Effects of SPI1-mediated transcriptome remodeling on Alzheimer's disease-related phenotypes in mouse models of Aß amyloidosis.

SPI1 was recently reported as a genetic risk factor for Alzheimer's disease (AD) in large-scale genome-wide association studies. However, it is unknown whether SPI1 should be downregulated or increased to have therapeutic benefits. To investigate the effect of modulating SPI1 levels on AD pathogenesis, we performed extensive biochemical, histological, and transcriptomic analyses using both Spi1-knockdown and Spi1-overexpression mouse models. Here, we show that the knockdown of Spi1 expression significantly exacerbates insoluble amyloid-ß (Aß) levels, amyloid plaque deposition, and gliosis. Conversely, overexpression of Spi1 significantly ameliorates these phenotypes and dystrophic neurites. Further mechanistic studies using targeted and single-cell transcriptomics approaches demonstrate that altered Spi1 expression modulates several pathways, such as immune response pathways and complement system. Our data suggest that transcriptional reprogramming by targeting transcription factors, like Spi1, might hold promise as a therapeutic strategy. This approach could potentially expand the current landscape of druggable targets for AD.

Data regarding M1 muscarinic receptor-mediated modulation of hepatic catalase activity in response to oxidative stress.

We recently demonstrated the role of M1 muscarinic receptors (M1R) in modulating oxidative stress in liver and hepatocytes (Urrunaga et al., 2015) [1]. Here we provide data regarding the effect of a novel M1R agonist, VU0357017 (Lebois et al., 2010) [2], on H2O2-mediated hepatocyte injury, the effect of an M1R antagonist VU0255035 (Sheffler et al., 2009) [3] on catalase and super oxide dismutase (SOD) activities in H2O2-treated hepatocytes in vitro, and finally, the effect of M1R ablation on hepatic catalase activity in acetaminophen (APAP)-treated mice.

M1 Muscarinic Receptor Deficiency Attenuates Azoxymethane-Induced Chronic Liver Injury in Mice.

Cholinergic nervous system regulates liver injury. However, the role of M1 muscarinic receptors (M1R) in modulating chronic liver injury is uncertain. To address this gap in knowledge we treated M1R-deficient and WT mice with azoxymethane (AOM) for six weeks and assessed liver injury responses 14 weeks after the last dose of AOM. Compared to AOM-treated WT mice, M1R-deficient mice had attenuated liver nodularity, fibrosis and ductular proliferation, α-SMA staining, and expression of α1 collagen, Tgfß-R, Pdgf-R, Mmp-2, Timp-1 and Timp-2. In hepatocytes, these findings were associated with reductions of cleaved caspase-3 staining and Tnf-α expression. In response to AOM treatment, M1R-deficient mice mounted a vigorous anti-oxidant response by upregulating Gclc and Nqo1 expression, and attenuating peroxynitrite generation. M1R-deficient mouse livers had increased expression of Trail-R2, a promotor of stellate cell apoptosis; dual staining for TUNNEL and α-SMA revealed increased stellate cells apoptosis in livers from M1R-deficient mice compared to those from WT. Finally, pharmacological inhibition of M1R reduced H2O2-induced hepatocyte apoptosis in vitro. These results indicate that following liver injury, anti-oxidant response in M1R-deficient mice attenuates hepatocyte apoptosis and reduces stellate cell activation, thereby diminishing fibrosis. Therefore, targeting M1R expression and activation in chronic liver injury may provide therapeutic benefit.

M1 muscarinic receptors modify oxidative stress response to acetaminophen-induced acute liver injury.

The role of muscarinic receptor subtypes in modulating acute liver injury is unknown. We detected M1 muscarinic receptor (M1R) expression in human and murine hepatocytes, and investigated the consequences of M1R deficiency on acute liver injury in vivo and inhibiting M1R activation on hepatocyte injury in vitro. Age-matched wild-type (WT) and M1R-deficient (Chrm1(-/-)) male mice were injected intraperitoneally with 200mg/kg acetaminophen (APAP) and euthanized 0, 2, 4, 16, 24, and 36h later. Biochemical and histological parameters indicated that liver injury peaked within 16h after APAP treatment and resolved by 24h. Compared to WT, M1R-deficient mice had reduced intrahepatic hemorrhage and hepatocyte necrosis, reflected by an attenuated rise in serum alanine aminotransferase levels. Livers of M1R-deficient mice showed reduced hepatocyte DNA fragmentation and attenuated expression of injury cytokines (Il-1α, Il-1ß, Il-6, and Fasl). In all mice hepatic glutathione levels decreased after APAP injection, but they recovered more quickly in M1R-deficient mice. During the course of APAP-induced liver injury in M1R-deficient compared to WT mice, hepatic Nrf-2, Gclc, and Nqo1 expressions increased and nitrotyrosine generation decreased. APAP metabolic pathways were not altered by M1R deficiency; expression of hepatic Cyp2e1, Cyp1a2, Cyp3a11, Cyp3a13, Car, and Pxr was similar in Chrm1(-/-) and WT mice. Finally, treatment of murine AML12 hepatocytes with a novel M1R antagonist, VU0255035, attenuated H2O2-induced oxidative stress, prevented GSH depletion, and enhanced viability. We conclude that M1R modify hepatocyte responses to oxidative stress and that targeting M1R has therapeutic potential for toxic liver injury.

Comparative whole genome sequencing reveals phenotypic tRNA gene duplication in spontaneous Schizosaccharomyces pombe La mutants.

We used a genetic screen based on tRNA-mediated suppression (TMS) in a Schizosaccharomyces pombe La protein (Sla1p) mutant. Suppressor pre-tRNA(Ser)UCA-C47:6U with a debilitating substitution in its variable arm fails to produce tRNA in a sla1-rrm mutant deficient for RNA chaperone-like activity. The parent strain and spontaneous mutant were analyzed using Solexa sequencing. One synonymous single-nucleotide polymorphism (SNP), unrelated to the phenotype, was identified. Further sequence analyses found a duplication of the tRNA(Ser)UCA-C47:6U gene, which was shown to cause the phenotype. Ninety percent of 28 isolated mutants contain duplicated tRNA(Ser)UCA-C47:6U genes. The tRNA gene duplication led to a disproportionately large increase in tRNA(Ser)UCA-C47:6U levels in sla1-rrm but not sla1-null cells, consistent with non-specific low-affinity interactions contributing to the RNA chaperone-like activity of La, similar to other RNA chaperones. Our analysis also identified 24 SNPs between ours and S. pombe 972h- strain yFS101 that was recently sequenced using Solexa. By including mitochondrial (mt) DNA in our analysis, overall coverage increased from 52% to 96%. mtDNA from our strain and yFS101 shared 14 mtSNPs relative to a 'reference' mtDNA, providing the first identification of these S. pombe mtDNA discrepancies. Thus, strain-specific and spontaneous phenotypic mutations can be mapped in S. pombe by Solexa sequencing.