Background Cell-to-cell propagation of α-synuclein (α-syn) aggregates is thought to contribute to the pathogenesis of Parkinson’s disease (PD) and underlie the spread of α-syn neuropathology. Increased pro-inflammatory cytokine levels and activated microglia are present in PD and activated microglia can promote α-syn aggregation. However, it is unclear how microglia influence α-syn cell-to-cell transfer. Methods We developed a clinically relevant mouse model to monitor α-syn prion-like propagation between cells; we transplanted wild-type mouse embryonic midbrain neurons into a mouse striatum overexpressing human α-syn (huα-syn) following adeno-associated viral injection into the substantia nigra. In this system, we depleted or activated microglial cells and determined the effects on the transfer of huα-syn from host nigrostriatal neurons into the implanted dopaminergic neurons, using the presence of huα-syn within the grafted cells as a readout. Results First, we compared α-syn cell-to-cell transfer between host mice with a normal number of microglia to mice in which we had pharmacologically ablated 80% of the microglia from the grafted striatum. With fewer host microglia, we observed increased accumulation of huα-syn in grafted dopaminergic neurons. Second, we assessed the transfer of α-syn into grafted neurons in the context of microglia activated by one of two stimuli, lipopolysaccharide (LPS) or interleukin-4 (IL-4). LPS exposure led to a strong activation of microglial cells (as determined by microglia morphology, cytokine production and an upregulation in genes involved in the inflammatory response in the LPS-injected mice by RNA sequencing analysis). LPS-injected mice had significantly higher amounts of huα-syn in grafted neurons. In contrast, injection of IL-4 did not change the proportion of grafted dopamine neurons that contained huα-syn relative to controls. As expected, RNA sequencing analysis on striatal tissue revealed differential gene expression between LPS and IL-4-injected mice; with the genes upregulated in tissue from mice injected with LPS including several of those involved in an inflammatory response. Conclusions The absence or the hyperstimulation of microglia affected α-syn transfer in the brain. Our results suggest that under resting, non-inflammatory conditions, microglia modulate the transfer of α-syn. Pharmacological regulation of neuroinflammation could represent a future avenue for limiting the spread of PD neuropathology. Electronic supplementary material The online version of this article (10.1186/s13024-019-0335-3) contains supplementary material, which is available to authorized users.

The assembly of Ag receptor genes by V(D)J recombination is regulated by transcriptional promoters and enhancers which control chromatin accessibility at Ig and TCR gene segments to the RAG-1/RAG-2 recombinase complex. Paradoxically, germline deletions of the IgH enhancer (Eμ) only modestly reduce DH→JH rearrangements when assessed in peripheral B cells. However, deletion of Eμ severely impairs recombination of VH gene segments, which are located over 100 kb away. We now test two alternative explanations for the minimal effect of Eμ deletions on primary DH→JH rearrangement: 1) Accessibility at the DHJH cluster is controlled by a redundant cis-element in the absence of Eμ. One candidate for this element lies 5′ to DQ52 (PDQ52) and exhibits promoter/enhancer activity in pre-B cells. 2) In contrast to endpoint B cells, DH→JH recombination may be significantly impaired in pro-B cells from enhancer-deficient mice. To elucidate the roles of PDQ52 and Eμ in the regulation of IgH locus accessibility, we generated mice with targeted deletions of these elements. We report that the defined PDQ52 promoter is dispensable for germline transcription and recombination of the DHJH cluster. In contrast, we demonstrate that Eμ directly regulates accessibility of the DHJH region. These findings reveal a significant role for Eμ in the control mechanisms that activate IgH gene assembly and suggest that impaired VH→DHJH rearrangement in enhancer-deficient cells may be a downstream consequence of the primary block in DH→JH recombination.

Antigen receptor gene assembly is governed by transcriptional promoters and enhancers that communicate over large distances and modulate chromatin accessibility to V(D)J recombinase. The precise role of these cis-acting elements in opening chromatin at recombinase targets and the mechanisms underlying their crosstalk remain unclear. We show that the TCRbeta enhancer (Ebeta) directs long-range chromatin opening over both DbetaJbeta clusters. Strikingly, chromatin associated with the Dbeta1 gene segment is refractory to Ebeta-mediated opening. Accessibility at Dbeta1 is accompanied by the formation of a stable holocomplex between a Dbeta-proximal promoter and Ebeta. These findings indicate a stepwise process for Dbeta --> Jbeta recombination that relies on distinct aspects of Ebeta activity: an intrinsic function that directs general chromatin opening and a cooperative function that facilitates the assembly of a promoter/enhancer holocomplex, unmasks the Dbeta1 gene segment, and triggers TCRbeta gene assembly.

The assembly of genes encoding antigen receptors is regulated by developmental changes in chromatin that either permit or deny access to a single variable-(diversity)-joining recombinase. These changes are guided by transcriptional promoters and enhancers, which serve as accessibility-control elements in antigen-receptor loci. The function of each accessibility-control element and the factors they recruit to remodel chromatin remain obscure. Here we show that the recruitment of SWI-SNF chromatin-remodeling complexes compensated for the accessibility-control element function of a promoter but not an enhancer of the T cell receptor-beta locus (Tcrb). Loss of SWI-SNF function in thymocytes inactivated recombinase targets at the endogenous Tcrb locus. Thus, initiation of Tcrb gene assembly and T cell development is contingent on the recruitment of SWI-SNF to promoters, which exposes gene segments to variable-(diversity)-joining recombinase.

SummaryLeishmania major and all other parasitic protozoa are unable to synthesize purines de novo and are therefore reliant upon uptake of preformed purines from their hosts via nucleobase and nucleoside transporters. L. major expresses two nucleobase permeases, NT3 that is a high affinity transporter for purine nucleobases and NT4 that is a low affinity transporter for adenine. nt3 (-/-) null mutant promastigotes were unable to replicate in medium containing 10 mM hypoxanthine, guanine, or xanthine and replicated slowly in 10 mM adenine due to residual low affinity uptake of that purine. The NT3 transporter mediated the uptake of the anti-leishmanial drug allopurinol, and the nt3 (-/-) mutants were resistant to killing by this drug. Expression of the NT3 permease was profoundly downregulated at the protein but not the mRNA level in stationary phase compared with logarithmic phase promastigotes. The nt4 (-/-) null mutant was quantitatively impaired in survival within murine bone marrow-derived macrophages. Extensive efforts to generate an nt3 (-/-) /nt4 (-/-) dual null mutant were not successful, suggesting that one of the two nucleobase permeases must be retained for robust growth of the parasite. The phenotypes of these null mutants underscore the importance of purine nucleobase transporters in the Leishmania life cycle and pharmacology.