A proteomics analysis of 5xFAD mouse brain regions reveals the lysosome-associated protein Arl8b as a candidate biomarker for Alzheimer’s disease

BACKGROUND: Alzheimer's disease (AD) is characterized by the intra- and extracellular accumulation of amyloid-ß (Aß) peptides. How Aß aggregates perturb the proteome in brains of patients and AD transgenic mouse models, remains largely unclear. State-of-the-art mass spectrometry (MS) methods can comprehensively detect proteomic alterations, providing relevant insights unobtainable with transcriptomics investigations. Analyses of the relationship between progressive Aß aggregation and protein abundance changes in brains of 5xFAD transgenic mice have not been reported previously. METHODS: We quantified progressive Aß aggregation in hippocampus and cortex of 5xFAD mice and controls with immunohistochemistry and membrane filter assays. Protein changes in different mouse tissues were analyzed by MS-based proteomics using label-free quantification; resulting MS data were processed using an established pipeline. Results were contrasted with existing proteomic data sets from postmortem AD patient brains. Finally, abundance changes in the candidate marker Arl8b were validated in cerebrospinal fluid (CSF) from AD patients and controls using ELISAs. RESULTS: Experiments revealed faster accumulation of Aß42 peptides in hippocampus than in cortex of 5xFAD mice, with more protein abundance changes in hippocampus, indicating that Aß42 aggregate deposition is associated with brain region-specific proteome perturbations. Generating time-resolved data sets, we defined Aß aggregate-correlated and anticorrelated proteome changes, a fraction of which was conserved in postmortem AD patient brain tissue, suggesting that proteome changes in 5xFAD mice mimic disease-relevant changes in human AD. We detected a positive correlation between Aß42 aggregate deposition in the hippocampus of 5xFAD mice and the abundance of the lysosome-associated small GTPase Arl8b, which accumulated together with axonal lysosomal membranes in close proximity of extracellular Aß plaques in 5xFAD brains. Abnormal aggregation of Arl8b was observed in human AD brain tissue. Arl8b protein levels were significantly increased in CSF of AD patients. CONCLUSIONS: We report a comprehensive biochemical and proteomic investigation of hippocampal and cortical brain tissue derived from 5xFAD transgenic mice, providing a valuable resource to the neuroscientific community. We identified Arl8b, with significant abundance changes in 5xFAD and AD patient brains. Arl8b might enable the measurement of progressive lysosome accumulation in AD patients and have clinical utility as a candidate biomarker

Cloning and expression analysis of the chick ortholog of TBX22, the gene mutated in X-linked cleft palate and ankyloglossia

T-box genes constitute a conserved gene family with important roles in many developmental processes. Several family members have been implicated in human congenital diseases. Recently, mutations in TBX22 were found to cause X-linked cleft palate (CPX and ankyloglossia), a semidominant X-linked disorder affecting formation of the secondary palate. Here, we have cloned the chick ortholog of human TBX22 and have analyzed its expression during embryogenesis. Expression is very prominent in the somites and ill the myotome, and in the mandible and maxilla of the developing jaw. Other sites of expression include the limbs, the cranial mesenchyme and the eye. Hence, Tbx22 expression domains encompass the regions important for the development of the disease phenotype. (C) 2002 Elsevier Science Ireland Ltd. All rights reserved

tom-1, a novel v-Myb target gene expressed in AMV- and E26-transformed myelomonocytic cells.

The retroviral oncogene v-myb is a mutated and truncated version of the c-myb proto-oncogene and encodes a transcription factor (v-Myb) that specifically transforms myelomonocytic cells. Two different variants of v-myb, transduced independently by the oncogenic chicken retroviruses AMV and E26, have been characterized. It is believed that both variants of v-Myb transform myelomonocytic cells by affecting the expression of specific genes; however, no target genes common to both oncogenic viruses have been identified. Here, we describe the identification of a novel v-Myb target gene, designated as tom-1 (target of myb 1). The tom-1 gene has two promoters, one of which is Myb-inducible. tom-1 is expressed at elevated levels in AMV-transformed as well as in E26-transformed myeloid cells. We show that tom-1 activation by v-Myb does not require de novo protein synthesis and that the Myb-inducible tom-1 promoter contains a functional Myb binding site. Thus, tom-1 is the first example of a direct target gene for both oncogenic forms of the v-myb gene. Further analysis of the Myb-inducible tom-1 promoter shows that a C/EBP binding site is juxtaposed to the Myb binding site and that C/EBP is required for the Myb-dependent activation of the promoter. Together with previous work our results suggest that C/EBP may be a general cooperation partner for v-Myb in myelomonocytic cells

The T-box transcription factor Tbx15 is required for skeletal development

10.1016/j.mod.2004.10.011MECHANISMS OF DEVELOPMENT1222131-14

Overuse of corticosteroids in patients with immune thrombocytopenia (ITP) between 2011 and 2017 in the United States

Abstract Corticosteroids (CSs) are standard first‐line therapy for immune thrombocytopenia (ITP). Prolonged exposure is associated with substantial toxicity; thus guidelines recommend avoidance of prolonged CS treatment and early use of second‐line therapies. However, real‐world evidence on ITP treatment patterns remains limited. We aimed to assess real‐world treatment patterns in patients with newly‐diagnosed ITP, using two large US healthcare databases (Explorys and MarketScan) between January 1, 2011 and July 31, 2017. Adults with ITP, ≥12 months of database registration prior to diagnosis, ≥1 ITP treatment, and ≥1 month enrollment following initiation of first ITP treatment were included (n = 4066 Explorys; n = 7837 MarketScan). Information on lines of treatment (LoTs) was collected. As expected, CSs were the most common first‐line treatment (Explorys, 87.9%; MarketScan, 84.5%). However, CSs remained by far the most common treatment (Explorys ≥77%; MarketScan ≥85%) across all subsequent LoTs. Second‐line treatments such as rituximab (12.0% Explorys; 24.5% MarketScan), thrombopoietin receptor agonists (11.3% Explorys; 15.6% MarketScan), and splenectomy (2.5% Explorys; 8.1% MarketScan) were used much less frequently. CS use is widespread in the US in patients with ITP across all LoTs. Quality improvement initiatives are needed to reduce CS exposure and bolster use of second‐line treatments

Identification of VCP/p97, CHIP and amphiphysin II interaction partners using membrane-based human proteome arrays

Proteins mediate their biological function through interactions with other proteins. Therefore, the systematic identification and characterization of protein-protein interactions have become a powerful proteomic strategy to understand protein function and comprehensive cellular regulatory networks. For the screening of valosin-containing protein, carboxyl terminus of Hsp70-interacting protein (CHIP), and amphiphysin II interaction partners, we utilized a membrane-based array technology that allows the identification of human protein-protein interactions with crude bacterial cell extracts. Many novel interaction pairs such as valosin-containing protein/autocrine motility factor receptor, CHIP/caytaxin, or amphiphysin II/DLP4 were identified and subsequently confirmed by pull-down, two-hybrid and co-immunoprecipitation experiments. In addition, assays were performed to validate the interactions functionally. CHIP e.g. was found to efficiently polyubiquitinate caytaxin in vitro, suggesting that it might influence caytaxin degradation in vivo. Using peptide arrays, we also identified the binding motifs in the proteins DLP4, XRCC4, and fructose-1,6-bisphosphatase, which are crucial for the association with the Src homology 3 domain of amphiphysin II. Together these studies indicate that our human proteome array technology permits the identification of protein-protein interactions that are functionally involved in neurodegenerative disease processes, the degradation of protein substrates, and the transport of membrane vesicles