Biology subjects

Shen, Z.

Publications and source records attributed to Shen, Z..

At least 127 records · Page 7Linked to original sources

bioRxiv2020

Cortical activity evoked by voice pitch changes: a combined fNIRS and EEG study

Numerous studies have investigated the cortical representation and time course of responses to auditory signals with and without pitch. However, little is known regarding the responses evoked by pitch changes. To study the spatial as well as temporal characteristics of cortical activity elicited by voice pitch changes, combined functional near-infrared spectroscopy (fNIRS) and electroencephalography (EEG) data were obtained from normal-hearing listeners presented with continuous vowel sequences in which the prosodic contours either varied between the individual vowels or were the same throughout. The fNIRS topographies and the EEG source reconstructions indicated additional right-hemispheric activity anterior to the primary auditory cortex and in the superior temporal sulcus for sequences with variable prosody. Additionally, the time courses of the fNIRS signals showed that activity was more sustained in response to sequences with changing prosodic contours. The EEG data revealed a similar pattern for the P2 amplitude, which was smaller during the second half for blocks with fixed prosody, whereas the P1 was consistently larger for sequences with variable prosody. Moreover, the sources of the P2, but not the P1, were lateralised to the right, suggesting that the hemispheric asymmetry in the processing of voice pitch changes develops over time.

neuroscience↗

bioRxiv2020

A robust and versatile nanobody platform for drug delivery

Therapeutic and diagnostic efficacies of numerous small biomolecules and chemical compounds are hampered by the short half-lives. Here we report the development of a repertoire of diverse, high-affinity albumin-nanobodies (NbHSA) to facilitate drug delivery. By integrating biophysics, and hybrid structural biology, we have systematically characterized the NbHSA for albumin binding, mapped the epitopes, and resolved the architecture of a tetrameric Nb-albumin complex. We employed quantitative proteomics for accurate, multiplex Nb pharmacokinetic analysis. Using a humanized albumin mouse model, we found that the NbHSA has outstanding pharmacokinetics; the most stable NbHSA has a 771-fold T1/2 improvement compared with a control Nb. Interestingly, the pharmacokinetics of NbHSA is related to their biophysical and structural properties. To demonstrate the utility of NbHSA, we developed a highly stable NbHSA-hIL-2 cytokine conjugate "Duraleukin" and confirmed its improved anticancer properties than hIL-2 alone. We envision that this high-quality Nb resource will advance research into novel biotherapeutics.

bioengineering↗

bioRxiv2020

Characterization of a novel gene, srpA, conferring resistance to streptogramin A, pleuromutilins, and lincosamides in Streptococcus suis

Antimicrobial resistance is undoubtedly one of the greatest global health threats. The emergence of multidrug-resistant (MDR) gram-positive pathogens, like methicillin-resistant Staphylococcus aureus, vancomycin-resistant Enterococcus faecium, and {beta}-lactamase-resistant Streptococcus pneumonia, has severely limited our antibiotic arsenal. Numerous ribosome-targeting antibiotics, especially pleuromutilins, oxazolidinones, and streptogramins, are viewed as promising alternatives against aggressive MDR pathogens. In this study, we identified a new ABC-F family determinant, srpA, in Streptococcus suis by a comparative analysis of whole genome sequences of tiamulin-resistant and -sensitive bacteria. Functional cloning confirmed that the deduced gene can mediate cross-resistance to pleuromutilins, lincosamides, and streptogramin A in S. suis and S. aureus. A sequence alignment revealed that srpA shares the highest amino acid identity with Vga(E) (36%) and shows canonical characteristics of ABC-F family members. In SrpA-ribosome docked compounds, the extended loop region of SrpA approaches the valnemulin binding pocket in the ribosome peptidyl-transferase center and competes with bound valnemulin. A detailed mutational analysis of the loop residues confirmed that this domain is crucial for SrpA activity, as substitutions or truncations of this region affect the efficiency and specificity of antibiotic resistance. A ribosome binding assay supported the protective effects of SrpA on the ribosome by preventing antibiotic binding as well as displacing bound drugs. These findings clarify the mechanisms underlying resistance to ribosomal antibiotics.

microbiology↗

bioRxiv2020

Hybrid capture-based sequencing enables unbiased recovery of SAR-CoV-2 genomes from fecal samples and characterization of the dynamics of intra-host variants

BackgroundIn response to the current COVID-19 pandemic, it is crucial to understand the origin, transmission, and evolution of SARS-CoV-2, which relies on close surveillance of genomic diversity in clinical samples. Although the mutation at the population level had been extensively investigated, how the mutations evolve at the individual level is largely unknown, partly due to the difficulty of obtaining unbiased genome coverage of SARS-CoV-2 directly from clinical samples. MethodsEighteen time series fecal samples were collected from nine COVID-19 patients during the convalescent phase. The nucleic acids of SARS-CoV-2 were enriched by the hybrid capture method with different rounds of hybridization. ResultsBy examining the sequencing depth, genome coverage, and allele frequency change, we demonstrated the impeccable performance of the hybrid capture method in samples with Ct value < 34, as well as significant improvement comparing to direct metatranscriptomic sequencing in samples with lower viral loads. We identified 229 intra-host variants at 182 sites in 18 fecal samples. Among them, nineteen variants presented frequency changes > 0.3 within 1-5 days, reflecting highly dynamic intra-host viral populations. Meanwhile, we also found that the same mutation showed different frequency changes in different individuals, indicating a strong random drift. Moreover, the evolving of the viral genome demonstrated that the virus was still viable in the gastrointestinal tract during the convalescent period. ConclusionsThe hybrid capture method enables reliable analyses of inter- and intra-host variants of SARS-CoV-2 genome, which changed dramatically in the gastrointestinal tract; its clinical relevance warrants further investigation.

genomics↗

bioRxiv2020

Phosphoproteomics after nitrate treatments reveal an important role for PIN2 phosphorylation in control of root system architecture.

Nitrate is an important signaling molecule that commands genome-wide gene expression changes that impact metabolism, physiology, plant growth and development. Although gene expression responses to nitrate at the mRNA level have been characterized in great detail, the impact of nitrate signaling at the proteome level has been much less explored. Most signaling pathways involve post-translational modifications of key protein factors and chiefly among these modifications is protein phosphorylation. In an effort to identify new components involved in nitrate responses in plants, we performed analyses of the Arabidopsis thaliana root phosphoproteome in response to nitrate treatments via liquid chromatography coupled to tandem mass spectrometry. We identified 268 phosphoproteins that show significant changes at 5 min or 20 min after nitrate treatments. The large majority of these proteins (96%) are coded by genes that are not modulated at the expression level in response to nitrate treatments in publicly available transcriptome data. Proteins identified by 5 min include potential signaling-components such as kinases or transcription factors. In contrast, by 20 min, proteins identified were associated with protein binding, transporter activity or hormone metabolism functions. Interestingly, the phosphorylation profile of NITRATE TRANSPORTER 1.1 (NRT1.1) mutant plants in response to nitrate at 5 min was significantly different (95%) as compared to wild-type plants. This result is consistent with the role of NRT1.1 as a key component of a nitrate signaling pathway that involves phosphoproteomic changes. Our integrative bioinformatics analysis highlights auxin transport as an important mechanism modulated by nitrate signaling at the post-translational level. We experimentally validated the role of PIN2 phosphorylation in both primary and lateral root growth responses to nitrate. Our data provide new insights into the phosphoproteome and identifies novel protein components that are regulated post-translationally, such as PIN2, in nitrate responses in Arabidopsis thaliana roots.

plant biology↗

bioRxiv2020

Transduction catalysis: doxorubicin accelerates and enhances rAAV-mediated gene expression in the cortex of mouse, cat and monkey

Rapid and efficient gene transduction via recombinant adeno-associated viruses (rAAVs) is highly desirable across many basic and clinical research domains. Here we report vector co-infusion with doxorubicin, a clinical anti-cancer drug, markedly enhanced rAAV-mediated gene expression in the cerebral cortex across mammalian species (cat, mouse, and macaque), acting throughout the time-period examined and detectable at just three days post-transfection. This enhancement showed serotype generality, being common to rAAV serotypes 2, 8, 9 and PHP.eB tested, and was observed both locally, and at remote locations consistent with doxorubicin undergoing retrograde axonal transport. All these effects were observed at doses matching human blood plasma levels in clinical therapy, and lacked detectable cytotoxicity as assessed by cell morphology, activity, apoptosis and behavioral testing. Altogether, this study identifies an effective means to improve the capability and scope of in vivo rAAV applications, accelerating and augmenting gene transduction at doxorubicin concentrations paralleling medical practice. HighlightsO_LIAnti-cancer drug doxorubicin doubles the rate of rAAV-mediated transgene expression C_LIO_LIDoxorubicin enhancement generalizes across rAAV serotypes and animal species C_LIO_LIThe effect is observed in both locally and retrogradely infected cortical neurons C_LIO_LIThe effective dosage is free from appreciable cytotoxicity and matches clinical settings C_LI

neuroscience↗

bioRxiv2020

Auditory cortex activity measured with functional near-infrared spectroscopy is susceptible to masking by cortical blood stealing

To validate the use of functional near-infrared spectroscopy (fNIRS) in auditory perception experiments, combined fNIRS and electroencephalography (EEG) data were obtained from normal-hearing subjects passively listening to speech-like stimuli without linguistic content. The fNIRS oxy-haemoglobin (HbO) results were found to be inconsistent with the deoxy-haemoglobin (HbR) and EEG data, as they were dominated by pronounced cerebral blood stealing in anterior- to-posterior direction. This large-scale bilateral gradient in the HbO data masked the right-lateralised neural activity in the auditory cortex that was clearly evident in the HbR data and EEG source reconstructions. When the subjects were subsequently split into subgroups with more positive or more negative HbO responses in the right auditory cortex, the former group surprisingly showed smaller event-related potentials, less activity in frontal cortex, and increased EEG alpha power, all indicating reduced attention and vigilance. These findings thus suggest that positive HbO responses in the auditory cortex may not necessarily be a favourable result when investigating auditory perception using fNIRS. More generally, the results show that the interpretation of fNIRS HbO signals can be misleading and demonstrate the benefits of combined fNIRS-EEG analyses in resolving this issue.

neuroscience↗

bioRxiv2020

Natural genetic variation affecting transcription factor spacing at regulatory regions is generally well tolerated

Regulation of gene expression requires the combinatorial binding of sequence-specific transcription factors (TFs) at promoters and enhancers. Prior studies showed that alterations in the spacing between TF binding sites can influence promoter and enhancer activity. However, the relative importance of TF spacing alterations resulting from naturally occurring insertions and deletions (InDels) has not been systematically analyzed. To address this question, we first characterized the genome-wide spacing relationships of 75 TFs in K562 cells as determined by ChIP-sequencing. We found a dominant pattern of a relaxed range of spacing between collaborative factors, including 46 TFs exclusively exhibiting relaxed spacing with their binding partners. Next, we exploited millions of InDels provided by genetically diverse mouse strains and human individuals to investigate the effects of altered spacing on TF binding and local histone acetylation. Spacing alterations resulting from naturally occurring InDels are generally tolerated in comparison to genetic variants directly affecting TF binding sites. A remarkable range of tolerance was further established for PU.1 and C/EBP{beta}, which exhibit relaxed spacing, by introducing synthetic spacing alterations ranging from 5-bp increase to >30-bp decrease using CRISPR/Cas9 mutagenesis. These findings provide implications for understanding mechanisms underlying enhancer selection and for the interpretation of non-coding genetic variation.

genomics↗

bioRxiv2020

Neuron-glia Integrity: Functional Assessment, Molecular Underpinnings, and Implication for Higher Brain Functions

We developed a theory of neuron-glia integrity to underline the fact that neurons and glia cells work together in the central nervous system. Here we substantiated this theory and exemplified the implication of intact neuron-glia integrity for higher brain functions. An animal model of maternal separation with early weaning (MSEW) was applied to neonatal rats to mimic early life neglect and abuse in humans. Behavioral performance of rats was evaluated at adulthood, followed by functional assessments of neuron-glia integrity in living rats, and the demonstration of molecular underpinnings of impaired neuron-glia integrity in MSEW rats. MSEW rats showed higher levels of anxiety and explorative activity, higher glutamate level, but lower GABA level in PFC and hippocampus. MSEW procedure down-regulated protein levels of GLT-1 and ATP-, but up-regulated GAD65 and GS, while had no effects on GLAST and PAG. Moreover, it reduced the fractional anisotropy values in various brain regions, in addition to increasing NAA levels. Concurrently, MSEW led to hypomyelination in PFC as evidenced by relevant cellular and molecular changes.

neuroscience↗

bioRxiv2020

Large-scale Proteomic and Phosphoproteomic Analysis of Maize Seedling Leaves During De-etiolation

De-etiolation consists of a series of developmental and physiological changes that a plant undergoes in response to light. During this process light, an important environmental signal, triggers the inhibition of mesocotyl elongation and the production of photosynthetically active chloroplasts, and etiolated leaves transition from the "sink" stage to the "source" stage. De-etiolation has been extensively studied in maize (Zea mays L). However, little is known about how this transition is regulated. In this study, we describe a quantitative proteomic and phosphoproteomic atlas of the de-etiolation process in maize. We identified 16,420 proteins and quantified 14,168. In addition, 8,746 phosphorylation sites within 3,110 proteins were identified. From the proteomic and phosphoproteomic data combined, we identified a total of 17,436 proteins, 27.6% of which are annotated protein coding genes in the Zea_mays AGPv3.28 database. Only 6% of proteins significantly changed in abundance during de-etiolation. In contrast, the phosphorylation levels of more than 25% of phosphoproteins significantly changed; these included proteins involved in gene expression and homeostatic pathways and rate-limiting enzymes involved in photosynthesis light and carbon reactions. Based on phosphoproteomic analysis, 34% (1,057) of all phosphoproteins identified in this study contained more than three phosphorylation sites, and 37 proteins contained more than 16 phosphorylation sites, which shows that multi-phosphorylation is ubiquitous during the de-etiolation process. Our results suggest that plants might preferentially regulate the level of PTMs rather than protein abundance for adapting to changing environments. The study of PTMs could thus better reveal the regulation of de-etiolation.

bioinformatics↗

bioRxiv2020

Novel and potent inhibitors targeting DHODH, a rate-limiting enzyme in de novo pyrimidine biosynthesis, are broad-spectrum antiviral against RNA viruses including newly emerged coronavirus SARS-CoV-2

Emerging and re-emerging RNA viruses occasionally cause epidemics and pandemics worldwide, such as the on-going outbreak of coronavirus SARS-CoV-2. Existing direct-acting antiviral (DAA) drugs cannot be applied immediately to new viruses because of virus-specificity, and the development of new DAA drugs from the beginning is not timely for outbreaks. Thus, host-targeting antiviral (HTA) drugs have many advantages to fight against a broad spectrum of viruses, by blocking the viral replication and overcoming the potential viral mutagenesis simultaneously. Herein, we identified two potent inhibitors of DHODH, S312 and S416, with favorable drug-like and pharmacokinetic profiles, which all showed broad-spectrum antiviral effects against various RNA viruses, including influenza A virus (H1N1, H3N2, H9N2), Zika virus, Ebola virus, and particularly against the recent novel coronavirus SARS-CoV-2. Our results are the first to validate that DHODH is an attractive host target through high antiviral efficacy in vivo and low virus replication in DHODH knocking-out cells. We also proposed the drug combination of DAA and HTA was a promising strategy for anti-virus treatment and proved that S312 showed more advantageous than Oseltamivir to treat advanced influenza diseases in severely infected animals. Notably, S416 is reported to be the most potent inhibitor with an EC50 of 17nM and SI value >5882 in SARS-CoV-2-infected cells so far. This work demonstrates that both our self-designed candidates and old drugs (Leflunomide/Teriflunomide) with dual actions of antiviral and immuno-repression may have clinical potentials not only to influenza but also to COVID-19 circulating worldwide, no matter such viruses mutate or not.

microbiology↗

bioRxiv2020

Genetic elucidation of complex biochemical traits mediating maize innate immunity

Specialized metabolites constitute key layers of immunity underlying crop resistance; however, challenges in resolving complex pathways limit our understanding of their functions and applications. In maize (Zea mays) the inducible accumulation of acidic terpenoids is increasingly considered as a defense regulating disease resistance. To understand maize antibiotic biosynthesis, we integrated association mapping, pan-genome multi-omic correlations, enzyme structure-function studies, and targeted mutagenesis. We now define ten genes in three zealexin (Zx) gene clusters comprised of four sesquiterpene synthases and six cytochrome P450s that collectively drive the production of diverse antibiotic cocktails. Quadruple mutants blocked in the production of {beta}-macrocarpene exhibit a broad-spectrum loss of disease resistance. Genetic redundancies ensuring pathway resiliency to single null mutations are combined with enzyme substrate-promiscuity creating a biosynthetic hourglass pathway utilizing diverse substrates and in vivo combinatorial chemistry to yield complex antibiotic blends. The elucidated genetic basis of biochemical phenotypes underlying disease resistance demonstrates a predominant maize defense pathway and informs innovative strategies for transferring chemical immunity between crops.

plant biology↗

bioRxiv2020

Distinct natural syllable-selective neuronal ensembles in the primary auditory cortex of awake marmosets

Marmosets are highly social non-human primates living in families. They exhibit rich vocalization, but neural basis underlying complex vocal communication is largely unknown. Here we report the existence of specific neuronal ensembles in marmoset A1 that respond selectively to distinct monosyllable or disyllable calls made by conspecific marmosets. These neurons were spatially dispersed within A1 but distinct from those responsive to pure tones. Syllable-selective responses were markedly diminished when individual domains of the syllable were deleted or the domain sequence was altered, indicating the importance of global rather than local spectral-temporal properties of the sound. Disyllable-selective responses also disappeared when the sequence of the two monosyllable components was reversed or their interval was extended beyond 1 second. Light anesthesia largely abolished disyllable-selective responses. Our findings demonstrate extensive inhibitory and facilitory interactions among syllable-evoked responses, and provide the basis for further study of circuit mechanisms underlying vocal communication in awake non-human primates.

neuroscience↗

bioRxiv2020

Targeting OCA-B/Pou2af1 blocks type-1 diabetes and reduces infiltration of activated, islet-reactive T cells

The transcriptional coregulator OCA-B promotes expression of T cell target genes in cases of repeated antigen exposure, a necessary feature of autoimmunity. We hypothesized that T cell-specific OCA-B deletion and pharmacologic OCA-B inhibition would protect mice from autoimmune diabetes. We developed an Ocab conditional allele and backcrossed it onto a diabetes-prone NOD/ShiLtJ strain background. T cell-specific OCA-B loss protected mice from spontaneous disease. Protection was associated with large reductions in islet CD8+ T cell receptor specificities associated with diabetes pathogenesis. CD4+ clones associated with diabetes were present, but associated with anergic phenotypes. The protective effect of OCA-B loss was recapitulated using autoantigen-specific NY8.3 mice, but diminished in monoclonal models specific to artificial or neoantigens. Rationally-designed membrane-penetrating OCA-B peptide inhibitors normalized glucose levels, and reduced T cell infiltration and proinflammatory cytokine expression in newly-diabetic NOD mice. Together, the results indicate that OCA-B is a potent autoimmune regulator and a promising target for pharmacologic inhibition. ~40-word summary statement for the online JEM table of contents and alertsKim and colleagues show that OCA-B in T cells is essential for the generation of type-1 diabetes. OCA-B loss leaves the pancreatic lymph nodes largely undisturbed, but associates autoreactive CD4+ T cells in the pancreas with anergy while deleting potentially autoreactive CD8+ T cells. SummaryKim et al. show that loss or inhibition of OCA-B in T cells protects mice from type-1 diabetes.

immunology↗

bioRxiv2020

MAGGIE: leveraging genetic variation to identify DNA sequence motifs mediating transcription factor binding and function

Genetic variation in regulatory elements can alter transcription factor (TF) binding by mutating a TF binding motif, which in turn may affect the activity of the regulatory elements. However, it is unclear which TFs are prone to be affected by a given variant. Current motif analysis tools either prioritize TFs based on motif enrichment without linking to a function or are limited in their applications due to the assumption of linearity between motifs and their functional effects. Here, we present MAGGIE, a novel method for identifying motifs mediating TF binding and function. By leveraging measurements from diverse genotypes, MAGGIE uses a statistical approach to link mutation of a motif to changes of an epigenomic feature without assuming a linear relationship. We benchmark MAGGIE across various applications using both simulated and biological datasets and demonstrate its improvement in sensitivity and specificity compared to the state-of-the-art motif analysis approaches. We use MAGGIE to reveal insights into the divergent functions of distinct NF-{kappa}B factors in the pro-inflammatory macrophages, showing its promise in discovering novel functions of TFs. The Python package for MAGGIE is freely available at https://github.com/zeyang-shen/maggie.

bioinformatics↗

bioRxiv2020

CG14906 (mettl4) mediates m6A methylation of U2 snRNA in Drosophila

Recent studies reported that METTL4 regulates DNA 6mA in vivo and therefore is a candidate DNA m6A methyltransfease. However, the enzymatic activity of METTL4 in vitro has not been demonstrated in part due to the difficulties of obtaining well-folded proteins. Here we show that mettl4 is a major methyltransfase responsible for m6A methylation of U2 snRNA both in vitro and in vivo in fly, and identify adenosine at 29th position as the site of m6A methylation. This study answered a long-standing question regarding the enzymatic activity of METTL4, and thus paved the way for further investigating the functions of METTL4 in different biological settings.

biochemistry↗

bioRxiv2019

Mismatch responses mediated by adaptation and deviance detection have complementary functional profiles that point to different auditory short-term memory systems

Mismatch negativity (MMN) is a macroscopic EEG deflection in response to rare or unexpected sounds. It has provided important insights into auditory short-term memory, pre-attentive guidance of attention, and their alteration in conditions such as schizophrenia. It remains unclear if MMN is caused by passive adaptation, active memory-comparison processes (deviance detection; DD), or a mix of both. To answer this question, macaque monkeys listened to a new paradigm that quantified both components of MMN. Micro- and macroscopic mismatch responses in the rhesus were dominated by adaptation at short latencies but included a smaller contribution of deviance detection at longer latencies. Most importantly, we show that mismatch responses mediated by adaptation have a short temporal scope and narrow frequency tuning while mismatch responses mediated by deviance detection have a longer temporal scope but broader frequency tuning. The different functional profiles point to the involvement of two distinct auditory short-term memory systems and complementary roles in the pre-attentive guidance of attention.

neuroscience↗

bioRxiv2019

Deletion of mouse Setd4 promotes the recovery of hematopoietic failure.

Acquired hematopoietic failure is commonly caused by therapeutic and accidental exposure to toxic agents to the bone marrow (BM). Efficient recovery from the BM failure is not only dictated by the intrinsic sensitivity and proliferation capacity of the hematopoietic stem and progenitor cells, but also nourished by the BM environment niche. Identification of genetic factors that improve the recovery from hematopoietic failure is essential. Vertebrate SETD4 is a poorly characterized, putative non-histone methyl-transferase whose physiological substrates have not yet been fully identified. By inducing Setd4 deletion in adult mice, we found that loss of Setd4 improved the survival of whole body irradiation induced BM failure. This was associated with improved recoveries of long-term and short-term hematopoietic stem cells (HSC), and early progenitor cells. BM transplantation analyses surprisingly showed that the improved recovery was not due to a radiation resistance of the Setd4 deficient HSC, but that Setd4 deficient HSC were actually more sensitive to radiation. However, the Setd4 deficient mice were better recipients for allogeneic HSC transplantation. Furthermore, there was an enhanced splenic erythropoiesis in Setd4 deficient mice. These findings not only revealed a previously unrecognized role of the Setd4 as a unique modulator of hematopoiesis, but also underscored the critical role of the BM niche in the recovery of hematopoietic failure. These studies also implicated Setd4 as a potential target for therapeutic inhibition to improve the conditioning of the BM niche prior to allogeneic transplantation. Key pointsO_LIDeletion of Setd4 in adult mice improved the survival from hematopoietic failure. C_LIO_LISetd4 deficiency sensitized HSCs to radiation, but improved bone marrow environment niche. C_LIO_LIThe study suggests that SETD4 as a potential inhibitory target to improve bone marrow niche function for recovery of bone marrow failure. C_LI

developmental biology↗