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In birds, males are the homogametic sex (ZZ) and females the heterogametic sex (ZW). Primary sex determination is thought to depend on a sex chromosome gene dosage mechanism, and the most likely sex determinant is the Z chromosome gene Doublesex and Mab-3-Related Transcription factor 1 (DMRT1). To clarify this issue, we used a CRISPR-Cas9-based monoallelic targeting approach and sterile surrogate hosts to generate birds with targeted mutations in the DMRT1 gene. The resulting chromosomally male (ZZ) chicken with a single functional copy of DMRT1 developed ovaries in place of testes, demonstrating the avian sex-determining mechanism is based on DMRT1 dosage. These ZZ ovaries expressed typical female markers and showed clear evidence of follicular development. However, these ZZ adult birds with an ovary in place of testes were indistinguishable in appearance to wild-type adult males, supporting the concept of cell-autonomous sex identity (CASI) in birds. In experiments where estrogen synthesis was blocked in control ZW embryos, the resulting gonads developed as testes. In contrast, if estrogen synthesis was blocked in ZW embryos that lacked DMRT1, the gonads invariably adopted an ovarian fate. Our analysis shows that DMRT1 is the key sex determination switch in birds and that it is essential for testis development, but that production of estrogen is also a key factor in primary sex determination in chickens, and that this production is linked to DMRT1 expression.Synucleinopathies are neurodegenerative diseases with both central and peripheral immune responses. However, whether the peripheral immune changes occur early in disease and their relation to brain events is yet unclear. Isolated rapid-eye-movement (REM) sleep behavior disorder (iRBD) can precede synucleinopathy-related parkinsonism and provides a prodromal phenotype to study early Parkinson's disease events. In this prospective case-control study, we describe monocytic markers in a cohort of iRBD patients that were associated with the brain-imaging markers of inflammation and neuronal dysfunction. Using 11C-PK11195 positron emission tomography (PET), we previously showed increased immune activation in the substantia nigra of iRBD patients, while 18F-DOPA PET detected reduced putaminal dopaminergic function. Here we describe that patients' blood monocytic cells showed increased expression of CD11b, while HLA-DR expression was decreased compared to healthy controls. The iRBD patients had increased classical monocytes and mature natural killer cells. Remarkably, the levels of expression of Toll-like receptor 4 (TLR4) on blood monocytes in iRBD patients were positively correlated with nigral immune activation measured by 11C-PK11195 PET and negatively correlated with putaminal 18F-DOPA uptake; the opposite was seen for the percentage of CD163+ myeloid cells. This suggesting a deleterious role for TLR4 and, conversely, a protective one for the CD163 expression. We show an association between peripheral blood monocytes and brain immune and dopaminergic changes in a synucleinopathy-related disorder, thus suggesting a cross-talk among periphery and brain during the disease.Both the small and large subunits of the ribosome, the molecular machine that synthesizes proteins, are complexes of ribosomal RNAs (rRNAs) and a number of proteins. In bacteria, the small subunit has a single 16S rRNA whose folding is the first step in its assembly. The central domain of the 16S rRNA folds independently, driven either by Mg2+ ions or by interaction with ribosomal proteins. To provide a quantitative description of ion-induced folding of the ∼350-nucleotide rRNA, we carried out extensive coarse-grained molecular simulations spanning Mg2+ concentration between 0 and 30 mM. The Mg2+ dependence of the radius of gyration shows that globally the rRNA folds cooperatively. Surprisingly, various structural elements order at different Mg2+ concentrations, indicative of the heterogeneous assembly even within a single domain of the rRNA. selleck inhibitor Binding of Mg2+ ions is highly specific, with successive ion condensation resulting in nucleation of tertiary structures. We also predict the Mg2+-dependent protection factors, measurable in hydroxyl radical footprinting experiments, which corroborate the specificity of Mg2+-induced folding. The simulations, which agree quantitatively with several experiments on the folding of a three-way junction, show that its folding is preceded by formation of other tertiary contacts in the central junction. Our work provides a starting point in simulating the early events in the assembly of the small subunit of the ribosome.Cadherin transmembrane proteins are responsible for intercellular adhesion in all biological tissues and modulate tissue morphogenesis, cell motility, force transduction, and macromolecular transport. The protein-mediated adhesions consist of adhesive trans interactions and lateral cis interactions. Although theory suggests cooperativity between cis and trans bonds, direct experimental evidence of such cooperativity has not been demonstrated. Here, the use of superresolution microscopy, in conjunction with intermolecular single-molecule Förster resonance energy transfer, demonstrated the mutual cooperativity of cis and trans interactions. Results further demonstrate the consequent assembly of large intermembrane junctions, using a biomimetic lipid bilayer cell adhesion model. Notably, the presence of cis interactions resulted in a nearly 30-fold increase in trans-binding lifetimes between epithelial-cadherin extracellular domains. In turn, the presence of trans interactions increased the lifetime of cis bonds. Importantly, comparison of trans-binding lifetimes of small and large cadherin clusters suggests that this cooperativity is primarily due to allostery. The direct quantitative demonstration of strong mutual cooperativity between cis and trans interactions at intermembrane adhesions provides insights into the long-standing controversy of how weak cis and trans interactions act in concert to create strong macroscopic cell adhesions.Regulation of circadian behavior and physiology by the Drosophila brain clock requires communication from central clock neurons to downstream output regions, but the mechanism by which clock cells regulate downstream targets is not known. We show here that the pars intercerebralis (PI), previously identified as a target of the morning cells in the clock network, also receives input from evening cells. We determined that morning and evening clock neurons have time-of-day-dependent connectivity to the PI, which is regulated by specific peptides as well as by fast neurotransmitters. Interestingly, PI cells that secrete the peptide DH44, and control restactivity rhythms, are inhibited by clock inputs while insulin-producing cells (IPCs) are activated, indicating that the same clock cells can use different mechanisms to drive cycling in output neurons. Inputs of morning cells to IPCs are relevant for the circadian rhythm of feeding, reinforcing the role of the PI as a circadian relay that controls multiple behavioral outputs.
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