-
Mashup Score: 2Spatiotemporal proteomics reveals the biosynthetic lysosomal membrane protein interactome in neurons - 1 hour(s) ago
Lysosomes are membrane-bound organelles critical for maintaining cellular homeostasis. Delivery of biosynthetic lysosomal proteins to lysosomes is crucial to orchestrate proper lysosomal function. However, it remains unknown how the delivery of biosynthetic lysosomal proteins to lysosomes is ensured in neurons, which are highly polarized cells. Here, we developed Protein Origin, Trafficking And Targeting to Organelle Mapping (POTATOMap), by combining trafficking synchronization and proximity-labelling based proteomics, to unravel the trafficking routes and interactome of the biosynthetic lysosomal membrane protein LAMP1 at specified time points. This approach, combined with advanced microscopy, enabled us to identify the neuronal domain-specific trafficking machineries of biosynthetic LAMP1. We revealed a role in replenishing axonal lysosomes, in delivery of newly synthesized axonal synaptic proteins, and interactions with RNA granules to facilitate hitchhiking in the axon. POTATOMap o
Source: www.biorxiv.orgCategories: General Medicine News, General HCPsTweet
-
Mashup Score: 0Neuronal phenotype defined by transcriptome-wide bursting kinetics in pyramidal and fast-spiking cells - 2 hour(s) ago
Single-cell sequencing revealed the transcriptional heterogeneity of neurons and introduced transcriptome-defined neuron types and subtypes. Temporal recordings of gene transcription showed that mRNA synthesis is arranged into transcriptional bursts which can be characterized by kinetic parameters. Here, we selected two distinct, functionally homogenous, and electrophysiologically well-defined neuronal cell types (the pyramidal cells and fast-spiking interneurons) for transcriptional burst analysis. Hierarchical clustering based on electrophysiological parameters recovered these cell types. In contrast, clustering based on transcripts failed to differentiate these neuronal phenotypes. We applied mathematical models to analyze burst kinetics of fast-spiking and pyramidal cells to explore the transcriptional underpinning of their distinct physiological functions. We found that inferred burst kinetic parameters are distinct measures of gene expression from average mRNA counts. Based on ou
Source: www.biorxiv.orgCategories: General Medicine News, General HCPsTweet
-
Mashup Score: 0Combinatorial characterization of bacterial taxa-driven differences in the microbiome of oyster reefs - 3 hour(s) ago
Oyster reefs are invaluable ecosystems that provide a wide array of critical ecosystem services, including water filtration, coastal protection, and habitat provision for various marine species. However, these essential habitats face escalating threats from climate change and anthropogenic stressors. To combat these challenges, numerous oyster restoration initiatives have been undertaken, representing a global effort to preserve and restore these vital ecosystems. A significant, yet poorly understood, component of oyster reefs is the microbial communities. These communities account for a substantial proportion of marine reefs and are pivotal in driving key biogeochemical processes. Particularly, the environmental microbiome plays a crucial role in supporting the health and resilience of oyster populations. In our study, we sought to shed light on the microbiome within oyster reef ecosystems by characterizing the abundance, and diversity of microorganisms in the soil, biofilm, and oyste
Source: www.biorxiv.orgCategories: General Medicine News, General HCPsTweet
-
Mashup Score: 2Split minireporters facilitate monitoring of gene expression and peptide production in linear cell-free transcription-translation systems - 3 hour(s) ago
Cell-free transcription-translation (TXTL) systems expressing genes from linear dsDNA enable rapid prototyping of genetic devices while avoiding cloning steps. However, repetitive inclusion of a reporter gene is an incompressible cost and sometimes accounts for most of the synthesized DNA length. Here we present minireporters based on split-GFP systems that reassemble into functional fluorescent proteins and can be used to monitor gene expression in E. coli TXTL. The 135 bp GFP10-11 fragment produces a fluorescent signal comparable to its full-length GFP counterpart when reassembling with its complementary protein synthesized from the 535 bp fragment expressed in TXTL. We show that minireporters can be used to characterize promoter libraries, with data qualitatively comparable to full-length GFP, and matching with in vivo expression measurements. We also use minireporters as small fusion tags to measure TXTL protein and peptide production yield. Finally, we generalize our concept by pr
Source: www.biorxiv.orgCategories: General Medicine News, General HCPsTweet
-
Mashup Score: 2Flawed machine-learning confounds coding sequence annotation - 3 hour(s) ago
Detecting protein coding genes in genomic sequences is a significant challenge for understanding genome functionality, yet the reliability of bioinformatic tools for this task remains largely unverified. This is despite some of these tools having been available for several decades, and being widely used for genome and transcriptome annotation. We perform an assessment of nucleotide sequence and alignment-based de novo protein-coding detection tools. The controls we use exclude any previous training dataset and include coding exons as a positive set and length-matched intergenic and shuffled sequences as negative sets. Our work demonstrates that several widely used tools are neither accurate nor computationally efficient for the protein-coding sequence detection problem. In fact, just three of nine tools significantly outperformed a naive scoring scheme. Furthermore, we note a high discrepancy between self-reported accuracies and the accuracy achieved in our study. Our results show that
Source: www.biorxiv.orgCategories: General Medicine News, General HCPsTweet
-
Mashup Score: 1
In-solution hybridisation enrichment of genetic markers is a method of choice in paleogenomic studies, where the DNA of interest is generally heavily fragmented and contaminated with environmental DNA, and where the retrieval of genetic data comparable between individuals is challenging. Here, we benchmarked the commercial “Twist Ancient DNA” reagent from Twist Biosciences using sequencing libraries from ancestrally diverse ancient human samples with low to high endogenous DNA content (0.1-44%). For each library, we tested one and two rounds of enrichment, and assessed performance compared to deep shotgun sequencing. We find that the “Twist Ancient DNA” assay provides robust enrichment of ~1.2M target SNPs without introducing allelic bias that may interfere with downstream population genetics analyses. Additionally, we show that pooling up to 4 sequencing libraries and performing two rounds of enrichment is both reliable and cost-effective for libraries with less than 27% endogenous DN
Source: www.biorxiv.orgCategories: General Medicine News, General HCPsTweet
-
Mashup Score: 0
Contrary to the common belief, the most commonly used laboratory mouse inbred strains are shaped by a distinctive genetic and phenotypic diversity. In the past 10 years next generation sequencing unveiled a wide spectrum of genetic variants in different mouse inbred strains and the meticulous observation of researchers pointed to a variegate intra-and inter-strain phenotypic diversity. Although a genotype-phenotype correlation has been described for some traits, the relationship between several endophenotypes and causative genetic variability remains still unknown. Recently, we characterized the brain collateral plasticity in two brain ischemia C57BL/6J mouse models (i.e bilateral common carotid artery stenosis [BCCAS] and 60-min transient unilateral middle cerebral artery occlusion [MCAO]) and observed a Mendelian-like fashion of inheritance of the posterior communicating artery (PcomA) plasticity. Interestingly, a copy number variant (CNV) spanning Ide locus was reported to segregate
Source: www.biorxiv.orgCategories: General Medicine News, General HCPsTweet
-
Mashup Score: 0A phosphate-binding pocket in cyclin B3 is essential for XErp1/Emi2 degradation in meiosis I - 4 hour(s) ago
To ensure the correct euploid state of embryos, it is essential that vertebrate oocytes await fertilization arrested at metaphase of meiosis II. This MII arrest is mediated by XErp1/Emi2, which inhibits the ubiquitin ligase APC/C (anaphase-promoting complex/cyclosome). Cyclin B3 in complex with Cdk1 (cyclin-dependent kinase 1) is essential to prevent an untimely arrest of vertebrate oocytes in meiosis I by targeting XErp1/Emi2 for degradation. Yet, the molecular mechanism of XErp1/Emi2 degradation in MI is not well understood. Here, by combining TRIM-Away in oocytes with egg extract and in vitro studies, we demonstrate that a hitherto unknown phosphate-binding pocket in cyclin B3 is essential for efficient XErp1/Emi2 degradation in meiosis I. This pocket enables Cdk1/cyclin B3 to bind pre-phosphorylated XErp1/Emi2 facilitating further phosphorylation events, which ultimately target XErp1/Emi2 for degradation in a Plk1 (Polo-like kinase 1) dependent manner. Key elements of this degradat
Source: www.biorxiv.orgCategories: General Medicine News, General HCPsTweet
-
Mashup Score: 0Development and Characterization of Recombinant ADP-Ribose Binding Reagents that Allow Simultaneous Detection of Mono and Poly ADP-Ribose - 4 hour(s) ago
ADP-ribosylation (ADPRylation) is a post-translational modification (PTM) of proteins mediated by the activity of a variety of ADP-ribosyltransferase (ART) enzymes, such as the Poly(ADP-ribose) Polymerase (PARP) family of proteins. This PTM is diverse in both form and biological functions, which makes it a highly interesting modification, but difficult to study due to limitations in reagents available to detect the diversity of ADP-ribosylation. Recently we developed a set of recombinant antibody-like ADP-ribose binding proteins, using naturally occurring ADPR binding domains (ARBDs) that include macrodomains and WWE domains, that have been functionalized by fusion to the constant Fc region of rabbit immunoglobulin. Herein, we present an expansion of this biological toolkit, where we have replaced the rabbit Fc sequence with two other species, the Fc for mouse and goat immunogloblulin. Characterization of the new reagents indicates that they can be detected in a species-dependent manne
Source: www.biorxiv.orgCategories: General Medicine News, General HCPsTweet
-
Mashup Score: 0Deep dive into the diversity and properties of rhodopsins in actinomycetes of the family Geodermatophilaceae - 4 hour(s) ago
Rhodopsin with the NTQ motif and heliorhodopsins, both discovered for the first time in actinomycetes of the family Geodermatophilaceae were characterized, along with characterization of the DTEW, DTEF and NDQ rhodopsins reported in members of this family previously. The identified absorption maxima ranged from 513 to 559 nm, of which the blue-shifted rhodopsin contained the NTQ motif and the red-shifted one were heliorhodopsin. An assessment of the pumping activity of the NTQ rhodopsin and heliorhodopsin showed that the former functioned as inward Cl–, Br–, I–pump with much weaker activity towards NO3-, while no any activity was detected for the later. The DTEW and DTEF rhodopsins had the outward H+-transport activity which was dependent on the presence of Ca2+ ions and on the particular E. coli expression strain. The outward pumping of H+ was also detected for NDQ rhodopsin both in NaCl and KCl solutions at pH 5 and 6, but not at pH 7. Weak Na+-efflux activity for this rhodopsin w
Source: www.biorxiv.orgCategories: General Medicine News, General HCPsTweet
Spatiotemporal proteomics reveals the biosynthetic lysosomal membrane protein interactome in neurons https://t.co/MxtLyHCcNy #bioRxiv