Methods of probing the interactions between small molecules and disordered proteins

It is generally recognized that a large fraction of the human proteome is made up of proteins that remain disordered in their native states. Despite the fact that such proteins play key biological roles and are involved in many major human diseases, they still represent challenging targets for drug discovery. A major bottleneck for the identification of compounds capable of interacting with these proteins and modulating their disease-promoting behaviour is the development of effective techniques to probe such interactions. The difficulties in carrying out binding measurements have resulted in a poor understanding of the mechanisms underlying these interactions. In order to facilitate further methodological advances, here we review the most commonly used techniques to probe three types of interactions involving small molecules: (1) those that disrupt functional interactions between disordered proteins; (2) those that inhibit the aberrant aggregation of disordered proteins, and (3) those that lead to binding disordered proteins in their monomeric states. In discussing these techniques, we also point out directions for future developments.Gabriella T. Heller is supported by the Gates Cambridge Trust Scholarship. Francesco A. Aprile is supported by a Senior Research Fellowship award from the Alzheimer’s Society, UK (grant number 317, AS-SF-16-003)

Measuring 129Xe transfer across the blood‐brain barrier using MR spectroscopy

Purpose This study develops a tracer kinetic model of xenon uptake in the human brain to determine the transfer rate of inhaled hyperpolarized 129Xe from cerebral blood to gray matter that accounts for the effects of cerebral physiology, perfusion and magnetization dynamics. The 129Xe transfer rate is expressed using a tracer transfer coefficient, which estimates the quantity of hyperpolarized 129Xe dissolved in cerebral blood under exchange with depolarized 129Xe dissolved in gray matter under equilibrium of concentration. Theory and Methods Time‐resolved MR spectra of hyperpolarized 129Xe dissolved in the human brain were acquired from three healthy volunteers. Acquired spectra were numerically fitted with five Lorentzian peaks in accordance with known 129Xe brain spectral peaks. The signal dynamics of spectral peaks for gray matter and red blood cells were quantified, and correction for the 129Xe T1 dependence upon blood oxygenation was applied. 129Xe transfer dynamics determined from the ratio of the peaks for gray matter and red blood cells was numerically fitted with the developed tracer kinetic model. Results For all the acquired NMR spectra, the developed tracer kinetic model fitted the data with tracer transfer coefficients between 0.1 and 0.14. Conclusion In this study, a tracer kinetic model was developed and validated that estimates the transfer rate of HP 129Xe from cerebral blood to gray matter in the human brain

Living apart together: crosstalk between the core and supernumerary genomes in a fungal plant pathogen

Background: Eukaryotes display remarkable genome plasticity, which can include supernumerary chromosomes that differ markedly from the core chromosomes. Despite the widespread occurrence of supernumerary chromosomes in fungi, their origin, relation to the core genome and the reason for their divergent characteristics are still largely unknown. The complexity of genome assembly due to the presence of repetitive DNA partially accounts for this. Results: Here we use single-molecule real-time (SMRT) sequencing to assemble the genome of a prominent fungal wheat pathogen, Fusarium poae, including at least one supernumerary chromosome. The core genome contains limited transposable elements (TEs) and no gene duplications, while the supernumerary genome holds up to 25 % TEs and multiple gene duplications. The core genome shows all hallmarks of repeat-induced point mutation (RIP), a defense mechanism against TEs, specific for fungi. The absence of RIP on the supernumerary genome accounts for the differences between the two (sub) genomes, and results in a functional crosstalk between them. The supernumerary genome is a reservoir for TEs that migrate to the core genome, and even large blocks of supernumerary sequence (> 200 kb) have recently translocated to the core. Vice versa, the supernumerary genome acts as a refuge for genes that are duplicated from the core genome. Conclusions: For the first time, a mechanism was determined that explains the differences that exist between the core and supernumerary genome in fungi. Different biology rather than origin was shown to be responsible. A "living apart together" crosstalk exists between the core and supernumerary genome, accelerating chromosomal and organismal evolution

Clinical and genetic delineation of autosomal recessive and dominant ACTL6B-related developmental brain disorders.

PURPOSE: This study aims to comprehensively delineate the phenotypic spectrum of ACTL6B-related disorders, previously associated with both autosomal recessive and autosomal dominant neurodevelopmental disorders. Molecularly, the role of the nucleolar protein ACTL6B in contributing to the disease has remained unclear. METHODS: We identified 105 affected individuals, including 39 previously reported cases, and systematically analysed detailed clinical and genetic data for all individuals. Additionally, we conducted knockdown experiments in neuronal cells to investigate the role of ACTL6B in ribosome biogenesis. RESULTS: Biallelic variants in ACTL6B are associated with severe-to-profound global developmental delay/intellectual disability (GDD/ID), infantile intractable seizures, absent speech, autistic features, dystonia, and increased lethality. De novo monoallelic variants result in moderate-to-severe GDD/ID, absent speech, and autistic features, while seizures and dystonia were less frequently observed. Dysmorphic facial features and brain abnormalities, including hypoplastic corpus callosum, parenchymal volume loss/atrophy, are common findings in both groups. We reveal that in the nucleolus, ACTL6B plays a crucial role in ribosome biogenesis, in particular in pre-rRNA processing. CONCLUSION: This study provides a comprehensive characterization of the clinical spectrum of both autosomal recessive and dominant forms of ACTL6B-associated disorders. It offers a comparative analysis of their respective phenotypes provides a plausible molecular explanation and suggests their inclusion within the expanding category of 'ribosomopathies'