The degradation of p53 and its major E3 ligase Mdm2 is differentially dependent on the proteasomal ubiquitin receptor S5a.

p53 and its major E3 ligase Mdm2 are both ubiquitinated and targeted to the proteasome for degradation. Despite the importance of this in regulating the p53 pathway, little is known about the mechanisms of proteasomal recognition of ubiquitinated p53 and Mdm2. In this study, we show that knockdown of the proteasomal ubiquitin receptor S5a/PSMD4/Rpn10 inhibits p53 protein degradation and results in the accumulation of ubiquitinated p53. Overexpression of a dominant-negative deletion of S5a lacking its ubiquitin-interacting motifs (UIM)s, but which can be incorporated into the proteasome, also causes the stabilization of p53. Furthermore, small-interferring RNA (siRNA) rescue experiments confirm that the UIMs of S5a are required for the maintenance of low p53 levels. These observations indicate that S5a participates in the recognition of ubiquitinated p53 by the proteasome. In contrast, targeting S5a has no effect on the rate of degradation of Mdm2, indicating that proteasomal recognition of Mdm2 can be mediated by an S5a-independent pathway. S5a knockdown results in an increase in the transcriptional activity of p53. The selective stabilization of p53 and not Mdm2 provides a mechanism for p53 activation. Depletion of S5a causes a p53-dependent decrease in cell proliferation, demonstrating that p53 can have a dominant role in the response to targeting S5a. This study provides evidence for alternative pathways of proteasomal recognition of p53 and Mdm2. Differences in recognition by the proteasome could provide a means to modulate the relative stability of p53 and Mdm2 in response to cellular signals. In addition, they could be exploited for p53-activating therapies. This work shows that the degradation of proteins by the proteasome can be selectively dependent on S5a in human cells, and that this selectivity can extend to an E3 ubiquitin ligase and its substrate

Measurements of double-polarized compton scattering asymmetries and extraction of the proton spin polarizabilities

The spin polarizabilities of the nucleon describe how the spin of the nucleon responds to an incident polarized photon. The most model-independent way to extract the nucleon spin polarizabilities is through polarized Compton scattering. Double-polarized Compton scattering asymmetries on the proton were measured in the Δ(1232) region using circularly polarized incident photons and a transversely polarized proton target at the Mainz Microtron. Fits to asymmetry data were performed using a dispersion model calculation and a baryon chiral perturbation theory calculation, and a separation of all four proton spin polarizabilities in the multipole basis was achieved. The analysis based on a dispersion model calculation yields γE1E1=−3.5±1.2, γM1M1=3.16±0.85, γE1M2=−0.7±1.2, and γM1E2=1.99±0.29, in units of 10−4  fm4

A new measurement of the neutron detection efficiency for the NaI Crystal Ball detector

We report on a measurement of the neutron detection efficiency in NaI crystals in the Crystal Ball detector obtained from a study of single p0 photoproduction on deuterium using the tagged photon beam at the Mainz Microtron. The results were obtained up to a neutron energy of 400 MeV. They are compared to previous measurements made more than 15 years ago at the pion beam at the BNL AGS

Amyloid polymorphisms constitute distinct clouds of conformational variants in different etiological subtypes of Alzheimer's disease

The molecular architecture of amyloids formed in vivo can be interrogated using luminescent conjugated oligothiophenes (LCOs), a unique class of amyloid dyes. When bound to amyloid, LCOs yield fluorescence emission spectra that reflect the 3D structure of the protein aggregates. Given that synthetic amyloid-β peptide (Aβ) has been shown to adopt distinct structural conformations with different biological activities, we asked whether Aβ can assume structurally and functionally distinct conformations within the brain. To this end, we analyzed the LCO-stained cores of β-amyloid plaques in postmortem tissue sections from frontal, temporal, and occipital neocortices in 40 cases of familial Alzheimer's disease (AD) or sporadic (idiopathic) AD (sAD). The spectral attributes of LCO-bound plaques varied markedly in the brain, but the mean spectral properties of the amyloid cores were generally similar in all three cortical regions of individual patients. Remarkably, the LCO amyloid spectra differed significantly among some of the familial and sAD subtypes, and between typical patients with sAD and those with posterior cortical atrophy AD. Neither the amount of Aβ nor its protease resistance correlated with LCO spectral properties. LCO spectral amyloid phenotypes could be partially conveyed to Aβ plaques induced by experimental transmission in a mouse model. These findings indicate that polymorphic Aβ-amyloid deposits within the brain cluster as clouds of conformational variants in different AD cases. Heterogeneity in the molecular architecture of pathogenic Aβ among individuals and in etiologically distinct subtypes of AD justifies further studies to assess putative links between Aβ conformation and clinical phenotype

Silencing of PTK7 in Colon Cancer Cells: Caspase-10-Dependent Apoptosis via Mitochondrial Pathway

Protein tyrosine kinase-7 (PTK7) is a catalytically inactive receptor tyrosine kinase (RTK). PTK7 is upregulated in many common human cancers, including colon cancer, lung cancer, gastric cancer and acute myeloid leukemia. The reason for this up-regulation is not yet known. To explore the functional role of PTK7, the expression of PTK7 in HCT 116 cells was examined using small interference (siRNA)-mediated gene silencing. Following transfection, the siRNA successfully suppressed PTK7 mRNA and protein expression. Knocking down of PTK7 in HCT 116 cells inhibited cell proliferation compared to control groups and induced apoptosis. Furthermore, this apoptosis was characterized by decreased mitochondrial membrane potential and activation of caspase-9 and -10. Addition of a caspase-10 inhibitor totally blocked this apoptosis, suggesting that caspase-10 may play a critical role in PTK7-knockdown-induced apoptosis, downstream of mitochondria. These observations may indicate a role for PTK7 in cell proliferation and cell apoptosis and may provide a potential therapeutic pathway for the treatment of a variety of cancers

Neurofilaments in spinocerebellar ataxia type 3: blood biomarkers at the preataxic and ataxic stage in humans and mice

With molecular treatments coming into reach for spinocerebellar ataxia type 3 (SCA3), easily accessible, cross-species validated biomarkers for human and preclinical trials are warranted, particularly for the preataxic disease stage. We assessed serum levels of neurofilament light (NfL) and phosphorylated neurofilament heavy (pNfH) in ataxic and preataxic subjects of two independent multicentric SCA3 cohorts and in a SCA3 knock-in mouse model. Ataxic SCA3 subjects showed increased levels of both NfL and pNfH. In preataxic subjects, NfL levels increased with proximity to the individual expected onset of ataxia, with significant NfL elevations already 7.5 years before onset. Cross-sectional NfL levels correlated with both disease severity and longitudinal disease progression. Blood NfL and pNfH increases in human SCA3 were each paralleled by similar changes in SCA3 knock-in mice, here also starting already at the presymptomatic stage, closely following ataxin-3 aggregation and preceding Purkinje cell loss in the brain. Blood neurofilaments, particularly NfL, might thus provide easily accessible, cross-species validated biomarkers in both ataxic and preataxic SCA3, associated with earliest neuropathological changes, and serve as progression, proximity-to-onset and, potentially, treatment-response markers in both human and preclinical SCA3 trials

Is music enriching for group-housed captive chimpanzees (Pan troglodytes)?

Many facilities that house captive primates play music for animal enrichment or for caregiver enjoyment. However, the impact on primates is unknown as previous studies have been inconclusive. We conducted three studies with zoo-housed chimpanzees (Pan troglodytes) and one with group-housed chimpanzees at the National Centre for Chimpanzee Care to investigate the effects of classical and pop/rock music on various variables that may be indicative of increased welfare. Study one compared the behaviour and use of space of 18 animals when silence, classical or pop/rock music was played into one of several indoor areas. Overall, chimpanzees did not actively avoid the area when music was playing but were more likely to exit the area when songs with higher beats per minute were broadcast. Chimpanzees showed significantly fewer active social behaviours when music, rather than silence, was playing. They also tended to be more active and engage in less abnormal behaviour during the music but there was no change to either self-grooming or aggression between music and silent conditions. The genre of music had no differential effects on the chimpanzees’ use of space and behaviour. In the second study, continuous focal observations were carried out on three individuals with relatively high levels of abnormal behaviour. No differences in behaviour between music and silence periods were found in any of the individuals. The final two studies used devices that allowed chimpanzees to choose if they wanted to listen to music of various types or silence. Both studies showed that there were no persistent preferences for any type of music or silence. When taken together, our results do not suggest music is enriching for group-housed captive chimpanzees, but they also do not suggest that music has a negative effect on welfare

Short-term effects of unilateral lesion of the primary motor cortex (M1) on ipsilesional hand dexterity in adult macaque monkeys

Although the arrangement of the corticospinal projection in primates is consistent with a more prominent role of the ipsilateral motor cortex on proximal muscles, rather than on distal muscles involved in manual dexterity, the role played by the primary motor cortex on the control of manual dexterity for the ipsilateral hand remains a matter a debate, either in the normal function or after a lesion. We, therefore, tested the impact of permanent unilateral motor cortex lesion on the manual dexterity of the ipsilateral hand in 11 macaque monkeys, within a time window of 60 days post-lesion. For comparison, unilateral reversible pharmacological inactivation of the motor cortex was produced in an additional monkey. Manual dexterity was assessed quantitatively based on three motor parameters derived from two reach and grasp manual tasks. In contrast to the expected dramatic, complete deficit of manual dexterity of the contralesional hand that persists for several weeks, the impact on the manual dexterity of the ipsilesional hand was generally moderate (but statistically significant) and, when present, lasted less than 20 days. Out of the 11 monkeys, only 3 showed a deficit of the ipsilesional hand for 2 of the 3 motor parameters, and 4 animals had a deficit for only one motor parameter. Four monkeys did not show any deficit. The reversible inactivation experiment yielded results consistent with the permanent lesion data. In conclusion, the primary motor cortex exerts a modest role on ipsilateral manual dexterity, most likely in the form of indirect hand postural control

Visualization of acetylcholine distribution in central nervous system tissue sections by tandem imaging mass spectrometry

Metabolite distribution imaging via imaging mass spectrometry (IMS) is an increasingly utilized tool in the field of neurochemistry. As most previous IMS studies analyzed the relative abundances of larger metabolite species, it is important to expand its application to smaller molecules, such as neurotransmitters. This study aimed to develop an IMS application to visualize neurotransmitter distribution in central nervous system tissue sections. Here, we raise two technical problems that must be resolved to achieve neurotransmitter imaging: (1) the lower concentrations of bioactive molecules, compared with those of membrane lipids, require higher sensitivity and/or signal-to-noise (S/N) ratios in signal detection, and (2) the molecular turnover of the neurotransmitters is rapid; thus, tissue preparation procedures should be performed carefully to minimize postmortem changes. We first evaluated intrinsic sensitivity and matrix interference using Matrix Assisted Laser Desorption/Ionization (MALDI) mass spectrometry (MS) to detect six neurotransmitters and chose acetylcholine (ACh) as a model for study. Next, we examined both single MS imaging and MS/MS imaging for ACh and found that via an ion transition from m/z 146 to m/z 87 in MS/MS imaging, ACh could be visualized with a high S/N ratio. Furthermore, we found that in situ freezing method of brain samples improved IMS data quality in terms of the number of effective pixels and the image contrast (i.e., the sensitivity and dynamic range). Therefore, by addressing the aforementioned problems, we demonstrated the tissue distribution of ACh, the most suitable molecular specimen for positive ion detection by IMS, to reveal its localization in central nervous system tissues

Micelles as Delivery Vehicles for Oligofluorene for Bioimaging

With the successful development of organic/polymeric light emitting diodes, many organic and polymeric fluorophores with high quantum efficiencies and optical stability were synthesized. However, most of these materials which have excellent optical properties are insoluble in water, limiting their applications in biological fields. Herein, we used micelles formed from an amino-group-containing poly(ε-caprolactone)-block-poly(ethylene glycol) (PCL-b-PEG-NH2) to incorporate a hydrophobic blue emitter oligofluorene (OF) to enable its application in biological conditions. Although OF is completely insoluble in water, it was successfully transferred into aqueous solutions with a good retention of its photophysical properties. OF exhibited a high quantum efficiency of 0.84 in a typical organic solvent of tetrahydrofuran (THF). In addition, OF also showed a good quantum efficiency of 0.46 after being encapsulated into micelles. Two cells lines, human glioblastoma (U87MG) and esophagus premalignant (CP-A), were used to study the cellular internalization of the OF incorporated micelles. Results showed that the hydrophobic OF was located in the cytoplasm, which was confirmed by co-staining the cells with nucleic acid specific SYTO 9, lysosome specific LysoTracker Red®, and mitochondria specific MitoTracker Red. MTT assay indicated non-toxicity of the OF-incorporated micelles. This study will broaden the application of hydrophobic functional organic compounds, oligomers, and polymers with good optical properties to enable their applications in biological research fields