To through light on the mechanisms underlying the stimulation and persistence of glial cell activation in Parkinsonism, we investigate the function of IFN-γ and TNF-α in experimental models of Parkinson's disease and analyze their relation with local glial cell activation. It was found that IFN-γ and TNF-α remained higher over the years in the serum and CNS of chronic Parkinsonian macaques than in untreated animals, accompanied by sustained glial activation (microglia and astroglia) in the substantia nigra pars compacta. Importantly, Parkinsonian monkeys showed persistent and increasing levels of IFN-γR signaling in both microglial and astroglial cells. In addition, experiments performed in IFN-γ and TNF-α KO mice treated with MPTP revealed that, even before dopaminergic cell death can be observed, the presence of IFN-γ and TNF-α is crucial for microglial and astroglial activation, and, together, they have an important synergistic role. Both cytokines were necessary for the full level of activation to be attained in both microglial and astroglial cells. These results demonstrate that IFN-γ signaling, together with the contribution of TNF-α, have a critical and cell-specific role in stimulating and maintaining glial cell activation in Parkinsonism.

The stress protein Nupr1 is a highly basic, multifunctional, intrinsically disordered protein (IDP). MSL1 is a histone acetyl transferase-associated protein, known to intervene in the dosage compensation complex (DCC). In this work, we show that both Nupr1 and MSL1 proteins were recruited and formed a complex into the nucleus in response to DNA-damage, which was essential for cell survival in reply to cisplatin damage. We studied the interaction of Nupr1 and MSL1, and their binding affinities to DNA by spectroscopic and biophysical methods. The MSL1 bound to Nupr1, with a moderate affinity (2.8 µM) in an entropically-driven process. MSL1 did not bind to non-damaged DNA, but it bound to chemically-damaged-DNA with a moderate affinity (1.2 µM) also in an entropically-driven process. The Nupr1 protein bound to chemically-damaged-DNA with a slightly larger affinity (0.4 µM), but in an enthalpically-driven process. Nupr1 showed different interacting regions in the formed complexes with Nupr1 or DNA; however, they were always disordered (“fuzzy”), as shown by NMR. These results underline a stochastic description of the functionality of the Nupr1 and its other interacting partners.

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Neuronal specification is regulated by the activity of transcription factors containing the basic helix-loop-helix motif (bHLH); these regulating proteins include, among others, the neurogenin (Ngn) family, related to the atonal family of genes. Neurogenin 1 (NGN1) is a 237-residue protein that contains a bHLH domain and is involved in neuronal differentiation. In this work, we synthesized the bHLH region of NGN1 (bHLHN) comprising residues 90-150 of the full-length NGN1. The domain is a monomeric natively unfolded protein with a pH-dependent premolten globule conformation, as shown by several spectroscopic techniques (namely, NMR, fluorescence, FTIR, and circular dichroism). The unfolded character of the domain also explains, first, the impossibility of its overexpression in several Escherichia coli strains and, second, its insolubility in aqueous buffers. To the best of our knowledge, this is the first extensive study of the conformational preferences of a bHLH domain under different solution conditions. Upon binding to two DNA E-boxes, the protein forms "fuzzy" complexes (that is, the complexes were not fully folded). The affinities of bHLHN for both DNA boxes were smaller than those of other bHLH domains, which might explain why the protein-DNA complexes were not fully folded.

Insulin-like growth factor-I (IGF-I) has protective effects against b-amyloid (Ab)-induced neuronal cell death. Because alterations of the somatostatinergic system have been described in Alzheimer's disease, we investigated the effects of the Ab peptide and the possible protective role of IGF-I on the somatostatinergic system of the rat temporal cortex and on cell death and phosphorylated (p)-Akt levels in this area. Ab25-35 was administered intracerebroventricularly to male rats via an osmotic minipump over 14 days (300 pmol/day). Another group received a subcutaneous IGF-I infusion (50 lg/kg/day), concomitant with Ab25-35 administration, whereas a third group received IGF-I alone. Ab25-35 significantly decreased the somatostatin (SRIF)-like immunoreactive content and the SRIF receptor density, as a result of a decrease in the levels of the SRIF receptor subtype 2. The inhibitory effect of SRIF on adenylyl cyclase activity was significantly lower after Ab25-35 infusion, whereas the levels of the inhibitory G protein subunit Gia1, Gia2 or Gia3 were unaltered. Cell death was increased and p-Akt levels decreased in Ab25-35-treated animals. IGF-I administration increased immunoreactive IGF-I levels in the temporal cortex and restored all parameters affected by Ab25-35 to baseline values. These findings suggest that IGF-I prevents the deleterious effect of Ab25-35 on the somatostatinergic system.

Since the publication of this article an error has been identified in one of the authors names. D Aguado-Yera should have been D Aguado-Llera. The correct list is shown above.

Alzheimer¢s disease (AD) is characterized by the presence of senile plaques containing b-amyloid (Ab) and neuronal loss (Walsh and Selkoe 2004). The most frequent alteration of a neuropeptide in the brain of patients with AD is the reduction of somatostatin (SRIF), implicated in learning and memory, in the temporal cortex (Reinikainen et al. 1990). The effects of SRIF are mediated via five SRIF receptor subtypes located in diverse brain areas, including the temporal cortex. These Abbreviations used: [Ca 2+ ] c , cytosolic calcium; AC, adenylyl cyclase; AD, Alzheimer¢s disease; Ab, b-amyloid; CREB, cAMP-responsive element binding protein; ECL, enhanced chemiluminescence; ERK, extracellular-signal regulated kinase; GPE, glycine-proline-glutamate; GSK3b, glycogen synthase kinase 3b; HSP27, heat-shock protein 27; IGF-I, insulin-like growth factor-I; PVDF, polyvinylidene fluoride; SDS, sodium dodecyl sulfate; SRIF, somatostatin; SRIF-LI, somatostatin-like immunoreactivity; sst, somatostatin receptor subtype; TTBS, Tween 20 Tris Buffered Saline; TUNEL, terminal deoxynucleotidyltransferasemediated dUTP nick end labeling. AbstractThe protective effects of insulin-like growth factor I on the somatostatin (SRIF) system in the temporal cortex after bamyloid (Ab) injury may be mediated through its N-terminal tripeptide glycine-proline-glutamate (GPE). GPE is cleaved to cyclo[Pro-Gly] (cPG), a metabolite suggested to mediate in neuroprotective actions. We evaluated the effects of GPE and cPG in the temporal cortex of Ab25-35-treated rats on SRIF and SRIF receptor protein and mRNA levels, adenylyl cyclase activity, cell death, Ab25-35 accumulation, cytosolic calcium levels ([Ca 2+ ] c ) and the intracellular signaling mechanisms involved. GPE and cPG did not change Ab25-35 levels, but GPE partially restored SRIF and SRIF receptor 2 protein content and mRNA levels and protected against cell death after Ab25-35 insult, which was coincident with Akt activation and glycogen synthase kinase 3b inhibition. In addition, GPE displaced glutamate from NMDA receptors and blocked the glutamate induced rise in cytosolic calcium in isolated rat neurons and moderately increased Ca 2+ influx per se. Our findings suggest that GPE, but not its metabolite, mimics insulin-like growth factor I effects on the SRIF system through a mechanism independent of Ab clearance that involves modulation of calcium and glycogen synthase kinase 3b signaling. Keywords: calcium, glycogen synthase kinase 3b, Gly-ProGlu, neuroprotection, somatostatin, b-amyloid. (Reisine and Bell 1995). A marked loss of SRIF receptors with unaltered levels of Gi proteins has been reported in the cortex of AD patients (Cowburn et al. 1991). Diverse approaches have been tested to block Ab toxicity and diminish the progression of AD. Insulin-like growth factor-I (IGF-I) that acts as a survival factor in the brain, inhibits glycogen synthase kinase 3b (GSK3b) through phosphorylation on Ser9 in a PI3K-dependent manner preventing apoptosis (Quevedo et al. 2000). However, its use...

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