Molecular correlates of trait anxiety: expanding biomarker discovery from protein expression to turnover

Depression and anxiety disorders affect a great number of people in the world. Although remarkable efforts have been devoted to understanding the clinical and biological basis of these disorders, progress has been relatively slow. Furthermore, no laboratory test currently is available for diagnosis of anxiety and depression. These disorders are mainly diagnosed empirically on the basis of a doctor’s personal observations and experiences. Hence, discovery of biomarkers for these psychiatric disorders deserves much scientific attention. The animal models investigated in the present study represent high, low, and normal anxiety-like phenotypes (HAB, LAB, NAB) and were established by selective inbreeding. To compare the protein expression levels between different animal lines, living animals were metabolically labeled with the 15N stable isotope and then investigated by quantitative mass spectrometry. In addition, metabolomic studies were performed to shed light on pathways affected in the trait anxiety mouse model. A number of proteins and metabolites were found to be significantly altered in their expression levels between the three mouse lines. Both protein and metabolite information was used for in silico network analysis to find pathways pertinent to the pathobiology of anxiety disorders. Another focus of this thesis was the development of new methodologies for the metabolic labeling approach. This includes improved identification of labeled proteins and the analysis of protein turnover. The latter represents another important aspect in the field of proteomics and adds a dynamic dimension to the field. The method allows the detection of protein expression alterations at a much earlier stage. The newly developed ProTurnyer (Protein Turnover Analyzer) algorithm is able to calculate in a high throughput manner turnover for individual proteins

Semantic-Aware Frame-Event Fusion based Pattern Recognition via Large Vision-Language Models

Pattern recognition through the fusion of RGB frames and Event streams has emerged as a novel research area in recent years. Current methods typically employ backbone networks to individually extract the features of RGB frames and event streams, and subsequently fuse these features for pattern recognition. However, we posit that these methods may suffer from key issues like sematic gaps and small-scale backbone networks. In this study, we introduce a novel pattern recognition framework that consolidates the semantic labels, RGB frames, and event streams, leveraging pre-trained large-scale vision-language models. Specifically, given the input RGB frames, event streams, and all the predefined semantic labels, we employ a pre-trained large-scale vision model (CLIP vision encoder) to extract the RGB and event features. To handle the semantic labels, we initially convert them into language descriptions through prompt engineering, and then obtain the semantic features using the pre-trained large-scale language model (CLIP text encoder). Subsequently, we integrate the RGB/Event features and semantic features using multimodal Transformer networks. The resulting frame and event tokens are further amplified using self-attention layers. Concurrently, we propose to enhance the interactions between text tokens and RGB/Event tokens via cross-attention. Finally, we consolidate all three modalities using self-attention and feed-forward layers for recognition. Comprehensive experiments on the HARDVS and PokerEvent datasets fully substantiate the efficacy of our proposed SAFE model. The source code will be made available at https://github.com/Event-AHU/SAFE_LargeVLM.Comment: In Peer Revie

CKS Proteins Promote Checkpoint Recovery by Stimulating Phosphorylation of Treslin

CKS proteins are small (9-kDa) polypeptides that bind to a subset of the cyclin-dependent kinases. The two paralogs expressed in mammals, Cks1 and Cks2, share an overlapping function that is essential for early development. However, both proteins are frequently overexpressed in human malignancy. It has been shown that CKS protein overexpression overrides the replication stress checkpoint, promoting continued origin firing. This finding has led to the proposal that CKS protein-dependent checkpoint override allows premalignant cells to evade oncogene stress barriers, providing a causal link to oncogenesis. Here, we provide mechanistic insight into how overexpression of CKS proteins promotes override of the replication stress checkpoint. We show that CKS proteins greatly enhance the ability of Cdk2 to phosphorylate the key replication initiation protein treslin in vitro. Furthermore, stimulation of treslin phosphorylation does not occur by the canonical adapter mechanism demonstrated for other substrates, as cyclin-dependent kinase (CDK) binding-defective mutants are capable of stimulating treslin phosphorylation. This effect is recapitulated in vivo, where silencing of Cks1 and Cks2 decreases treslin phosphorylation, and overexpression of wild-type or CDK binding-defective Cks2 prevents checkpoint-dependent dephosphorylation of treslin. Finally, we provide evidence that the role of CKS protein-dependent checkpoint override involves recovery from checkpoint-mediated arrest of DNA replication

Nicotinamide mononucleotide adenylyltransferase uses its NAD+ substrate-binding site to chaperone phosphorylated Tau.

Funder: Science and Technology Commission of Shanghai Municipality; FundRef: http://dx.doi.org/10.13039/501100003399Funder: Dr. John T. MacDonald Foundation; FundRef: http://dx.doi.org/10.13039/100010239Tau hyper-phosphorylation and deposition into neurofibrillary tangles have been found in brains of patients with Alzheimer's disease (AD) and other tauopathies. Molecular chaperones are involved in regulating the pathological aggregation of phosphorylated Tau (pTau) and modulating disease progression. Here, we report that nicotinamide mononucleotide adenylyltransferase (NMNAT), a well-known NAD+ synthase, serves as a chaperone of pTau to prevent its amyloid aggregation in vitro as well as mitigate its pathology in a fly tauopathy model. By combining NMR spectroscopy, crystallography, single-molecule and computational approaches, we revealed that NMNAT adopts its enzymatic pocket to specifically bind the phosphorylated sites of pTau, which can be competitively disrupted by the enzymatic substrates of NMNAT. Moreover, we found that NMNAT serves as a co-chaperone of Hsp90 for the specific recognition of pTau over Tau. Our work uncovers a dedicated chaperone of pTau and suggests NMNAT as a key node between NAD+ metabolism and Tau homeostasis in aging and neurodegeneration

Differential sensitivity of target genes to translational repression by miR-17~92

MicroRNAs (miRNAs) are thought to exert their functions by modulating the expression of hundreds of target genes and each to a small degree, but it remains unclear how small changes in hundreds of target genes are translated into the specific function of a miRNA. Here, we conducted an integrated analysis of transcriptome and translatome of primary B cells from mutant mice expressing miR-17~92 at three different levels to address this issue. We found that target genes exhibit differential sensitivity to miRNA suppression and that only a small fraction of target genes are actually suppressed by a given concentration of miRNA under physiological conditions. Transgenic expression and deletion of the same miRNA gene regulate largely distinct sets of target genes. miR-17~92 controls target gene expression mainly through translational repression and 5’UTR plays an important role in regulating target gene sensitivity to miRNA suppression. These findings provide molecular insights into a model in which miRNAs exert their specific functions through a small number of key target genesCX is a Pew Scholar in Biomedical Sciences. This study is supported by the PEW Charitable Trusts, Cancer Research Institute, National Institute of Health (R01AI087634, R01AI089854, RC1CA146299, R56AI110403, and R01AI121155 to CX), National Natural Science Foundation of China (31570882 to WHL, 31570883 to NX, 31570911 to GF, 91429301 to JH, 31671428 and 31500665 to YZ), 1000 Young Talents Program of China (K08008 to NX), 100 Talents Program of The Chinese Academy of Sciences (YZ), National Program on Key Basic Research Project of China (2016YFA0501900 to YZ), the Fundamental Research Funds for the Central Universities of China (20720150065 to NX and GF), Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Science, ICT & Future Planning (NRF-2015R1C1A1A01052387 to SGK, NRF-2016R1A4A1010115 to SGK and PHK), and 2016 Research Grant from Kangwon National University (SGK)

Stable Isotope Metabolic Labeling with a Novel 15N-Enriched Bacteria Diet for Improved Proteomic Analyses of Mouse Models for Psychopathologies

The identification of differentially regulated proteins in animal models of psychiatric diseases is essential for a comprehensive analysis of associated psychopathological processes. Mass spectrometry is the most relevant method for analyzing differences in protein expression of tissue and body fluid proteomes. However, standardization of sample handling and sample-to-sample variability are problematic. Stable isotope metabolic labeling of a proteome represents the gold standard for quantitative mass spectrometry analysis. The simultaneous processing of a mixture of labeled and unlabeled samples allows a sensitive and accurate comparative analysis between the respective proteomes. Here, we describe a cost-effective feeding protocol based on a newly developed 15N bacteria diet based on Ralstonia eutropha protein, which was applied to a mouse model for trait anxiety. Tissue from 15N-labeled vs. 14N-unlabeled mice was examined by mass spectrometry and differences in the expression of glyoxalase-1 (GLO1) and histidine triad nucleotide binding protein 2 (Hint2) proteins were correlated with the animals' psychopathological behaviors for methodological validation and proof of concept, respectively. Additionally, phenotyping unraveled an antidepressant-like effect of the incorporation of the stable isotope 15N into the proteome of highly anxious mice. This novel phenomenon is of considerable relevance to the metabolic labeling method and could provide an opportunity for the discovery of candidate proteins involved in depression-like behavior. The newly developed 15N bacteria diet provides researchers a novel tool to discover disease-relevant protein expression differences in mouse models using quantitative mass spectrometry

Degradation of HK2 by chaperone-mediated autophagy promotes metabolic catastrophe and cell death

Hexokinase II (HK2), a key enzyme involved in glucose metabolism, is regulated by growth factor signaling and is required for initiation and maintenance of tumors. Here we show that metabolic stress triggered by perturbation of receptor tyrosine kinase FLT3 in non–acute myeloid leukemia cells sensitizes cancer cells to autophagy inhibition and leads to excessive activation of chaperone-mediated autophagy (CMA). Our data demonstrate that FLT3 is an important sensor of cellular nutritional state and elucidate the role and molecular mechanism of CMA in metabolic regulation and mediating cancer cell death. Importantly, our proteome analysis revealed that HK2 is a CMA substrate and that its degradation by CMA is regulated by glucose availability. We reveal a new mechanism by which excessive activation of CMA may be exploited pharmacologically to eliminate cancer cells by inhibiting both FLT3 and autophagy. Our study delineates a novel pharmacological strategy to promote the degradation of HK2 in cancer cells

A critical review of microplastic pollution in urban freshwater environments and legislative progress in China: Recommendations and insights

Freshwater systems are vitally important, supporting diversity and providing a range of ecosystem services. In China, rapid urbanization (over 800 million urban population) has led to multiple anthropogenic pressures that threaten urban freshwater environments. Microplastics (<5 mm) result from intensive production and use of plastic materials, but their effects in urban freshwater environments remain poorly understood. Rising concerns over the ecological effects of microplastics have resulted in increased attention being given to this contaminant in Chinese freshwater systems. Some studies provide quantitative data on contamination loads, but in general relevant knowledge in freshwater environment remains narrow in China, and lacking adequate understanding of threshold levels for detrimental effects. Notably, non-standardized sample collection and processing techniques for point and non-point sources have hindered comparisons of contamination loads and associated risk. Meanwhile, legislative frameworks for managing microplastics in China remain in their infancy. This manuscript critically reviews what is known of the nature and magnitude of microplastic pollution in Chinese freshwater environments, and summarizes relevant Chinese legislation. It provides recommendations for improving the legislative framework in China and identifies research gaps that need to be addressed to improve management and regulatory strategies for dealing with microplastic pollution in Chinese urban freshwater environments