Dap12 and Trem2, molecules involved in innate immunity and neurodegeneration, are co-expressed in the CNS

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Abstract

Polycystic lipomembranous osteodysplasia with sclerosing leukoencephalopathy (PLOSL) is a recessively inherited disease characterized by early onset dementia associated with bone cysts. Our group has recently established the molecular background of PLOSL by identifying mutations in DAP12 and TREM2 genes. To understand how loss of function of the immune cell activating DAP12/TREM2 signaling complex leads to dementia and loss of myelin, we have analyzed here Dap12 and Trem2 expression in the mouse CNS. We show that Dap12 and Trem2 are expressed from embryonic stage to adulthood, and demonstrate a highly similar expression pattern. In addition, we identify microglial cells and oligodendrocytes as the major Dap12/Trem2-producing cells in the CNS and, consequently, as the predominant cell types involved in PLOSL pathogenesis. These findings provide a good starting point for the study of the molecular mechanisms of this inherited dementia and new evidence for the involvement of the immune system in neuronal degeneration.

Introduction

Polycystic lipomembranous osteodysplasia with sclerosing leukoencephalopathy (PLOSL), also known as Nasu–Hakola disease, is a recessively inherited disease characterized by a combination of bone cysts and early onset progressive dementia. Pain and fractures in the ankles and wrists appear as first symptoms during the 3rd decade of life. Frontal lobe syndrome (euphoria and loss of social inhibitions) and dementia due to degradation of the white matter follow. The disease leads to severe dementia and finally death before age 50 (Hakola, 1972, Paloneva et al., 2001, Verloes et al., 1997). PLOSL has a global distribution, but it is enriched in the Finnish and Japanese populations. We have recently characterized the molecular background of PLOSL by identifying mutations in two genes: DAP12 and TREM2 (Paloneva et al., 2000, Paloneva et al., 2002). The disease phenotype is identical regardless of the defective gene.

Somewhat surprisingly, both of the genes behind PLOSL encode proteins that were originally identified as essential elements of the immune system. The first PLOSL gene DAP12 (also known as KARAP or TYROBP) encodes a transmembrane adapter protein, which plays an important role in myeloid and natural killer (NK) cell activation and thus in triggering and amplifying inflammatory responses (Lanier et al., 1998). DAP12 forms homodimers, which upon complex formation with ligand-bound receptors trigger an activating signal transduction pathway (Lanier and Bakker, 2000, McVicar et al., 1998). DAP12 interacts with many different receptors depending on cell type. TREM2, encoded by the second PLOSL gene, is one of the DAP12-associated receptors. The DAP12/TREM2 membrane complex plays a role in maturation and survival of human dendritic cells (Bouchon et al., 2001).

Interestingly, patients with PLOSL do not clinically manifest any immunological defects, but their symptoms appear only in the central nervous system (CNS) and bone. The function of DAP12 and TREM2 in these target tissues of PLOSL is poorly understood. Defects in microglia and osteoclasts have been proposed as the primary cause of PLOSL (Paloneva et al., 2002), since they represent cell types originating from the same myeloid lineage. Indeed, recent studies by our group and others show that in vitro-induced DAP12- or TREM2-deficient human osteoclasts differentiate inefficiently and have aberrant morphology (Cella et al., 2003, Paloneva et al., 2003). In addition, in vitro-induced osteoclasts from Dap12-deficient mice show an immature phenotype with reduced bone resorption activity (Kaifu et al., 2003).

Very little is known about the function of DAP12/TREM2 in the CNS. Studies of mRNA signal intensities by Northern blot from different regions of the human brain show that DAP12 and TREM2 follow a similar expression pattern (Paloneva et al., 2002). Mouse microglial cells have been demonstrated to express Trem2 transcripts (Schmid et al., 2002) and Dap12 protein (Kaifu et al., 2003). Dap12 protein was also detected in mouse oligodendrocytes (Kaifu et al., 2003). However, no comprehensive study of DAP12/TREM2 expression in the CNS has been conducted.

In order to gain insight into how DAP12/TREM2 deficiency leads to the CNS symptoms of PLOSL, we have here compared, in detail, the spatial and temporal expression of these genes in mouse CNS, as well as specified the CNS cell types expressing both these gene products.

Section snippets

Ethical aspects

This study was approved by the Chancellor's Animal Research Committee at the University of California Los Angeles as well as the ethics committee for the use of laboratory animals in the National Public Health Institute, Helsinki.

Northern blot analysis

Northern blots containing mRNA from different aged mice brains (Rnway laboratories Inc., Seoul, Korea) were probed for Dap12 and Trem2 expression according to the manufacturer's protocol. The 313-bp Dap12 and 310-bp Trem2 probes were produced by reverse

Dap12 and Trem2 are already expressed in mouse brain at embryonic day 17

Northern blots containing poly(A) RNA from mouse brain at different stages of development were probed with Dap12- and Trem2-specific probes. Expression of both Dap12 and Trem2 was already detected at embryonic day 18. Mouse brain at postnatal day 1, day 3, 1 week, 2 weeks, 3 weeks, 1 month, 2 months, 3 months, 6 months, and 1 year also showed expression of both mRNAs (Fig. 1). In Northern analysis, the steady-state transcript levels did not show significant variation during development. The

Discussion

In mouse, Dap12 and Trem2 expression in the CNS is obvious already at the embryonic stage, and the steady-state expression levels do not change significantly during development. Dap12 and Trem2 transcripts are expressed in the same regions of the brain with similar distributions. Dap12, however, is systematically expressed at a higher level than Trem2. In neonatal mice, expression of both transcripts is concentrated into subcortical regions whereas later in life, the expression is more

Acknowledgments

We thank Paula Hakala and Tuula Manninen for technical assistance and members of Dr. Daniel Geschwind's lab at University of California Los Angeles for their help with the in situ hybridizations. We are grateful to Aimee Trudeau for reviewing the language. This study was supported by the Centre of Excellence in Disease Genetics of the Academy of Finland and the Helsinki Graduate School in Biotechnology and Molecular Biology.

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