Alteration of gene expression and protein solubility of the PI 5-phosphatase SHIP2 are correlated with Alzheimer's disease pathology progression.

A recent large genome-wide association study has identified EGFR (encoding the epidermal growth factor EGFR) as a new genetic risk factor for late-onset AD. SHIP2, encoded by INPPL1, is taking part in the signalling and interactome of several growth factor receptors, such as the EGFR. While INPPL1 has been identified as one of the most significant genes whose RNA expression correlates with cognitive decline, the potential alteration of SHIP2 expression and localization during the progression of AD remains largely unknown. Here we report that gene expression of both EGFR and INPPL1 was upregulated in AD brains. SHIP2 immunoreactivity was predominantly detected in plaque-associated astrocytes and dystrophic neurites and its increase was correlated with amyloid load in the brain of human AD and of 5xFAD transgenic mouse model of AD. While mRNA of INPPL1 was increased in AD, SHIP2 protein undergoes a significant solubility change being depleted from the soluble fraction of AD brain homogenates and co-enriched with EGFR in the insoluble fraction. Using FRET-based flow cytometry biosensor assay for tau-tau interaction, overexpression of SHIP2 significantly increased the FRET signal while siRNA-mediated downexpression of SHIP2 significantly decreased FRET signal. Genetic association analyses suggest that some variants in INPPL1 locus are associated with the level of CSF pTau. Our data support the hypothesis that SHIP2 is an intermediate key player of EGFR and AD pathology linking amyloid and tau pathologies in human AD.

CAG repeat mosaicism is gene specific in spinocerebellar ataxias.

Expanded CAG repeats in coding regions of different genes are the most common cause of dominantly inherited spinocerebellar ataxias (SCAs). These repeats are unstable through the germline, and larger repeats lead to earlier onset. We measured somatic expansion in blood samples collected from 30 SCA1, 50 SCA2, 74 SCA3, and 30 SCA7 individuals over a mean interval of 8.5 years, along with postmortem tissues and fetal tissues from SCA1, SCA3, and SCA7 individuals to examine somatic expansion at different stages of life. We showed that somatic mosaicism in the blood increases over time. Expansion levels are significantly different among SCAs and correlate with CAG repeat lengths. The level of expansion is greater in individuals with SCA7 who manifest disease compared to that of those who do not yet display symptoms. Brain tissues from SCA individuals have larger expansions compared to the blood. The cerebellum has the lowest mosaicism among the studied brain regions, along with a high expression of ATXNs and DNA repair genes. This was the opposite in cortices, with the highest mosaicism and lower expression of ATXNs and DNA repair genes. Fetal cortices did not show repeat instability. This study shows that CAG repeats are increasingly unstable during life in the blood and the brain of SCA individuals, with gene- and tissue-specific patterns.

Somatic copy number variant load in neurons of healthy controls and Alzheimer's disease patients.

The possible role of somatic copy number variations (CNVs) in Alzheimer's disease (AD) aetiology has been controversial. Although cytogenetic studies suggested increased CNV loads in AD brains, a recent single-cell whole-genome sequencing (scWGS) experiment, studying frontal cortex brain samples, found no such evidence. Here we readdressed this issue using low-coverage scWGS on pyramidal neurons dissected via both laser capture microdissection (LCM) and fluorescence activated cell sorting (FACS) across five brain regions: entorhinal cortex, temporal cortex, hippocampal CA1, hippocampal CA3, and the cerebellum. Among reliably detected somatic CNVs identified in 1301 cells obtained from the brains of 13 AD patients and 7 healthy controls, deletions were more frequent compared to duplications. Interestingly, we observed slightly higher frequencies of CNV events in cells from AD compared to similar numbers of cells from controls (4.1% vs. 1.4%, or 0.9% vs. 0.7%, using different filtering approaches), although the differences were not statistically significant. On the technical aspects, we observed that LCM-isolated cells show higher within-cell read depth variation compared to cells isolated with FACS. To reduce within-cell read depth variation, we proposed a principal component analysis-based denoising approach that significantly improves signal-to-noise ratios. Lastly, we showed that LCM-isolated neurons in AD harbour slightly more read depth variability than neurons of controls, which might be related to the reported hyperploid profiles of some AD-affected neurons.

The lipid phosphatase Synaptojanin 1 undergoes a significant alteration in expression and solubility and is associated with brain lesions in Alzheimer's disease.

Synaptojanin 1 (SYNJ1) is a brain-enriched lipid phosphatase critically involved in autophagosomal/endosomal trafficking, synaptic vesicle recycling and metabolism of phosphoinositides. Previous studies suggest that SYNJ1 polymorphisms have significant impact on the age of onset of Alzheimer's disease (AD) and that SYNJ1 is involved in amyloid-induced toxicity. Yet SYNJ1 protein level and cellular localization in post-mortem human AD brain tissues have remained elusive. This study aimed to examine whether SYNJ1 localization and expression are altered in post-mortem AD brains. We found that SYNJ1 is accumulated in Hirano bodies, plaque-associated dystrophic neurites and some neurofibrillary tangles (NFTs). SYNJ1 immunoreactivity was higher in neurons and in the senile plaques in AD patients carrying one or two ApolipoproteinE (APOE) ε4 allele(s). In two large cohorts of APOE-genotyped controls and AD patients, SYNJ1 transcripts were significantly increased in AD temporal isocortex compared to control. There was a significant increase in SYNJ1 transcript in APOEε4 carriers compared to non-carriers in AD cohort. SYNJ1 was systematically co-enriched with PHF-tau in the sarkosyl-insoluble fraction of AD brain. In the RIPA-insoluble fraction containing protein aggregates, SYNJ1 proteins were significantly increased and observed as a smear containing full-length and cleaved fragments in AD brains. In vitro cleavage assay showed that SYNJ1 is a substrate of calpain, which is highly activated in AD brains. Our study provides evidence of alterations in SYNJ1 mRNA level and SYNJ1 protein degradation, solubility and localization in AD brains.

Author Correction: TLR9 activation via microglial glucocorticoid receptors contributes to degeneration of midbrain dopamine neurons.

The originally published version of this Article contained an error in the subheading "Microglial GR does not affect DN loss triggered by TLR4 and TLR7," which was incorrectly given as "Microglial GR does affect DN loss triggered by TLR2 and TLR4". This has now been corrected in both the PDF and HTML versions of the Article.

TLR9 activation via microglial glucocorticoid receptors contributes to degeneration of midbrain dopamine neurons.

Inflammation is a characteristic feature of Parkinson's disease (PD). We examined the role of TLR9 and its regulation by glucocorticoid receptors (GRs) in degeneration of substantia nigra dopamine neurons (DNs). TLR9 agonist, CpG-ODN, induced DN degeneration in mice lacking GR in microglia but not in controls. TLR9 deletion reduced DN loss in neurotoxin, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) mouse model of PD. GR regulates TLR9 activation during MPTP neurotoxicity as TLR9 antagonist suppressed increased DN loss in microglia/macrophage GR mutant mice. GR absence in microglia enhanced TLR9 translocation to endolysosomes and facilitated its cleavage leading to pro-inflammatory gene expression. GR-dependent TLR9 activation also triggered DN loss following intranigral injection of mitochondrial DNA. Finally, microglial GR sensitivity to A53T-alpha-synuclein induced DN degeneration as well as decreased microglial GR expression observed in SN of PD brain samples, all suggest that reduced microglial GR activity in SN can stimulate TLR9 activation and DN loss in PD pathology.

Aging, but not tau pathology, impacts olfactory performances and somatostatin systems in THY-Tau22 mice.

Somatostatin (SOM) cortical levels decline in Alzheimer's disease (AD) in correlation with cognitive impairment severity, the latter being closely related to the presence of neurofibrillary tangles. Impaired olfaction is another hallmark of AD tightly related to tau pathology in the olfactory pathways. Recent studies showed that SOM modulates olfactory processing, suggesting that alterations in SOM levels participate to olfactory deficits in AD. Herein, we first observed that human olfactory peduncle and cortex are enriched in SOM cells and fibers, in aged postmortem brains. Then, the possible link between SOM alterations and olfactory deficits was evaluated by exploring the impact of age and tau hyperphosphorylation on olfactory SOM networks and behavioral performances in THY-Tau22 mice, a tauopathy transgenic model. Distinct molecular repertoires of SOM peptide and receptors were associated to sensory or cortical olfactory processing structures. Aging mainly affected SOM neurotransmission in piriform and entorhinal cortex in wild-type mice, although olfactory performances decreased. However, no further olfactory impairment was evidenced in THY-Tau22 mice until 12 months although tau pathology early affected olfactory cortical structures. Thus, tau hyperphosphorylation per se has a limited impact on olfactory performances in THY-Tau22 mice.

Cynomolgus macaques immunized with two HIV-1 Tat stabilized proteins raise strong and long-lasting immune responses with a pattern of Th1/Th2 response differing from that in mice.

The transcriptional transactivator (Tat) of HIV-1 is regarded as an attractive target for the development of an AIDS vaccine. However, investigations have suggested that Tat is poorly immunogenic and prompted us to develop a strategy to increase its ability to raise an immune response. The strategy is based on stabilization of Tat using sulfated sugars, previously proven to boost its humoral immunogenic potency, in mice. We have now examined the pattern of immune response raised by two Tat-stabilized proteins previously mixed with Alum, in mice and macaques. We found that BALB/c and DBA/2 mice raise a Th2 immune response that contrasts with the mixed Th1/Th2 response observed in cynomolgus macaques indicating that the profile of response found in mice cannot be extrapolated to macaques. We thoroughly analyzed the macaque anti-Tat immune response and observed that the anti-Tat antibodies appear after only one immunization and that their titers remain elevated up to 8 weeks after the last injection. We found a similar behavior for the cellular immune response. Furthermore, we observed that approximately 50% of the IFN-gamma-secreting Tat-specific T-cells are CD8 lymphocytes. Last, we observed that the macaque sera neutralize the transactivating activity of Tat. This analysis shows that the two Tat-stabilized preparations raise a potent and long-lasting humoral and cellular immune response in cynomolgus macaque and opens an avenue to investigate whether a strong anti-Tat immune response can protect non-human primates against a viral challenge.