The amyloid-ß1-42-oligomer interacting peptide D-AIP possesses favorable biostability, pharmacokinetics, and brain region distribution.

We have previously developed a unique 8-amino acid Aß42 oligomer-Interacting Peptide (AIP) as a novel anti-amyloid strategy for the treatment of Alzheimer's disease. Our lead candidate has successfully progressed from test tubes (i.e., in vitro characterization of protease-resistant D-AIP) to transgenic flies (i.e., in vivo rescue of human Aß42-mediated toxicity via D-AIP-supplemented food). In the present study, we examined D-AIP in terms of its stability in multiple biological matrices (i.e., ex-vivo mouse plasma, whole blood, and liver S9 fractions) using MALDI mass spectrometry, pharmacokinetics using a rapid and sensitive LC-MS method, and blood brain barrier (BBB) penetrance in WT C57LB/6 mice. D-AIP was found to be relatively stable over 3 h at 37 °C in all matrices tested. Finally, label-free MALDI imaging showed that orally administered D-AIP can readily penetrate the intact BBB in both male and female WT mice. Based upon the favorable stability, pharmacokinetics, and BBB penetration outcomes for orally administered D-AIP in WT mice, we then examined the effect of D-AIP on amyloid "seeding" in vitro (i.e., freshly monomerized versus preaggregated Aß42). Complementary biophysical assays (ThT, TEM, and MALDI-TOF MS) showed that D-AIP can directly interact with synthetic Aß42 aggregates to disrupt primary and/or secondary seeding events. Taken together, the unique mechanistic and desired therapeutic potential of our lead D-AIP candidate warrants further investigation, that is, testing of D-AIP efficacy on the altered amyloid/tau pathology in transgenic mouse models of Alzheimer's disease.

Identification and mapping of central pair proteins by proteomic analysis.

Cilia or flagella of eukaryotes are small micro-hair like structures that are indispensable to single-cell motility and play an important role in mammalian biological processes. Cilia or flagella are composed of nine doublet microtubules surrounding a pair of singlet microtubules called the central pair (CP). Together, this arrangement forms a canonical and highly conserved 9+2 axonemal structure. The CP, which is a unique structure exclusive to motile cilia, is a pair of structurally dimorphic singlet microtubules decorated with numerous associated proteins. Mutations of CP-associated proteins cause several different physical symptoms termed as ciliopathies. Thus, it is crucial to understand the architecture of the CP. However, the protein composition of the CP was poorly understood. This was because the traditional method of identification of CP proteins was mostly limited by available Chlamydomonas mutants of CP proteins. Recently, more CP protein candidates were presented based on mass spectrometry results, but most of these proteins were not validated. In this study, we re-evaluated the CP proteins by conducting a similar comprehensive CP proteome analysis comparing the mass spectrometry results of the axoneme sample prepared from Chlamydomonas strains with and without CP complex. We identified a similar set of CP protein candidates and additional new 11 CP protein candidates. Furthermore, by using Chlamydomonas strains lacking specific CP sub-structures, we present a more complete model of localization for these CP proteins. This work has established a new foundation for understanding the function of the CP complex in future studies.

Facilitated aggregation of FG nucleoporins under molecular crowding conditions.

Intrinsically disordered and phenylalanine-glycine-rich nucleoporins (FG Nups) form a crowded and selective transport conduit inside the NPC that can only be transited with the help of nuclear transport receptors (NTRs). It has been shown in vitro that FG Nups can assemble into two distinct appearances, amyloids and hydrogels. If and how these phenomena are linked and if they have a physiological role still remains unclear. Using a variety of high-resolution fluorescence and electron microscopic (EM) tools, we reveal that crowding conditions mimicking the NPC environment can accelerate the aggregation and amyloid formation speed of yeast and human FG Nups by orders of magnitude. Aggregation can be inhibited by NTRs, providing a rationale on how the cell might control amyloid formation of FG Nups. The superb spatial resolving power of EM also reveals that hydrogels are enlaced amyloid fibres, and these findings have implications for existing transport models and for NPC assembly.