Evaluating Nuclei Concentration in Amyloid Fibrillation Reactions Using Back-Calculation Approach

Background: In spite of our extensive knowledge of the more than 20 proteins associated with different amyloid diseases, we do not know how amyloid toxicity occurs or how to block its action. Recent contradictory reports suggest that the fibrils and/or the oligomer precursors cause toxicity. An estimate of their temporal concentration may broaden understanding of the amyloid aggregation process. Methodology/Principal Findings: Assuming that conversion of folded protein to fibril is initiated by a nucleation event, we back-calculate the distribution of nuclei concentration. The temporal in vitro concentration of nuclei for the model hormone, recombinant human insulin, is estimated to be in the picomolar range. This is a conservative estimate since the back-calculation method is likely to overestimate the nuclei concentration because it does not take into consideration fibril fragmentation, which would lower the amount of nuclei Conclusions: Because of their propensity to form aggregates (non-ordered) and fibrils (ordered), this very low concentration could explain the difficulty in isolating and blocking oligomers or nuclei toxicity and the long onset time for amyloid diseases

A Mitochondrial Kinase Complex Is Essential to Mediate an ERK1/2-Dependent Phosphorylation of a Key Regulatory Protein in Steroid Biosynthesis

ERK1/2 is known to be involved in hormone-stimulated steroid synthesis, but its exact roles and the underlying mechanisms remain elusive. Both ERK1/2 phosphorylation and steroidogenesis may be triggered by cAMP/cAMP-dependent protein kinase (PKA)-dependent and-independent mechanisms; however, ERK1/2 activation by cAMP results in a maximal steroidogenic rate, whereas canonical activation by epidermal growth factor (EGF) does not. We demonstrate herein by Western blot analysis and confocal studies that temporal mitochondrial ERK1/2 activation is obligatory for PKA-mediated steroidogenesis in the Leydig-transformed MA-10 cell line. PKA activity leads to the phosphorylation of a constitutive mitochondrial MEK1/2 pool with a lower effect in cytosolic MEKs, while EGF allows predominant cytosolic MEK activation and nuclear pERK1/2 localization. These results would explain why PKA favors a more durable ERK1/2 activation in mitochondria than does EGF. By means of ex vivo experiments, we showed that mitochondrial maximal steroidogenesis occurred as a result of the mutual action of steroidogenic acute regulatory (StAR) protein –a key regulatory component in steroid biosynthesis-, active ERK1/2 and PKA. Our results indicate that there is an interaction between mitochondrial StAR and ERK1/2, involving a D domain with sequential basic-hydrophobic motifs similar to ERK substrates. As a result of this binding and only in the presence of cholesterol, ERK1/2 phosphorylates StAR at Ser232. Directed mutagenesis of Ser232 to a non-phosphorylable amino acid such as Ala (StAR S232A) inhibited in vitro StAR phosphorylation by active ERK1/2. Transient transfection of MA-10 cells with StAR S232A markedly reduced the yield of progesterone production. In summary, here we show that StAR is a novel substrate of ERK1/2, and that mitochondrial ERK1/2 is part of a multimeric protein kinase complex that regulates cholesterol transport. The role of MAPKs in mitochondrial function is underlined

Chacras of the pukara: the late occupation (tenth-sixteenth centuries) of Mesada de Andalhuala Banda, Yocavil (Catamarca, Argentina)

This chapter presents the results of archaeological survey undertaken in the Mesada de Andalhuala Banda, Southeast Yocavil Valley, Catamarca, Northwest Argentina (NWA). Three hundred and eighty-two architectural units (AU) were registered, which suggests a prolonged occupation during the first and second millennia AD. In this opportunity, I will focus on the Late Period occupation tenth?sixteenth centuries) and the evidence reflecting activities of the agricultural production cycle, including structures for cultivation, mounds, and longitudinal accumulation of stones that were the product of land clearing, irrigation systems, milling tools, and circular storage structures. Spatially associated with these features are simple and compound residential units with double-faced walls filled with rubble and sediment, morphology assignable to the Santa María culture, which could be the dwelling sector of the population tasked with food production. This information allowed us to hypothesize that the Mesada, as a main productive area, was occupied during the entire Late Period, a time that included climate change and interethnic conflict, and played an important role in relation to the nearby pukara settlement.Fil: Alvarez Larrain, Alina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Saavedra 15. Instituto de las Culturas. Universidad de Buenos Aires. Instituto de las Culturas; Argentina. Universidad Nacional Autónoma de México; Méxic