Biology subjects

Yao, D.

Publications and source records attributed to Yao, D..

46 records · Page 3Linked to original sources

bioRxiv2020

Phytophthora sojae effector Avr1d functions as E2 competitor and inhibits ubiquitination activity of GmPUB13 to facilitate infection

Oomycete pathogens such as Phytophthora secrete a repertoire of effectors to host cells to manipulate host immunity and benefit infection. In this study, we found that an RxLR effector, Avr1d, promoted Phytophthora sojae infection in soybean hairy-roots. Using a yeast two-hybrid screen, we identified the soybean E3 ubiquitin ligase GmPUB13 as a host target for Avr1d. By co-immunoprecipitation, gel infiltration and ITC assays, we confirmed that Avr1d interacts with GmPUB13 both in vivo and in vitro. Furthermore, we found that Avr1d inhibits the E3 ligase activity of GmPUB13. The crystal structure of Avr1d in complex with GmPUB13 was solved and revealed that Avr1d occupies the binding site for E2 ubiquitin conjugating enzyme on GmPUB13. In line with this, Avr1d competed with E2 ubiquitin conjugating enzymes for GmPUB13 binding in vitro, thereby decreasing the E3 ligase activity of GmPUB13. Meanwhile, we found that inactivation of the ubiquitin ligase activity of GmPUB13 stabilized GmPUB13 by blocking GmPUB13 degradation. Silencing of GmPUB13 in soybean hairy-roots decreased P. sojae infection, suggesting that GmPUB13 acts as a susceptibility factor, negatively regulating soybean resistance against P. sojae. Altogether, this study highlights a novel virulence mechanism of Phytophthora effectors, by which Avr1d competes with E2 for GmPUB13 binding to repress the GmPUB13 E3 ligase activity and thereby stabilizing the susceptibility factor GmPUB13 to facilitate Phytophthora infection. This is the first study to unravel the structural basis for modulation of host targets by Phytophthora effectors and will be instrumental for boosting plant resistance breeding. Significance StatementUbiquitination acts as a crucial regulator in plant immunity. Accordingly, microbial pathogens secrete effectors to hijak host ubiquitination system. However, the molecular mechanisms by which microbial effectors modulate host ubiquitination system are not yet clear. Here, we found that the Phytophthora sojae effector Avr1d physically binds to the U-box type E3 ligase GmPUB13, a susceptibility factor in soybean. The crystal structure of Avr1d in complex with GmPUB13 revealed that Avr1d occupies the binding site in GmPUB13 for the E2 ubiquitin conjugating enzyme and competes with E2 for physical binding to GmPUB13. Avr1d stabilized GmPUB13 by suppressing the self-ubiquitination activity of GmPUB13 and thereby promoting Phytophthora infection. This study provides structural basis for modulation of host targets by Phytophthora effectors.

microbiology↗

bioRxiv2020

High-throughput discovery and characterization of human transcriptional effectors

Thousands of proteins localize to the nucleus; however, it remains unclear which contain transcriptional effectors. Here, we develop HT-recruit - a pooled assay where protein libraries are recruited to a reporter, and their transcriptional effects are measured by sequencing. Using this approach, we measure gene silencing and activation for thousands of domains. We find a relationship between repressor function and evolutionary age for the KRAB domains, discover Homeodomain repressor strength is collinear with Hox genetic organization, and identify activities for several Domains of Unknown Function. Deep mutational scanning of the CRISPRi KRAB maps the co-repressor binding surface and identifies substitutions that improve stability/silencing. By tiling 238 proteins, we find repressors as short as 10 amino acids. Finally, we report new activator domains, including a divergent KRAB. Together, these results provide a resource of 600 human proteins containing effectors and demonstrate a scalable strategy for assigning functions to protein domains.

genetics↗

bioRxiv2020

Linking cerebellar functional gradients to transdiagnostic behavioral dimensions of psychopathology

High co-morbidity and substantial overlap across psychiatric disorders encourage a transition in psychiatry research from categorical to dimensional approaches that integrate neuroscience and psychopathology. Cerebellum is involved in a wide range of nonmotor cognitive functions and mental disorders. An important question thus centers on the extent to which cerebellar function can be linked to transdiagnostic dimensions of psychopathology. Here, this question is investigated using partial least squares to identify latent dimensions linking cerebellar connectome properties as assessed by macroscale spatial gradients of connectivity to a large set of clinical and behavioral measures in 198 participants across diagnostic categories. This analysis reveals significant correlated patterns of cerebellar connectivity gradients and behavioral measures that could be represented into four latent dimensions: general psychopathology, general lack of attention regulation, internalizing symptoms, and dysfunctional memory. Each dimension is associated with a distinct spatial pattern of cerebellar connectivity gradients. These findings highlight the relevance of cerebellar connectivity as a necessity for the study and classification of transdiagnostic dimensions of psychopathology.

neuroscience↗

bioRxiv2020

Human H-ferritin presenting RBM of spike glycoprotein as potential vaccine of SARS-CoV-2

The outbreak of COVID-19 has so far inflicted millions of people all around the world and will have a long lasting effect on every aspect of everyones life. Yet there is no effective approved treatment for the disease. In an effort of utilizing human ferritin as nanoplatform for drug delivery, we engineered a fusion protein by presenting receptor-binding motif (RBM) of SARS-CoV-2 virus spike glycoprotein on the N-terminus of ferritin subunits. The designed fusion protein with a cage-like structure, similar to that of corona virus, is a potential anti-SARS-CoV-2 vaccine. We hereby show the construction, preparation, and characterization of the fusion protein RBM-HFtn. Our initial affinity study confirmed its biological activity towards ACE2 receptor which suggests its mode of action against SARS-CoV-2 could be either through vaccine therapy or blocking the cellular entry of virus as antagonist of ACE2 receptor.

bioengineering↗

bioRxiv2020

Altered Sensorimotor-to-Transmodal Hierarchical Organization in Schizophrenia

For decades, schizophrenia has been primarily conceptualized as a disorder of high-order cognitive functions with deficits in executive brain regions. Yet due to the increasing reports of early sensory processing deficit, recent models focus more on the developmental effects of impaired sensory process on high-order functions. The present study examined whether this pathological interaction relates to an overarching system-level imbalance, specifically a disruption in macroscale hierarchy affecting integration and segregation of unimodal and transmodal networks. We applied a novel combination of connectome gradient and stepwise connectivity analysis to resting-state functional magnetic resonance imaging (rsfMRI) to characterize the sensorimotor-to-transmodal cortical hierarchy organization (96 patients vs. 122 controls). Using these techniques, we demonstrated compression of the cortical hierarchy organization in schizophrenia, with a prominent compression from the sensorimotor region and a less prominent compression from the frontal-parietal region, resulting in a diminished separation between sensory and fronto-parietal cognitive systems. Further analyses suggested reduced differentiation related to atypical functional connectome transition from unimodal to transmodal brain areas. Specifically, we found hypo-connectivity within unimodal regions and hyper-connectivity between unimodal regions and frontoparietal and ventral attention regions along the classical sensation-to-cognition continuum established in prior neuroanatomical work. The compression of cortical hierarchy organization represents a novel and integrative system-level substrate underlying the pathological interaction of early sensory and cognitive function in schizophrenia. This abnormal cortical hierarchy organization suggests cascaded impairments stemming from the disrupted somatosensory-motor system and inefficient integration of bottom-up sensory information with attentional demands and executive control processes partially account for high-level cognitive deficits characteristic of schizophrenia.

neuroscience↗

bioRxiv2019

Dimensional neuroimaging: (Internet) Gaming Disorder symptoms according to the WHO and APA frameworks associate with lower striatal volume

Growing concerns about the addictive nature of internet and computer games led to the endorsement of Internet Gaming Disorder (IGD) as an emerging disorder by the American Psychological Association (APA) in 2013 and the recognition of Gaming Disorder (GD) as a new diagnosis by the World Health Organization (WHO) in 2019. While the definition of clear diagnostic criteria for IGD is beneficial for clinicians and those in need of treatment, it may also stigmatize normal behavior. Furthermore, the neurobiological mechanisms underlying IGD symptoms, and whether they resemble those of other addictive disorders remains highly debated. To this end the present study employed a dimensional imaging approach to determine associations between IGD symptom-load according to the APA and WHO diagnostic frameworks and brain structure in a comparably large sample of healthy subjects. It was found that higher symptom-load on psychometric tests assessing the APA and WHO diagnostic frameworks convergently associated with lower volumes of the striatum. The present results suggest for the first time a neurobiological foundation of the proposed diagnostic criteria for IGD according to both diagnostic classification systems, further indicating that the transition from non-disordered to disordered gaming is accompanied by progressive neuroplastic changes in the striatum, thus resembling progressive changes in other addictive disorders.

neuroscience↗

bioRxiv2019

Objects seen as scenes: neural circuitry for attending whole or parts

Depending on our goals, we pay attention to the global shape of an object or to the local shape of its parts, since its difficult to do both at once. This typically effortless process can be impaired in disease. However, it is not clear which cortical regions carry the information needed to constrain shape processing to a chosen global/local level. Here, novel stimuli were used to dissociate functional MRI responses to global and local shapes. This allowed identification of cortical regions containing information about level (independent from shape). Crucially, these regions overlapped part of the cortical network implicated in scene processing. As expected, shape information (independent of level) was mainly located in category-selective areas specialized for object- and face-processing. Regions with the same informational profile were strongly linked (as measured by functional connectivity), but were weak when the profiles diverged. Specifically, in the ventral-temporal-cortex (VTC) regions favoring level and shape were consistently separated by the mid-fusiform sulcus (MFS). These regions also had limited crosstalk despite their spatial proximity, thus defining two functional pathways within VTC. We hypothesize that object hierarchical level is processed by neural circuitry that also analyses spatial layout in scenes, contributing to the control of the spatial-scale used for shape recognition. Use of level information tolerant to shape changes could guide whole/part attentional selection but facilitate illusory shape/level conjunctions under impoverished vision. Significance statementOne daily engages hierarchically organized objects (e.g. face-eyes-eyelashes). Their perception is commonly studied with global shapes composed by of local shapes. Seeing shape at one level is easy, but difficult for both at once. How can the brain guide attention to one level? Here using novel stimuli that dissociate different levels over time and examining local patterns of brain-activity, we found that the level and shape of visual objects were represented into segregated sets of cortical regions, each connected into their own pathway. Level information was found in part of the cortical network known to process scenes. Coding of object-level independently from shape could participate in guiding sustained attention within objects, eliminating interference from irrelevant levels. It could also help produce "illusory conjunctions" (perceptual migration of a shape to the wrong level) when attention is limited. HighlightsO_LIModified Navon figures allow dissociation in time of fMRI responses for the global/local levels. C_LIO_LIShape-invariant hierarchical level information was found in scenes selective areas, whereas level-invariant shape information was found in object- and faces- selective areas. C_LIO_LILevel and shape regions were divided by the mid-fusiform sulcus (MFS) in VTC cortex, and each type of region connected into its own pathway. C_LIO_LIHaving separate level/shape pathways could facilitate selective-attention, but foster illusory conjunctions. C_LI

neuroscience↗

bioRxiv2019

Identification and mitigation of pervasive off-target activity in CRISPR-Cas9 screens for essential non-coding elements

Pooled CRISPR-Cas9 screens have recently emerged as a powerful method for functionally characterizing regulatory elements in the non-coding genome, but off-target effects in these experiments have not been systematically evaluated. Here, we conducted a genome-scale screen for essential CTCF loop anchors in the K562 leukemia cell line. Surprisingly, the primary drivers of signal in this screen were single guide RNAs (sgRNAs) with low specificity scores. After removing these guides, we found that there were no CTCF loop anchors critical for cell growth. We also observed this effect in an independent screen fine-mapping the core motifs in enhancers of the GATA1 gene. We then conducted screens in parallel with CRISPRi and CRISPRa, which do not induce DNA damage, and found that an unexpected and distinct set of off-targets also caused strong confounding growth effects with these epigenome-editing platforms. Promisingly, strict filtering of CRISPRi libraries using GuideScan specificity scores removed these confounded sgRNAs and allowed for the identification of essential enhancers, which we validated extensively. Together, our results show off-target activity can severely limit identification of essential functional motifs by active Cas9, while strictly filtered CRISPRi screens can be reliably used for assaying larger regulatory elements.

genomics↗

bioRxiv2018

Valection: Design Optimization for Validation and Verification Studies

BackgroundPlatform-specific error profiles necessitate confirmatory studies where predictions made on data generated using one technology are additionally verified by processing the same samples on an orthogonal technology. In disciplines that rely heavily on high-throughput data generation, such as genomics, reducing the impact of false positive and false negative rates in results is a top priority. However, verifying all predictions can be costly and redundant, and testing a subset of findings is often used to estimate the true error profile. To determine how to create subsets of predictions for validation that maximize inference of global error profiles, we developed Valection, a software program that implements multiple strategies for the selection of verification candidates.\n\nResultsTo evaluate these selection strategies, we obtained 261 sets of somatic mutation calls from a single-nucleotide variant caller benchmarking challenge where 21 teams competed on whole-genome sequencing datasets of three computationally-simulated tumours. By using synthetic data, we had complete ground truth of the tumours mutations and, therefore, we were able to accurately determine how estimates from the selected subset of verification candidates compared to the complete prediction set. We found that selection strategy performance depends on several verification study characteristics. In particular the verification budget of the experiment (i.e. how many candidates can be selected) is shown to influence estimates.\n\nConclusionsThe Valection framework is flexible, allowing for the implementation of additional selection algorithms in the future. Its applicability extends to any discipline that relies on experimental verification and will benefit from the optimization of verification candidate selection.

bioinformatics↗

bioRxiv2017

Large-scale screening of rare genetic variants in humans reveals frequent splicing disruptions

Any individuals genome contains [~]4-5 million genetic variants that differ from reference, and understanding how these variants give rise to trait diversity and disease susceptibility is a central goal of human genetics1. A vast majority (96-99%) of an individuals variants are common, though at a population level the overwhelming majority of variants are rare2-5. Because of their scarcity in an individuals genome, rare variants that play important roles in complex traits are likely to have large functional effects6,7. Mutations that cause an exon to be skipped can have severe functional consequences on gene function, and many known disease-causing mutations reduce or eliminate exon recognition8. Here we explore the extent to which rare genetic variation in humans results in near complete loss of exon recognition. We developed a Multiplexed Functional Assay of Splicing using Sort-seq (MFASS) that allows us to measure exon inclusion in thousands of human exons and surrounding intronic sequence simultaneously. We assayed 27,733 extant variants in the Exome Aggregation Consortium (ExAC)9 within or adjacent to 2,339 human exons, and found that 3.8% (1,050) of the variants, almost all of which were extremely rare, led to large-effect defects in exon recognition. Importantly, we find that 83% of these splice-disrupting variants (SDVs) are located outside of canonical splice sites, are distributed evenly across distinct exonic and intronic regions, and are difficult to predict a priori. Our results indicate that loss of exon recognition is an important and underappreciated means by which rare variants exert large functional effects, and that MFASS enables their empirical assessment for splicing defects at scale.

genetics↗