showed that SYK deficiency impaired generation of the DAM response, Wang and colleagues also uncovered that SYK deletion led to accumulation of an Apoe-expressing microglial cell population that is prodromal to DAM. Furthermore, comparison of single-cell RNA-Seq datasets from 5xFAD;Syk-deficient and 5xFAD;Trem2-deficient microglia by Wang et al. suggested that Aβ pathology induced a TREM2-dependent but SYK-independent pathway in microglia that was sufficient to advance these cells to a prodromal stage of DAM. Biochemical analysis using bone-marrow-derived macrophage cultures from mice deficient for Syk, Trem2, or Dap10 showed that DAP10 (which also binds TREM2) sustained microglial cell proliferation, metabolic fitness, and Aβ phagocytosis independently of SYK. However, both non-redundant SYK and DAP10 signaling pathways were required for the implementation of full DAM response. Thus, TREM2 mediates microglial cell response to Aβ accumulation via SYK-dependent and -independent (e.g., DAP10) signaling pathways.

    Our recent observations suggest a critical role for the innate immune receptor TREM2 in modulating microglial pathology in AD (Griciuc at al., 2019). Although previous studies showed that SYK acts downstream of TREM2 (Yao et al., 2019), additional studies are required to investigate whether TREM2 relies partially or exclusively on SYK to control AD pathogenesis. To this end, Ennerfelt et al. compared their RNA-Seq dataset with our previously published dataset analyzing 5xFAD;Trem2-deficient microglia (Griciuc et al., 2019). They found that 25 percent of genes upregulated and 60 percent of genes downregulated in 5xFAD;Trem2-deficient microglia were shared with 5xFAD;Syk-deficient microglia. However, 5xFAD;Syk-deficient microglia also showed uniquely expressed genes not observed in 5xFAD;Trem2-deficient microglia. These important findings suggest that TREM2 signaling through SYK partially mediates the DAM response, and that SYK acts downstream of other receptors in addition to TREM2 in the 5xFAD mouse model.

    One such receptor, CLEC7A, which was originally defined as a fungal pathogen receptor, directly activates SYK. Ennerfelt and colleagues found that SYK was required for the CLEC7A-mediated phagocytic response in microglia, following injection of pustulan, a β-D-glucan that acts as a ligand for CLEC7A. These data suggest that CLEC7A signals through SYK to promote microglial phagocytosis in 5xFAD mice. Moreover, Wang et al. showed that antibody-mediated cross-linking and activation of CLEC7A in microglia expressing a TREM2 hypofunctional variant (TREM2R47H) on a 5xFAD background improved microglial activation.

    In summary, both papers suggest the exciting prospect that in addition to TREM2 signaling, the CLEC7A-SYK signaling pathway in microglia may represent a novel therapeutic target for AD.

    References:

    Keren-Shaul H, Spinrad A, Weiner A, Matcovitch-Natan O, Dvir-Szternfeld R, Ulland TK, David E, Baruch K, Lara-Astaiso D, Toth B, Itzkovitz S, Colonna M, Schwartz M, Amit I. A Unique Microglia Type Associated with Restricting Development of Alzheimer's Disease. Cell. 2017 Jun 15;169(7):1276-1290.e17. Epub 2017 Jun 8 PubMed.

    Krasemann S, Madore C, Cialic R, Baufeld C, Calcagno N, El Fatimy R, Beckers L, O'Loughlin E, Xu Y, Fanek Z, Greco DJ, Smith ST, Tweet G, Humulock Z, Zrzavy T, Conde-Sanroman P, Gacias M, Weng Z, Chen H, Tjon E, Mazaheri F, Hartmann K, Madi A, Ulrich JD, Glatzel M, Worthmann A, Heeren J, Budnik B, Lemere C, Ikezu T, Heppner FL, Litvak V, Holtzman DM, Lassmann H, Weiner HL, Ochando J, Haass C, Butovsky O. The TREM2-APOE Pathway Drives the Transcriptional Phenotype of Dysfunctional Microglia in Neurodegenerative Diseases. Immunity. 2017 Sep 19;47(3):566-581.e9. PubMed.

    Griciuc A, Patel S, Federico AN, Choi SH, Innes BJ, Oram MK, Cereghetti G, McGinty D, Anselmo A, Sadreyev RI, Hickman SE, El Khoury J, Colonna M, Tanzi RE. TREM2 Acts Downstream of CD33 in Modulating Microglial Pathology in Alzheimer's Disease. Neuron. 2019 Sep 4;103(5):820-835.e7. Epub 2019 Jul 10 PubMed.

    Yao H, Coppola K, Schweig JE, Crawford F, Mullan M, Paris D. Distinct Signaling Pathways Regulate TREM2 Phagocytic and NFκB Antagonistic Activities. Front Cell Neurosci. 2019;13:457. Epub 2019 Oct 10 PubMed.

    View all comments by Rudy Tanzi

  3. SYK is an essential regulator of several cellular responses and, in microglia, serves as downstream signaling molecule for many important receptors with innate immune function, including TREM2, CD37, Clec7A, as well as several members of the Fc receptor family. Mutations in several of these genes have been linked to increased risk for developing Alzheimer’s disease (AD) and/or multiple sclerosis (MS). In particular, TREM2 has been a focus of a great deal of recent research because it dramatically impacts the ability of microglia to interact with amyloid plaques in AD, and clear cellular debris and enhance recovery in models of MS. With that said, a role for SYK in the microglia response to AD or MS has not been established.

    These two new papers by Ennerfelt et al. and Wang et al. have described an essential role for SYK signaling in the normal microglial response to Aβ plaques and oligodendrocyte loss. Ennerfelt et al. utilized the 5xFAD mouse model of AD and the experimental autoimmune encephalomyelitis (EAE) model of MS, to show that SYK is an essential regulator of the microglial response to neurodegeneration. In 5xFAD mice, SYK-negative microglia were less likely to cluster around plaques, were less proliferative, less activated, and less phagocytic. The plaques in these mice were more prominent and neurotoxic. 

    Meanwhile, in EAE, silencing Syk in microglia resulted in mice that were less able to recover post-challenge and were unable to mount a normal disease-associated microglial (DAM) response. 

    These results show that SYK is an essential downstream regulator of the microglial response to challenge, and excitingly identifies SYK as a new target for possible therapeutic intervention. Wang et al. confirmed that SYK signaling was necessary for the normal DAM response in 5XFAD mice. Wang et al. also confirmed that loss of SYK signaling recapitulated the autophagy phenotype observed in microglia from Trem2-/- 5XFAD mice (Ulland  et al., 2017). Finally, Wang et al. showed that in a mouse model with decreased TREM2 signaling—the Trem2R47H 5xFAD mice—engaging Clec7A with an activating antibody could increase SYK signaling, leading to activation of microglia independently of TREM2 signaling. 

    The findings in these two new papers are exciting and could lead to the development of new therapeutics targeting the SYK pathway.

    References:

    Ulland TK, Song WM, Huang SC, Ulrich JD, Sergushichev A, Beatty WL, Loboda AA, Zhou Y, Cairns NJ, Kambal A, Loginicheva E, Gilfillan S, Cella M, Virgin HW, Unanue ER, Wang Y, Artyomov MN, Holtzman DM, Colonna M. TREM2 Maintains Microglial Metabolic Fitness in Alzheimer's Disease. Cell. 2017 Aug 10;170(4):649-663.e13. PubMed.

    View all comments by Tyler Ulland

  4. These are very relevant papers given the central role of microglia in Alzheimer’s disease and other neurodegenerative disease. It is also very nice to see that research performed independently and in parallel yields the same results.

    From a scientific point of view, the role of Syk in governing macrophage activation via ITAM receptors is not new—you can find literature on that over more than a decade ago. So, to me it is perhaps not surprising that this mechanism is conserved in microglia and TREM2 signaling. Having said that, it is very nice and relevant to see how targeting the intracellular pathway downstream of TREM2 produces a potent effect, not only in microglial responses, but also by changing the course of disease.

    Whereas these two papers focus on TREM2, Syk also acts downstream of other ITAM receptors, such as AXL and MERTK, that play a crucial role in interactions with Ab plaques, so it would be very interesting to investigate beyond TREM2 in follow up studies.

    Along the same lines, it would be important to determine how other microglial functions are altered—I see for example that microglial numbers and proliferation show changes—and to explore whether there could be any potential therapeutic window in modulating microglial intracellular signaling pathways. Whereas genetics clearly point to a role for microglia in AD, we see that manipulating microglial function has a strong impact in other models of disease, as well, such as multiple sclerosis.

    Regarding TREM2, it seems that whereas activation of microglia helps remove amyloid deposits, there is evidence that it may exacerbate tau pathology. While this may be related to timing, disease stages, etc., I think it is crucial to understand those fundamental differences before we can use these strategies in the clinic.

    View all comments by Renzo Mancuso

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