The fatty acid binding protein FABP7 is required for optimal oligodendrocyte differentiation during myelination but not during remyelination.

The major constituents of the myelin sheath are lipids, which are made up of fatty acids (FAs). The hydrophilic environment inside the cells requires FAs to be bound to proteins, preventing their aggregation. Fatty acid binding proteins (FABPs) are one class of proteins known to bind FAs in a cell. Given the crucial role of FAs for myelin sheath formation we investigated the role of FABP7, the major isoform expressed in oligodendrocyte progenitor cells (OPCs), in developmental myelination and remyelination. Here, we show that the knockdown of Fabp7 resulted in a reduction of OPC differentiation in vitro. Consistent with this result, a delay in developmental myelination was observed in Fabp7 knockout animals. This delay was transient with full myelination being established before adulthood. FABP7 was dispensable for remyelination, as the knockout of Fapb7 did not alter remyelination efficiency in a focal demyelination model. In summary, while FABP7 is important in OPC differentiation in vitro, its function is not crucial for myelination and remyelination in vivo.This work was supported by grants from the UK Multiple Sclerosis Society, the Adelson Medical Research Foundation, the Japan Society for the Promotion of Science (JSPS) KAKENHI Grant and a core support grant from the Wellcome Trust and MRC to the Wellcome Trust-Medical Research Council Cambridge Stem Cell Institute. AGF was also supported by an ECTRIMS postdoctoral fellowship from July 2018. SF and TB were also supported by a Wellcome-Trust PhD studentship

Genome-Scale Networks Link Neurodegenerative Disease Genes to α-Synuclein through Specific Molecular Pathways

Numerous genes and molecular pathways are implicated in neurodegenerative proteinopathies, but their inter-relationships are poorly understood. We systematically mapped molecular pathways underlying the toxicity of alpha-synuclein (α-syn), a protein central to Parkinson's disease. Genome-wide screens in yeast identified 332 genes that impact α-syn toxicity. To “humanize” this molecular network, we developed a computational method, TransposeNet. This integrates a Steiner prize-collecting approach with homology assignment through sequence, structure, and interaction topology. TransposeNet linked α-syn to multiple parkinsonism genes and druggable targets through perturbed protein trafficking and ER quality control as well as mRNA metabolism and translation. A calcium signaling hub linked these processes to perturbed mitochondrial quality control and function, metal ion transport, transcriptional regulation, and signal transduction. Parkinsonism gene interaction profiles spatially opposed in the network (ATP13A2/PARK9 and VPS35/PARK17) were highly distinct, and network relationships for specific genes (LRRK2/PARK8, ATXN2, and EIF4G1/PARK18) were confirmed in patient induced pluripotent stem cell (iPSC)-derived neurons. This cross-species platform connected diverse neurodegenerative genes to proteinopathy through specific mechanisms and may facilitate patient stratification for targeted therapy. Keywords: alpha-synuclein; iPS cell; Parkinson’s disease; stem cell; mRNA translation; RNA-binding protein; LRRK2; VPS35; vesicle trafficking; yeas

A Timescale for Evolution, Population Expansion, and Spatial Spread of an Emerging Clone of Methicillin-Resistant Staphylococcus aureus

Due to the lack of fossil evidence, the timescales of bacterial evolution are largely unknown. The speed with which genetic change accumulates in populations of pathogenic bacteria, however, is a key parameter that is crucial for understanding the emergence of traits such as increased virulence or antibiotic resistance, together with the forces driving pathogen spread. Methicillin-resistant Staphylococcus aureus (MRSA) is a common cause of hospital-acquired infections. We have investigated an MRSA strain (ST225) that is highly prevalent in hospitals in Central Europe. By using mutation discovery at 269 genetic loci (118,804 basepairs) within an international isolate collection, we ascertained extremely low diversity among European ST225 isolates, indicating that a recent population bottleneck had preceded the expansion of this clone. In contrast, US isolates were more divergent, suggesting they represent the ancestral population. While diversity was low, however, our results demonstrate that the short-term evolutionary rate in this natural population of MRSA resulted in the accumulation of measurable DNA sequence variation within two decades, which we could exploit to reconstruct its recent demographic history and the spatiotemporal dynamics of spread. By applying Bayesian coalescent methods on DNA sequences serially sampled through time, we estimated that ST225 had diverged since approximately 1990 (1987 to 1994), and that expansion of the European clade began in 1995 (1991 to 1999), several years before the new clone was recognized. Demographic analysis based on DNA sequence variation indicated a sharp increase of bacterial population size from 2001 to 2004, which is concordant with the reported prevalence of this strain in several European countries. A detailed ancestry-based reconstruction of the spatiotemporal dispersal dynamics suggested a pattern of frequent transmission of the ST225 clone among hospitals within Central Europe. In addition, comparative genomics indicated complex bacteriophage dynamics

Astrocyte layers in the mammalian cerebral cortex revealed by a single-cell in situ transcriptomic map.

Although the cerebral cortex is organized into six excitatory neuronal layers, it is unclear whether glial cells show distinct layering. In the present study, we developed a high-content pipeline, the large-area spatial transcriptomic (LaST) map, which can quantify single-cell gene expression in situ. Screening 46 candidate genes for astrocyte diversity across the mouse cortex, we identified superficial, mid and deep astrocyte identities in gradient layer patterns that were distinct from those of neurons. Astrocyte layer features, established in the early postnatal cortex, mostly persisted in adult mouse and human cortex. Single-cell RNA sequencing and spatial reconstruction analysis further confirmed the presence of astrocyte layers in the adult cortex. Satb2 and Reeler mutations that shifted neuronal post-mitotic development were sufficient to alter glial layering, indicating an instructive role for neuronal cues. Finally, astrocyte layer patterns diverged between mouse cortical regions. These findings indicate that excitatory neurons and astrocytes are organized into distinct lineage-associated laminae.The study was supported by the Paul G. Allen Foundation Distinguished Investigator Program (E.M.U. and D.H.R.), the Dr. Miriam and Sheldon G. Adelson Medical Research Foundation (D.H.R., D.G. and G. C.), BRAIN initiative (1U01 MH105991 to D.G.) and National Institute of Health (1R01 MH109912 to D.G.; P01NS08351 to D.H.R.), National Institute of Health Research and the European Union Seventh Framework (to P.H.), NINDS Informatics Center for Neurogenetics and Neurogenomics (P30 NS062691 to G.C.), Wellcome Trust core support (M.H., O.A.B.), European Research Council (281961 to M.G.H.), Fonds Wetenschappelijk Onderzoek (G066715N and 1523014N to M.G.H.), Stichting Alzheimer Onderzoek (S#16025 to M.G.H.) and VIB Institutional Support and Tech Watch funding (to M.G.H.), Howard Hughes Medical Institute and the Wellcome Trust (to D.H.R.)

Quantitative spatial transcriptomics of the developing brain

Developmental and cellular heterogeneity can be studied at the transcriptomic level using RNA sequencing methods, but techniques that quantitatively investigate the spatial dynamics of cell heterogeneity across large tissue areas at a single-cell level are lacking. Applying this capability to the developing brain would enable reconstruction of a full spatial transcriptomic map of neural heterogeneity and the discovery of novel regional neural subtypes with potential functional implications. Such capability would ultimately culminate in the creation of single cell-level tissue atlases with preserved cellular topology. My goal has been to combine single-molecule fluorescence in situ hybridisation (smFISH), automated confocal imaging and analytical tools to derive ‘Quantitative Spatial Transcriptomics (QST)’ for the spatiotemporal characterisation of gene expression across the developing mammalian forebrain. My thesis research has focused in two main areas: 1) Novel methodology for insights into mammalian cortex architecture: I have co-developed (with Dr Omer Bayraktar) an automated, multiplexed smFISH and imaging pipeline for screening brain-wide gene expression at cellular resolution, termed Large-area Spatial Transcriptomics (LaST). LaST allows “mapping back” of transcriptome data from single-cell and single-nuclei RNA sequencing for high-quality in situ validation in a regional qualitative and quantitative manner. In addition, this tool can uncover novel cell heterogeneity and discover unique cell identities based on precisely validated combinations of cellular gene expression. For example, the organisation of neurons into six distinct layers is a hallmark of the mammalian neocortex but it is not known if glial cells also possess any diversified laminar features. Using LaST, the single-neuron expression of layer markers was mapped across the mouse cerebral cortex and identified diversified glutamatergic neuron subclasses and their laminar distribution in the neocortex, including rare transcriptomic types. Moreover, applying LaST to cortical astrocytes, I identified molecular distinctions that were associated with three dorso-ventral astrocyte layers in the somatosensory cortex, which deviate from known layer organisation of glutamatergic neurons. These published results (Bayraktar, Bartels et al., 2020) identify a previously unrecognised spatial complexity to cortical architecture when considering combined patterns of neuron and astrocyte heterogeneity, and are likely to have functional implications. 2) High-resolution transcriptomic developmental biology of Cdkl5: I further developed LaST into QST for cell type-specific quantifications of a single subject gene and analysed the expression of Cyclin-dependent kinase-like 5 (Cdkl5) during forebrain development. CDKL5 mutations cause a severe human neurodevelopmental disorder called CDKL5 deficiency disorder (CDD) that is currently incurable. The expression of CDKL5 in developing neural cells, especially glia, is unclear, resulting in an incomplete understanding of the pathogenesis of the disease. Using QST, I quantified the spatiotemporal expression pattern of Cdkl5 mRNA in developing mouse brain and uncovered novel dynamic patterns of Cdkl5 enrichment. I discovered that Cdkl5 is initially enriched in upper layer neurons perinatally, followed by deep layer neurons during late postnatal development. Using QST and data derived from single-cell RNA sequencing, I found evidence that Cdkl5/CDKL5 is expressed by macroglia during early postnatal development and is enriched in distinct oligodendrocyte transcriptomic types in the mature mouse and human neocortex. An investigation of CDKL5 protein expression in mouse brain development and in human post-mortem tissue is yet to be fully characterised. These data, combined with future investigation of the function of CDKL5 in these cell populations, will improve our understanding of CDD pathogenesis and may uncover novel cellular targets for therapeutic intervention in CDD. In addition, the QST screen provides a quantitative reference map of physiological Cdkl5 expression levels during postnatal development that is instructive for future development of gene and protein replacement therapies. Combined, the novel spatial transcriptomics techniques developed in this thesis identified higher-order cerebral cortex organisation, and allowed characterisation of Cdkl5 gene expression, significantly advancing our understanding of mammalian neocortical development and Cdkl5 gene function therein.Wellcome (109142/Z/15/Z

Rape supportive cognition, sexual fantasies and implicit offence scripts: A comparison between high and low rape prone men

It is widely accepted that sexual fantasy can play a role in the aetiology of rape, and it has often been shown that dominant fantasies correlate positively with rape supportive attitudes in men with a proclivity to rape. Furthermore, it has been suggested that frequent use of deviant fantasy can lead to the creation of an implicit-offence script that, if acted out, would constitute a sexual offence. Thus, the aim of the present study was to provide support for the hypotheses that: (1) high rape prone men would report more rape supportive attitudes and dominant fantasies in contrast to men with a low rape proclivity; (2) dominant fantasies are influenced by rape supportive attitudes; and (3) high rape prone men will show more evidence of harbouring implicit-offence scripts. The results supported the first two hypotheses but not the third. Various explanations are offered for these results alongside a discussion of the potential limitations of the study and future research suggestions