Species: Mouse
Genes: Trem2
Mutations: TREM2 R47H
Modification: Trem2: Knock-In
Disease Relevance: Alzheimer's Disease
Strain Name: C57BL/6J-Trem2^em1Adiuj/J
Genetic Background: C57BL/6J
Availability: The Jackson Lab:Stock# 027918; Cryorecovery.

Summary

R47H is a rare variant in TREM2 that triples the risk of Alzheimer’s disease in heterozygous carriers. Several murine Trem2 R47H knock-in lines have been created using CRISPR/Cas9 gene editing, but early models, including Jackson Lab’s Trem2 R47H KI, were found to show decreased expression of Trem2 not seen in human carriers of the variant (Xiang et al., 2018; see Sep 2018 news). This lower Trem2 expression in R47H knock-in mice was traced to aberrant splicing of the mutant allele, which introduces a premature stop codon and could promote nonsense-mediated decay (Xiang et al., 2018). Nonetheless, Trem2 R47H KI mice are useful for studying the effects of partial loss of TREM2 function. This line was intercrossed with Jackson Lab’s APOE4 Knock-In line to create the APOE4/Trem2*R47H line that serves as a standard background, or “platform strain,” for the introduction of additional alleles associated with late-onset AD in the MODEL-AD project.

Although the vast majority of documented human R47H carriers are heterozygotes, characterization of this R47H knock-in model has focused on homozygotes.

Trem2 expression | Survival | Behavior | Neuropathology | Transcriptomics | Alterations in splicing | Response to demyelination | Synaptic pruning | Modification Details | Related Strains

Trem2 expression. Mice homozygous for the R47H allele exhibited an approximately 50 percent reduction in the level of Trem2 transcripts compared with wild-type mice (Xiang et al., 2018; Kotredes et al., 2021; Tran et al., 2023). In addition to the three Trem2 isoforms seen in wild-type brains, Trem2 R47H KI generate a transcript truncated within exon 2. The appearance of this aberrant transcript is accompanied by a greater than 50 percent reduction in levels of the major Trem2 isoform (Tran et al., 2023). Levels of TREM2 protein immunoprecipitated from the cortices of R47H KI mice are also approximately 50 percent those of wild-type mice (Kotredes et al., 2021).

Survival. Homozygous Trem2 R47H KI mice are viable and fertile. However, aged female mice show increased mortality, compared with mice carrying the wild-type alleles: The mortality rate for wild-type C57BL/6J animals was less than 5 percent at 24 months, but approximately 20 percent for female Trem2 R47H KI mice (Kotredes et al., 2021).

Behavior. The behavioral data summarized here were collected by the MODEL-AD Consortium and reported at the 2018 AAIC meeting. At 2 and 12 months of age, locomotor activity (distance traveled in an open-field test) and motor coordination (latency to fall on the rotarod test) are similar in Trem2 R47H KI and C57BL/6J control mice, with both genotypes showing an age-dependent decline in these measures. Two- and 12-month-old Trem2 R47H KI mice also performed similarly to control mice in a Y-maze test of working memory.

Neuropathology. No gross anatomical changes, hippocampal or cortical neuron loss, amyloid plaques, or neurofibrillary tangles were observed in mice up to 24 months of age (Kotredes et al., 2021). Consistent with the neuroanatomical findings, levels of plasma neurofilament-light chain, a putative biomarker for neuronal injury, were similar in Trem2 R47H KI and wild-type mice (Tran et al., 2023).

Transcriptomics. Alterations in the transcriptomes of Trem2 R47H KI mice, compared with mice homozygous for wild-type Trem2, are age- and sex-dependent (Kotredes et al., 2021). In year-old mice, males showed upregulation of genes related to RNA transport and spliceosomes and downregulation of genes involved in oxidative phosphorylation. Genes involved in RNA transport were also upregulated in females, while downregulated genes included those related to lysosomal pathways. At 24 months, downregulated genes were enriched in pathways related to inflammation, lysosomes, and phagosomes.

Alterations in splicing. Following up on their finding of upregulation of spliceosome genes in middle-aged R47H KI mice, MODEL-AD investigators evaluated genome-wide alterations in splicing (Pandey et al., 2023). They found that differentially spliced genes in the brains of 12-month R47H KI mice were enriched for neuronal functions, including glutamatergic and GABAergic synaptic transmission. Similar changes in splicing were found in the brains of Trem2 knockout mice, suggesting that these alterations may be a consequence of the reduced expression of Trem2 in the brains of the Trem2 R47H KI mice, rather than the R47H variant per se.

Response to demyelination. Degenerating myelin is a TREM2 ligand. To test whether R47H acts as a loss-of-function allele that impairs microglial responses to stimuli, homozygous Trem2 R47H KI mice were subjected to experimental demyelination (Tran et al., 2023). Administration of the copper chelator cuprizone caused similar degrees of demyelination in Trem2 R47H KI and wild-type mice and led to pronounced microgliosis in white-matter tracts. Trem2 expression in R47H KI mice increased in response to cuprizone, but to a lesser degree than in wild-type animals. Upregulation of inflammatory genes occurred in both wild-type and Trem2 R47H KI mice. Additionally, a number of genes were upregulated in the knock-in animals that were not altered in wild-type mice, Trem2 knockout mice, or mice carrying Trem2 R47H alleles engineered to express normal levels of Trem2.

Synaptic pruning. Trem2 R47H KI mice have been used to investigate synaptic pruning by microglia (Rueda‐Carrasco et al., 2023). When synapses become hyperactive (e.g., when exposed to Aβ oligomers), phosphatidylserine is exposed on the outer face of the plasma membrane and serves as an “eat-me” signal for microglia (see Sep 2023 news). As phosphatidylserine and Aβ are both TREM2 ligands, Trem2 R47H KI mice, with their reduced levels of TREM2, might be expected to show impaired synaptic pruning in the context of elevated Aβ, and this was indeed the case.

To study synaptic pruning in situ, Trem2 R47H KI mice were crossed with APP NL-F knock-in mice. In the latter line, the murine App gene was modified to contain a humanized Aβ region along with two AD-linked mutations. Aβ begins to accumulate in these animals around 6 months of age. Compared with wild-type mice, 6-month-old APP NL-F knock-in mice (with wild-type TREM2) had fewer excitatory synapses, a higher percentage of synapses with exposed phosphatidylserine, and more microglial engulfment of synapses. Mice carrying both APP NL-F and TREM2 R47H did not differ from wild-type mice in the degree of microglial engulfment of synapses or number of excitatory synapses and had an even higher percentage of synapses with exposed phosphatidylserine than APP NL-F mice with wild-type TREM2. These findings suggest that microglia expressing TREM2 R47H do not efficiently remove synapses tagged with phosphatidylserine.

Microglia cultured from neonatal mice were used to study synapse elimination in vitro. While microglia from wild-type mice preferentially engulfed synaptosomes that bound Aβ oligomers (oAβ), compared with synaptosomes devoid of oAβ, microglia from Trem2 R47H KI mice exhibited no such preference. In cultured neurons, oAβ evoked hyperactivity in dendritic spines—as indicated by an increased frequency of Ca²⁺ transients—associated with increased exposure of phosphatidylserine. Co-culture with wild-type microglia led to a resolution of these changes within 48 hours, while Trem2 R47H microglia did not ameliorate spine hyperactivity.

Modification Details

CRISPR/Cas9 was used to introduce an R47H missense mutation and two silent mutations (lysine AAG>AAA and alanine GCC>GCA) into the mouse Trem2 gene.  The silent mutations aid in genotyping and increase gene-editing efficiency.

Related Strains

This model was used to generate a double-mutant line, Trem2 R47H KI x APOE4, by crossing with a line carrying a humanized APOE4 gene (Jackson Lab Stock #027894).

Phenotype Characterization

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References

Mutations Citations

1. TREM2 R47H

News Citations

1. Model Morass? R47H Mutation Scuttles TREM2 Expression in Mice, Not People 13 Sep 2018
2. In Alzheimer Brain, Can Synaptic Pruning Be Good? 8 Sep 2023

Research Models Citations

1. APOE4 Knock-In (JAX)
2. Trem2 KO (KOMP)
3. Trem2 KO (JAX)
4. Trem2*R47H(NSS)
5. APP NL-F Knock-in

Paper Citations

1. Xiang X, Piers TM, Wefers B, Zhu K, Mallach A, Brunner B, Kleinberger G, Song W, Colonna M, Herms J, Wurst W, Pocock JM, Haass C. The Trem2 R47H Alzheimer's risk variant impairs splicing and reduces Trem2 mRNA and protein in mice but not in humans. Mol Neurodegener. 2018 Sep 6;13(1):49. PubMed.
2. Kotredes KP, Oblak A, Pandey RS, Lin PB, Garceau D, Williams H, Uyar A, O'Rourke R, O'Rourke S, Ingraham C, Bednarycek D, Belanger M, Cope Z, Foley KE, Logsdon BA, Mangravite LM, Sukoff Rizzo SJ, Territo PR, Carter GW, Sasner M, Lamb BT, Howell GR. Uncovering Disease Mechanisms in a Novel Mouse Model Expressing Humanized APOEε4 and Trem2*R47H. Front Aging Neurosci. 2021;13:735524. Epub 2021 Oct 11 PubMed. Correction.
3. Tran KM, Kawauchi S, Kramár EA, Rezaie N, Liang HY, Sakr JS, Gomez-Arboledas A, Arreola MA, Cunha CD, Phan J, Wang S, Collins S, Walker A, Shi KX, Neumann J, Filimban G, Shi Z, Milinkeviciute G, Javonillo DI, Tran K, Gantuz M, Forner S, Swarup V, Tenner AJ, LaFerla FM, Wood MA, Mortazavi A, MacGregor GR, Green KN. A Trem2R47H mouse model without cryptic splicing drives age- and disease-dependent tissue damage and synaptic loss in response to plaques. Mol Neurodegener. 2023 Feb 17;18(1):12. PubMed.
4. Pandey RS, Kotredes KP, Sasner M, Howell GR, Carter GW. Differential splicing of neuronal genes in a Trem2*R47H mouse model mimics alterations associated with Alzheimer's disease. BMC Genomics. 2023 Apr 4;24(1):172. PubMed.
5. Rueda-Carrasco J, Sokolova D, Lee SE, Childs T, Jurčáková N, Crowley G, De Schepper S, Ge JZ, Lachica JI, Toomey CE, Freeman OJ, Hardy J, Barnes SJ, Lashley T, Stevens B, Chang S, Hong S. Microglia-synapse engulfment via PtdSer-TREM2 ameliorates neuronal hyperactivity in Alzheimer's disease models. EMBO J. 2023 Oct 4;42(19):e113246. Epub 2023 Aug 14 PubMed. Correction.

Other Citations

1. APOE4/Trem2*R47H

External Citations

1. MODEL-AD
2. MODEL-AD Consortium
3. reported
4. The Jackson Lab:Stock# 027918

Further Reading