Activation of archaeal transcription mediated by recruitment of transcription factor B.

Archaeal promoters consist of a TATA box and a purine-rich adjacent upstream sequence (transcription factor B (TFB)-responsive element (BRE)), which are bound by the transcription factors TATA box-binding protein (TBP) and TFB. Currently, only a few activators of archaeal transcription have been experimentally characterized. The best studied activator, Ptr2, mediates activation by recruitment of TBP. Here, we present a detailed biochemical analysis of an archaeal transcriptional activator, PF1088, which was identified in Pyrococcus furiosus by a bioinformatic approach. Operon predictions suggested that an upstream gene, pf1089, is polycistronically transcribed with pf1088. We demonstrate that PF1088 stimulates in vitro transcription by up to 7-fold when the pf1089 promoter is used as a template. By DNase I and hydroxyl radical footprinting experiments, we show that the binding site of PF1088 is located directly upstream of the BRE of pf1089. Mutational analysis indicated that activation requires the presence of the binding site for PF1088. Furthermore, we show that activation of transcription by PF1088 is dependent upon the presence of an imperfect BRE and is abolished when the pf1089 BRE is replaced with a BRE from a strong archaeal promoter. Gel shift experiments showed that TFB recruitment to the pf1089 operon is stimulated by PF1088, and TFB seems to stabilize PF1088 operator binding even in the absence of TBP. Taken together, these results represent the first biochemical evidence for a transcriptional activator working as a TFB recruitment factor in Archaea, for which the designation TFB-RF1 is suggested.

Long-term in vivo imaging of fibrillar tau in the retina of P301S transgenic mice.

Tauopathies are widespread neurodegenerative disorders characterised by the intracellular accumulation of hyperphosphorylated tau. Especially in Alzheimer's disease, pathological alterations in the retina are discussed as potential biomarkers to improve early diagnosis of the disease. Using mice expressing human mutant P301S tau, we demonstrate for the first time a straightforward optical approach for the in vivo detection of fibrillar tau in the retina. Longitudinal examinations of individual animals revealed the fate of single cells containing fibrillar tau and the progression of tau pathology over several months. This technique is most suitable to monitor therapeutic interventions aimed at reducing the accumulation of fibrillar tau. In order to evaluate if this approach can be translated to human diagnosis, we tried to detect fibrillar protein aggregates in the post-mortem retinas of patients that had suffered from Alzheimer's disease or Progressive Supranuclear Palsy. Even though we could detect hyperphosphorylated tau, we did not observe any fibrillar tau or Aß aggregates. In contradiction to previous studies, our observations do not support the notion that Aß or tau in the retina are of diagnostic value in Alzheimer's disease.

Multiple events lead to dendritic spine loss in triple transgenic Alzheimer's disease mice.

The pathology of Alzheimer's disease (AD) is characterized by the accumulation of amyloid-ß (Aß) peptide, hyperphosphorylated tau protein, neuronal death, and synaptic loss. By means of long-term two-photon in vivo imaging and confocal imaging, we characterized the spatio-temporal pattern of dendritic spine loss for the first time in 3xTg-AD mice. These mice exhibit an early loss of layer III neurons at 4 months of age, at a time when only soluble Aß is abundant. Later on, dendritic spines are lost around amyloid plaques once they appear at 13 months of age. At the same age, we observed spine loss also in areas apart from amyloid plaques. This plaque independent spine loss manifests exclusively at dystrophic dendrites that accumulate both soluble Aß and hyperphosphorylated tau intracellularly. Collectively, our data shows that three spatio-temporally independent events contribute to a net loss of dendritic spines. These events coincided either with the occurrence of intracellular soluble or extracellular fibrillar Aß alone, or the combination of intracellular soluble Aß and hyperphosphorylated tau.