Post-Aire Maturation of Thymic Medullary Epithelial Cells Involves Selective Expression of Keratinocyte-Specific Autoantigens

The autoimmune regulator (Aire)-directed ectopic expression of tissue-specific antigens (TSAs) by mature medullary thymic epithelial cells (mTECs) has been viewed as an essential mechanism in the induction of central tolerance. Recent data suggest that the survival of mTECs extends beyond the Aire+ cell population to form the post-Aire mTEC population and Hassall’s corpuscles (HCs). The nature and function of these post-Aire epithelial cells and structures, however, have remained unidentified. In this study, we characterized in detail the end-stage development of mTECs and HCs in both Aire-sufficient and Aire-deficient mice. In addition, using a transgenic mouse model in which the LacZ reporter gene is under the control of the endogenous Aire promoter, we purified and analyzed the post-Aire mTECs to characterize their function. We showed that the end-stage maturation of mTECs closely resembles that of keratinocytes and that the lack of Aire results in a marked block of mTEC differentiation, which is partially overcome by ligands for RANK and CD40. We also provide evidence that, during mTEC development, Aire is expressed only once and during a limited 1–2 day period. The following loss of Aire expression is accompanied by a quick downregulation of MHC class II and CD80, and of most of the Aire-dependent and Aire-independent TSAs, with the exception of keratinocyte-specific genes. In the final stage of maturation, the mTECs lose their nuclei to become HCs and specifically express desmogleins (DGs) 1 and 3, which, via cross-presentation by APCs, may contribute to tolerance against these pemphigus vulgaris-related TSAs

The small ethylene response factor ERF96 is involved in the regulation of the abscisic acid response in Arabidopsis

Ethylene regulates many aspects of plant growth and development including seed germination, leaf senescence, and fruit ripening, and of plant responses to environmental stimuli including both biotic and abiotic stresses. Ethylene Response Factors (ERFs) are plant-specific transcription factors and are a subfamily of the AP2 (APETALA2)/ERF transcription factor family. The function of many members in this large gene family remains largely unknown. ERF96, a member of the Group IX ERF family transcription factors, has recently been shown to be a transcriptional activator that is involved in plant defense response in Arabidopsis. Here we provide evidence that ERF96 is a positive regulator of abscisic acid (ABA) responses. Bioinformatics analysis indicated that there are a total four small ERFs in Arabidopsis including ERF95, ERF96, ERF97 and ERF98, and that ERF96 forms a cluster with ERF95 and ERF97. By using quantitative RT-PCR, we found that ERF96 is expressed in all tissues and organs examined except roots, with relatively high expression in flowers and seeds. Results from the protoplast transfection assay results indicated that the EDLL motif-containing C-terminal domain is responsible for ERF96’s transcriptional activity. Although loss-of-function mutant of ERF96 was morphologically similar to wild type plants, transgenic plants overexpressing ERF96 had smaller rosette size and were delayed in flowering time. In ABA sensitivity assays, we found that ERF96 overexpression plants were hypersensitive to ABA in terms of ABA inhibition of seed germination, early seedling development and root elongation. Consistent with these observations, elevated transcript levels of some ABA-responsive genes including RD29A, ABI5, ABF3, ABF4, P5CS and COR15A were observed in the transgenic plants in the presence of ABA. However, in the absence of ABA treatment, the transcript levels of these ABA-responsive genes remained largely unchanged. Our experiments also showed that water loss in ERF96 overexpression plants was slower than that in Col wild type plants. Stomatal closure assays indicated that ERF96 overexpression plants had reduced stomatal aperture in the presence of ABA. Taken together, our results suggest that AtERF96 positively regulates ABA responses in Arabidopsis

Lower bone mineral density in Patients with Parkinson’s disease: a cross-sectional study from Chinese Mainland

Objectives Although several lines of evidence have suggested that patients with Parkinson’s disease (PD) have a higher risk of osteoporosis and fracture, the association between bone mineral density and severity of PD patients is unknown. Methods We performed a cross-sectional study of 54 patients with PD and 59 healthy age-matched controls. Multiple clinical scales were used to evaluate the severity of PD, and serum levels of calcium, phosphorus, and homocysteine were measured to determine bone mineral density’s association with PD severity. Results BMD in PD patients was significantly lower than in healthy controls. The BMD scores of the spine, femoral neck, and hip were lower in females than in males in the healthy group. In the PD group, BMD in the hip was significantly lower in females compared to males. There was a negative correlation between daily L-dopa dosage and BMD in the spine and hip in the PD group, while BMD in the spine, neck, and hip was significantly correlated with severity of PD. Besides, we found that among the lumbar spine, femoral neck and hip, bone loss in the lumbar spine was the most severe in PD patients based on the T-scores. Conclusions Our findings support the hypothesis that patients with PD have a higher risk of osteoporosis, and that low BMD in the spine, femoral neck, and hip may indirectly reflect the severity of PD. Our findings have prompted us to pay more attention to osteoporosis in the lumbar spine in Chinese PD patients

Identification and characterization of a cleavage site in the proteolysis of orf virus 086 protein

The ORF virus (ORFV) is among the parapoxvirus genus of the poxviridae family, but little is known about the proteolytic pathways of ORFV encoding proteins. By contrast, the proteolysis mechanism of the vaccinia virus has been extensively explored. Vaccinia virus core protein P4a undergoes a proteolytic process that takes place at a conserved cleavage site Ala-Gly-X (where X is any amino acid) and participates in virus assembly. Bioinformatics analysis revealed that an ORFV encoding protein, ORFV086, has a similar structure to the Vaccinia virus P4a core protein. In this study, we focus on the kinetic analysis and proteolysis mechanism of ORFV086. We found, via kinetic analysis, that ORFV086 is a late gene that starts to express at 8 hours post infection at mRNA level and 12 to 24 hours post infection at the protein level. The ORFV086 precursor and a 21kDa fragment can be observed in mature ORFV virions. The same bands were detected at only 3 hours post infection, suggesting that both the ORFV086 precursor and the 21kDa fragment are viral structural proteins. ORFV086 was cleaved from 12 to 24 hours post infection. The cleavage took place at different sites，resulting in seven bands with differing molecular weights. Sequence alignment revealed that five putative cleavage sites were predicted at C-terminal and internal regions of ORFV086. To investigate whether those cleavage sites are involved in proteolytic processing, full length and several deletion mutant ORFV086 recombinant proteins were expressed and probed. The GGS site that produced a 21kDa cleavage fragment was confirmed by identification of N/C-terminal FLAG epitope recombinant proteins, site-directed mutagenesis and Pulse-chase analysis. Interestingly, chase results demonstrated that, at late times, ORFV086 is partially cleaved. Taken together, we concluded that GGS is a cleavage site in ORFV086 and produces a 21kDa fragment post infection. Both ORFV086 precursor and the 21kDa fragment are structural proteins of mature ORFV virions. ORFV086 and its cleaved products are indispensable for correct assembly of mature viral particles and this proteolytic processing of ORFV086 may play an essential role in viral morphogenic transition