Through receptor mediated transcytosis comparable to that of undifferentiated THP-1 cells

A less stringent definition of the DR3-type consensus sequence, i.e. considering sequences with a lower HOMER score, or inclusion of hits immediately proximal to the peak summit, or both, can considerably increase the DR3 rate of a dataset. The ranking of the cell types for their DR3 frequency was largely independent of the chosen threshold for the specificity of the DR3type sequence. However, this should not be taken as generally applicable, because it is largely the product of our systematic and uniform analysis. Moreover, even assuming a lower binding specificity of VDR-RXR heterodimers, on average less than 50% of VDR peaks contain a DR3-type binding site. This suggests that in every 1,252D3-responsive cell type VDR is found not only in classical VDR-RXR heterodimers binding DR3-type sequences but also in alternative protein-DNA complexes. The latter contain a larger variety of transcription factors, such as GABPA, JUN, NFY, PU.1 and STAT5. The selection for these alternative VDR partners may largely depend on the cell-specific expression levels of these proteins. In all ligand-stimulated samples the rate of DR3-type sequences is higher than in the respective unstimulated samples, i.e. the latter contain a higher rate of non-DR3-type VDR binding sites. Although also unliganded VDR is functional, for example actively repressing some target genes via the interaction with co-repressor proteins, a number of the VDR binding sites in unstimulated cells may serve as lower affinity nuclear storage sites from where the receptor, upon ligand stimulation, moves to the binding sites of activated genes as we already hypothesized earlier. In conclusion, a new VDR ChIP-seq dataset from M1-type macrophage-like cells was the initiation for performing the first meta-analysis of all publicly available VDR ChIP-seq datasets. In contrast to general expectations, only some 11.5% of all 23,409 VDR binding sites contain a DR3-type sequence. Moreover, the number of identified VDR binding sites inversely correlates with the percentage of DR3-type sequences found below the peak summits. This context allowed a better understanding of the VDR binding pattern in LPS-differentiated THP-1 cells, which with 1,318 VDR peaks is rather small, but shows for ligand-stimulated samples a DR3 rate of close to 50%.Despite decades of intensive research in the labs of academic institutions and the pharmaceutical industry, the blood-brain barrier has remained a significant hurdle for treatment of CNS diseases with growing unmet medical need. Limiting brain access to small, predominantly hydrophobic molecules, this barrier is especially insurmountable to therapeutic proteins. Although antibodies are in clinical development for several CNS indications, brain exposure of these potential medicines is low and may in many cases be insufficient for therapeutic efficacy. The most important physiological entry route for proteins into the brain.

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