Axonopathy is a pronounced attribute of many neurodegenerative diseases. In Alzheimer's Disease (AD), axonal swellings and degeneration are prevalent and may contribute to the symptoms of AD senile dementia. Current limitations in identifying the contribution of axonal damage to AD include the inability to detect when this damage occurs in relation to other identifiers of AD because of the invasiveness of existing methods. To overcome this, we further developed the MRI methodology Manganese Enhanced MRI (MEMRI) to assess in vivo axonal transport rates. Prior to amyloid-beta (Aβ) deposition, the axonal transport rates in the Tg2576 mouse model of AD were normal. As Aβlevels increased and before plaque formation, we observed a significant decrease in axonal transport rates of the Tg2576 mice compared to controls. After plaque formation, the decline in the transport rate in the Tg2576 mice became even more pronounced. These data indicate that in vivo axonal transport rates decrease prior to plaque formation in the Tg2576 mouse model of AD.

The PutA flavoprotein from Escherichia coli is a transcriptional repressor and a bifunctional enzyme that regulates and catalyzes proline oxidation. PutA represses transcription of genes putA and putP by binding to the control DNA region of the put regulon. The objective of this study is to define and characterize the DNA binding domain of PutA. The DNA binding activity of PutA, a 1320 amino acid polypeptide, has been localized to N-terminal residues 1-261. After exploring a potential DNA-binding region and an N-terminal deletion mutant of PutA, residues 1-90 (PutA90) were determined to contain DNA binding activity and stabilize the dimeric structure of PutA. Cell-based transcriptional assays demonstrate that PutA90 functions as a transcriptional repressor in vivo. The dissociation constant of PutA90 with the put control DNA was estimated to be 110 nM, which is slightly higher than that of the PutA-DNA complex (K d ϳ 45 nM). Primary and secondary structure analysis of PutA90 suggested the presence of a ribbonhelix-helix DNA binding motif in residues 1-47. To test this prediction, we purified and characterized PutA47. PutA47 is shown to purify as an apparent dimer, to exhibit in vivo transcriptional activity, and to bind specifically to the put control DNA. In gel-mobility shift assays, PutA47 was observed to bind cooperatively to the put control DNA with an overall dissociation constant of 15 nM for the PutA47-DNA complex. Thus, Nterminal residues 1-47 are critical for DNA-binding and the dimeric structure of PutA. These results are consistent with the ribbon-helix-helix family of transcription factors.

␥-Secretase processing of the amyloid precursor protein (APP) generates A␤ 40 and A␤ 42 , peptides that constitute the principal components of the ␤-amyloid plaque pathology of Alzheimer's disease (AD). The ␥-secretase activity is executed by a high-molecular-weight complex of which presenilin 1 (PS1) is an essential component. PS1 is a multi-pass membrane protein, and the large hydrophilic loop domain between transmembrane domains 6 and 7 has been shown to interact with various proteins. To determine the physiological function of the loop domain, we created a strain of PS1 knock-in mice in which the exon 10, which encodes most of the hydrophilic loop sequence, was deleted from the endogenous PS1 gene. We report here that the homozygous exon 10-deleted mice are viable but exhibit drastically reduced ␥-secretase cleavage at the A␤ 40 , but not the A␤ 42 , site. Surprisingly, this reduction of A␤ 40 is associated with exacerbated plaque pathology when expressed on APP transgenic background. Thus, the PS1 loop plays a regulatory role in ␥-secretase processing, and decreased A␤ 40 , not increased A␤ 42 is likely the cause for the accelerated plaque deposition in these animals. Our finding supports a protective role of A␤ 40 against amyloid pathology and raises the possibility that impaired ␥-secretase activity could be the basis for AD pathogenesis in general.

Abstractα-synuclein is a small soluble, cytosolic protein which associates with vesicular membranes. It is a component of intracellular Lewy bodies present in Parkinson's disease and a subset of Alzheimer's disease (AD). In addition, early studies identified a fragment of α-synuclein in the amyloid plaques of AD patients. Hypothesizing that α-synuclein might modify the AD pathogenic process, we crossed the Tg2576 strain of APP transgenic mice onto an α-synuclein knockout background to determine the effects of α-synuclein on Aβ production and plaque deposition. We found that α-synuclein deficiency does not affect the Aβ levels, nor does it alter the age of onset of plaque pathology. To our surprise, however, loss of α-synuclein leads to a significant increase in plaque load in all areas of the forebrain at 18 months of age. This is associated with an increase in another synaptic protein, synaptophysin. We thus conclude that α-synuclein is not involved in seeding of the plaques, but rather suppresses the progression of plaque pathology at advanced stages.