Biology subjects

Tian, H.

Publications and source records attributed to Tian, H..

85 records · Page 5Linked to original sources

bioRxiv2019

Novel patterns of complex structural variation revealed across thousands of cancer genome graphs

Cancer genomes often harbor hundreds of somatic DNA rearrangement junctions, many of which cannot be easily classified into simple (e.g. deletion, translocation) or complex (e.g. chromothripsis, chromoplexy) structural variant classes. Applying a novel genome graph computational paradigm to analyze the topology of junction copy number (JCN) across 2,833 tumor whole genome sequences (WGS), we introduce three complex rearrangement phenomena: pyrgo, rigma, and tyfonas. Pyrgo are "towers" of low-JCN duplications associated with early replicating regions and superenhancers, and are enriched in breast and ovarian cancers. Rigma comprise "chasms" of low-JCN deletions at late-replicating fragile sites in esophageal and other gastrointestinal (GI) adenocarcinomas. Tyfonas are "typhoons" of high-JCN junctions and fold back inversions that are enriched in acral but not cutaneous melanoma and associated with a previously uncharacterized mutational process of non-APOBEC kataegis. Clustering of tumors according to genome graph-derived features identifies subgroups associated with DNA repair defects and poor prognosis.

genomics↗

bioRxiv2019

Nanopore sequencing of DNA concatemers reveals higher-order features of chromatin structure

Higher-order chromatin structure arises from the combinatorial physical interactions of many genomic loci. To investigate this aspect of genome architecture we developed Pore-C, which couples chromatin conformation capture with Oxford Nanopore Technologies (ONT) long reads to directly sequence multi-way chromatin contacts without amplification. In GM12878, we demonstrate that the pairwise interaction patterns implicit in Pore-C multi-way contacts are consistent with gold standard Hi-C pairwise contact maps at the compartment, TAD, and loop scales. In addition, Pore-C also detects higher-order chromatin structure at 18.5-fold higher efficiency and greater fidelity than SPRITE, a previously published higher-order chromatin profiling technology. We demonstrate Pore-Cs ability to detect and visualize multi-locus hubs associated with histone locus bodies and active / inactive nuclear compartments in GM12878. In the breast cancer cell line HCC1954, Pore-C contacts enable the reconstruction of complex and aneuploid rearranged alleles spanning multiple megabases and chromosomes. Finally, we apply Pore-C to generate a chromosome scale de novo assembly of the HG002 genome. Our results establish Pore-C as the most simple and scalable assay for the genome-wide assessment of combinatorial chromatin interactions, with additional applications for cancer rearrangement reconstruction and de novo genome assembly.

genomics↗

bioRxiv2019

A secreted LysM effector protects fungal hyphae through chitin-dependent homodimer polymerization

Plants trigger immune responses upon recognition of fungal cell wall chitin, followed by the release of various antimicrobials, including chitinase enzymes that hydrolyze chitin. In turn, many fungal pathogens secrete LysM effectors that prevent chitin recognition by the host through scavenging of chitin oligomers. We previously showed that intrachain LysM dimerization of the Cladosporium fulvum effector Ecp6 confers an ultrahigh-affinity binding groove that competitively sequesters chitin oligomers from host immune receptors. Additionally, particular LysM effectors are found to protect fungal hyphae against chitinase hydrolysis during host colonization. However, the molecular basis for the protection of fungal cell walls against hydrolysis remained unclear. Here, we determined a crystal structure of the single LysM domain-containing effector Mg1LysM of the wheat pathogen Zymoseptoria tritici and reveal that Mg1LysM is involved in the formation of two kinds of dimers; a chitin-dependent dimer as well as a chitin-independent homodimer. In this manner, Mg1LysM gains the capacity to form a supramolecular structure by chitin-induced oligomerization of chitin-independent Mg1LysM homodimers, a property that confers protection to fungal cell walls against host chitinases.

microbiology↗

bioRxiv2019

HELLS and PRDM9 form a Pioneer Complex to Open Chromatin at Meiotic Recombination Hotspots

Chromatin barriers prevent spurious interactions between regulatory elements and DNA-binding proteins. One such barrier, whose mechanism for overcoming is poorly understood, is access to recombination hotspots during meiosis. Here we show that the chromatin remodeler HELLS and DNA-binding protein PRDM9 function together to open chromatin at hotspots and provide access for the DNA double-strand break (DSB) machinery. Recombination hotspots are decorated by a unique combination of histone modifications, not found at other regulatory elements. HELLS is recruited to hotspots by PRDM9, and is necessary for both histone modifications and DNA accessibility at hotspots. In male mice lacking HELLS, DSBs are retargeted to other sites of open chromatin, leading to germ cell death and sterility. Together, these data provide a model for hotspot activation where HELLS and PRDM9 function as a pioneer complex to create a unique epigenomic environment of open chromatin, permitting correct placement and repair of DSBs.

genetics↗

bioRxiv2019

The glycine locating at random coil of picornaviruses VP3 enhances viral pathogenicity by targeting p53 to promote apoptosis and autophagy

Picornaviruses, comprising important and widespread pathogens of humans and animals, have evolved to control apoptosis and autophagy for their replication and spread. However, the underlying mechanism of the association between apoptosis/autophage and viral pathogenicity remains unclear. In the present study, VP3 of picornaviruses was demonstrated to induce apoptosis and autophagy. Foot-and-mouth disease virus (FMDV), which served as a research model here, can strongly induce both apoptosis and autophagy in the skin lesions. By directly interacting with p53, FMDV-VP3 facilitates its phosphorylation and translocation, resulting in Bcl-2 family-mediated apoptosis and LC3-dependent autophagy. The single residue Gly129 of FMDV-VP3 plays a crucial role in apoptosis and autophagy induction and the interaction with p53. Consistently, the comparison of rescued FMDV with mutated Gly129 and parental virus showed that the Gly129 is indispensable for viral replication and pathogenicity. More importantly, the Gly129 locates at a bend region of random coil structure, the mutation of Gly to Ala remarkably shrunk the volume of viral cavity. Coincidentally, the Gly is conserved in the similarly location of other picornaviruses, including poliovirus (PV), enterovirus 71 (EV71), coxsackievirus (CV) and seneca valley virus (SVA). This study demonstrates that picornaviruses induce apoptosis and autophagy to facilitate its pathogenicity and the Gly is functional site, providing novel insights into picornavirus biology.

microbiology↗

bioRxiv2019

Lentivirus-mediated long-term overexpression of specific microRNA in mammalian cells

The establishment of a method that would overexpress or suppress of specific microRNA activity is essential for the functional analysis of these molecules and for the development of miRNA therapeutic applications. There already exist excellent ways to inhibit miRNA function in vitro and in vivo by overexpressing miRNA target sequences, which include miRNA decoys, sponges, or antagomirs that are complementary to an miRNA seed region. Conversely, no methods to induce stable gain-of-function phenotypes for specific miRNAs have, as yet, been reported. Furthermore, the discovery of complementary miRNA pairs raises suspicion regarding the existing methods used for miRNA overexpression. In our study, we will study whether the traditional methods for miRNA overexpression can be used for specific miRNA overexpression while complementary miRNA pairs exist. In addition, we test various miRNA-expression cassettes that were designed to efficiently overexpress specific miRNA through the shRNA lentivirus expression system. We report the optimal conditions that were established for the design of such miRNA-expression cassettes. We finally demonstrate that the miRNA-expression cassettes achieve efficient and long-term overexpression of specific miRNAs. Meanwhile, our results also support the notion that miRNA-miRNA interactions are implicated in potential, mutual regulatory patterns and beyond the seed sequence of miRNA, extensive pairing interactions between a miRNA and its target also lead to target-directed miRNA degradation. Our results indicate that our method offers a simple and efficient means to over-express the specific miRNA with long-term which will be very useful for future studies in miRNA biology, as well as contributed to the development of miRNA-based therapy for clinical applications.

molecular biology↗

bioRxiv2019

High-Affinity Binding of Chemokine Analogs that Display Ligand Bias at the HIV-1 Co-receptor CCR5

The chemokine receptor CCR5 is a drug target to prevent transmission of HIV/AIDS. We studied four analogs of the native chemokine RANTES (CCL5) that have anti-HIV potencies of around 25 pM, which is more than four orders-of-magnitude higher than that of RANTES itself. It has been hypothesized that the ultra-high potency of the analogs is due to their ability to bind populations of receptors not accessible to native chemokines. To test this hypothesis, we developed a homogeneous dual-color fluorescence cross-correlation spectroscopy (FCCS) assay for saturation and competition binding experiments. The FCCS assay has the advantage that it does not rely on competition with radioactively labeled native chemokines used in conventional assays. We prepared site-specifically labeled fluorescent analogs using native chemical ligation of synthetic peptides, followed by bioorthogonal fluorescent labeling. We engineered a mammalian cell expression construct to provide fluorescently labeled CCR5, which was purified using a tandem immunoaffinity and size-exclusion chromatography approach to obtain monomeric fluorescent CCR5 in detergent solution. We found subnanomolar binding affinities for the two analogs 5P12-RANTES and 5P14-RANTES, and about twenty-fold reduced affinities for PSC-RANTES and 6P4-RANTES. Using homologous and heterologous competition experiments with unlabeled chemokine analogs, we conclude that the analogs all bind at the same binding site; whereas, the native chemokines (RANTES and MIP1) fail to displace bound fluorescent analogs even at tens of micromolar concentrations. Our results can be rationalized with de novo structural models of the N-terminal tails of the synthetic chemokines that adopt a different binding mode as compared to the parent compound.

biophysics↗

bioRxiv2019

PRH1 mediates ARF7-LBD dependent auxin signaling to regulate lateral root development in Arabidopsis thaliana

The development of lateral roots in Arabidopsis thaliana is strongly dependent on signaling directed by the AUXIN RESPONSE FACTOR7 (ARF7), which in turn activates LATERAL ORGAN BOUNDARIES DOMAIN (LBD) transcription factors (LBD16, 18, 29 and 33). Here, the product of PRH1, a PR-1 homolog annotated previously as encoding a pathogen-responsive protein, was identified as a target of ARF7-mediated auxin signaling and also as participating in the development of lateral roots. PRH1 was shown to be strongly induced by auxin treatment, and plants lacking a functional copy of PRH1 formed fewer lateral roots. The transcription of PRH1 was controlled by the binding of both ARF7 and LBDs to its promoter region. An interaction was detected between PRH1 and GATA23, a protein which regulates cell identity in lateral root founder cells. Author SummaryIn Arabidopsis thaliana AUXIN RESPONSE FACTOR7 (ARF7)-mediated auxin signaling plays a key role in lateral roots (LRs) development. The LATERAL ORGAN BOUNDARIES DOMAIN (LBD) transcription factors (LBD16, 18, 29 and 33) act downstream of ARF7-mediated auxin signaling to control LRs formation. Here, the PR-1 homolog PRH1 was identified as a novel target of both ARF7 and LBDs (especially the LBD29) during auxin induced LRs formation, as both ARF7 and LBDs were able to bind to the PRH1 promoter. More interestingly, PRH1 has a physical interaction with GATA23, which has been also reported to be up-regulated by auxin and influences LR formation through its regulation of LR founder cell identity. Whether the interaction between GATA23 and PRH1 affects the stability and/or the activity of either (or both) of these proteins remains an issue to be explored. This study provides improves new insights about how auxin regulates lateral root development.

plant biology↗

bioRxiv2018

EWSR1 affects PRDM9-dependent histone 3 methylation and provides a link between recombination hotspots and the chromosome axis

Meiotic recombination in most mammals requires recombination hotspot activation through the action of the histone 3 lysine-4 and lysine-36 methyltransferase PRDM9 to ensure successful double-strand break initiation and repair. Here we show that EWSR1, a protein whose role in meiosis was not previously clarified in detail, binds to both PRDM9 and pREC8, a phosphorylated meiosis-specific cohesin, in male meiotic cells. We created a Ewsr1 conditional knockout mouse models to deplete EWSR1 before the onset of meiosis, and found that absence of EWSR1 causes meiotic arrest with decreased histone trimethylation at meiotic hotspots, impaired DNA double-strand break repair, and reduced crossover number. Our results demonstrate that EWSR1 is essential for promoting PRDM9-dependent histone methylation and normal meiotic progress, possibly by facilitating the linking between PRDM9-bound hotspots and the nascent chromosome axis. Author SummaryIn most mammals, including humans and mice, genetic recombination initiates when the meiosis-specific protein PRDM9 binds specific DNA sequences, known as hotspots, at the beginning of the extended prophase I of meiosis, and activates them by trimethylating histone 3 at lysine-4 and lysine-36 on nearby nucleosomes. Although this activation of hotspots is believed to occur on the chromatin loops, the subsequent double-strand break formation and repair occur on a proteinaceous structure known as the chromosome axis. We now show that Ewing sarcoma RNA binding protein 1 (EWSR1) is a key player in early recombination events, binding to PRDM9, promoting PRDM9-dependent histone methylation, and facilitating the linking between PRDM9-bound hotspots and the nascent chromosome axis through the meiosis-specific cohesion REC8. As a result of these activities, EWSR1 assures sufficient numbers of properly positioned crossovers in each meiosis.

developmental biology↗

bioRxiv2018

Tissue-specific trans regulation of the mouse epigenome

Although a variety of writers, readers, and erasers of epigenetic modifications are known, we have little information about the underlying regulatory systems controlling the establishment and maintenance of the epigenetic landscape, which varies greatly among cell types. Here, we have explored how natural genetic variation impacts the epigenome in mice. Studying levels of H3K4me3, a histone modification at sites such as promoters, enhancers, and recombination hotspots, we found tissue-specific trans-regulation of H3K4me3 levels in four highly diverse cell types: male germ cells, embryonic stem (ES) cells, hepatocytes and cardiomyocytes. To identify the genetic loci involved, we measured H3K4me3 levels in male germ cells in a mapping population of 60 BXD recombinant inbred lines, identifying extensive trans-regulation primarily controlled by six major histone quantitative trait loci (hQTL). These chromatin regulatory loci act dominantly to suppress H3K4me3, which at hotspots reduces the likelihood of subsequent DNA double-strand breaks. QTL locations do not correspond with enzyme known to metabolize chromatin features. Instead their locations match clusters of zinc finger genes, making these possible candidates that explain the dominant suppression of H3K4me3. Collectively, these data describe an extensive, tissue-specific set of chromatin regulatory loci that control functionally related chromatin sites.

genomics↗

bioRxiv2018

CXXC1 is redundant for normal DNA double-strand break formation and meiotic recombination in mouse

In most mammals, including mice and humans, meiotic recombination is determined by the meiosis specific histone methytransferase PRDM9, which binds to specific DNA sequences and trimethylates histone 3 at lysine-4 and lysine-36 at the adjacent nucleosomes. These actions ensure successful DNA double strand break initiation and repair that occur on the proteinaceous structure forming the chromosome axis. The process of hotspot association with the axis after their activation by PRDM9 is poorly understood. Previously, we and others have identified CXXC1, an ortholog of S. cerevisiae Spp1 in mammals, as a PRDM9 interactor. In yeast, Spp1 is a histone methyl reader that links H3K4me3 sites with the recombination machinery, promoting DSB formation. Here we investigated whether CXXC1 has a similar function in mouse meiosis. We found that CXXC1 is co-expressed and interacts with PRDM9 in mouse spermatocytes. To investigate the meiotic function of CXXC1, we created a Cxxc1 conditional knockout mouse to deplete CXXC1 before the onset of meiosis. Surprisingly, knockout mice were fertile, and the loss of CXXC1 in spermatocytes had no effect on hotspot trimethylation activity, double-strand break formation or repair. Our results demonstrate that CXXC1 is not an essential link between recombination hotspot sites and DSB machinery and that the hotspot recognition pathway in mouse is independent of CXXC1.\n\nAuthor SummaryMeiotic recombination increases genetic diversity by ensuring novel combination of alleles passing onto the next generation correctly. In most mammals, the meiotic recombination sites are determined by histone methyltransferase PRDM9. These sites subsequently become associated with the chromosome axis with the participation of additional proteins and undergo double strand breaks, which are repaired by homologous recombination. In Saccharomyces cerevisiae, Spp1 (ortholog of CXXC1) binds to methylated H3K4 and connects these sites with chromosome axis promoting DSB formation. However, our data suggest that even though CXXC1 interacts with PRDM9 in male germ cells, it does not play a crucial role in mouse meiotic recombination. These results indicate that, unlike in S. cerevisiae, a recombination initiation pathway that includes CXXC1 could only serve as a non-essential pathway in mouse meiotic recombination.

cell biology↗

bioRxiv2018

The functional reach of the hippocampal memory system to the oculomotor system.

Visual exploration is related to activity in the hippocampus (HC) and/or extended medial temporal lobe system (MTL), is influenced by stored memories, and is altered in amnesic cases. An extensive set of polysynaptic connections exists both within and between the HC and oculomotor systems such that investigating how HC responses ultimately influence neural activity in the oculomotor system, and the timing by which such neural modulation could occur is not trivial. We leveraged TheVirtualBrain, a software platform for large-scale network simulations, to model the functional dynamics that govern the interactions between the two systems in the macaque cortex. Evoked responses following the stimulation of the MTL and some, but not all, subfields of the HC resulted in observable responses in oculomotor regions, including the frontal eye fields (FEF), within the time of a gaze fixation. Modeled lesions to some MTL regions slowed the dissipation of HC signal to oculomotor regions, whereas HC lesions generally did not affect the rapid MTL activity propagation to oculomotor regions. These findings provide a framework for investigating how information represented by the HC/MTL may influence the oculomotor system during a fixation and predict how HC lesions may affect visual exploration. Author SummaryNo major account of oculomotor (eye movement) guidance considers the influence of the hippocampus (HC) and broader medial temporal lobe (MTL) system, yet it is clear that information is exchanged between the two systems. Prior experience influences current viewing, and cases of amnesia due to compromised HC/MTL function show specific alterations in viewing behaviour. By modeling large-scale network dynamics, we show that stimulation of subregions of the HC, and of the MTL, rapidly results in observable responses in oculomotor control regions, and that HC/MTL lesions alter signal propagation. These findings suggest that information from memory may readily guide visual exploration, and calls for a reconsideration of the neural circuitry involved in oculomotor guidance.

neuroscience↗

bioRxiv2018

From 1D sequence to 3D chromatin dynamics and cellular functions: a phase separation perspective

The high-order chromatin structure plays a non-negligible role in gene regulation. However, the mechanism for the formation of different chromatin structures in different cells and the sequence dependence of this process remain to be elucidated. As the nucleotide distributions in human and mouse genomes are highly uneven, we identified CGI forest and prairie genomic domains based on CGI density, which better segregates genomic elements along the genome than GC content. The genome is then divided into two sequentially, epigenetically, and transcriptionally distinct regions. These two types of megabase-sized domains spatially segregate, but to a different extent in different cell types. Overall, the forests and prairies gradually segregate from each other in development, differentiation, and senescence. The multi-scale forest-prairie spatial intermingling is cell-type specific and increases in differentiation, thus helps define the cell identity. We propose that the phase separation of the 1D mosaic sequence in space, serving as a potential driving force, together with cell type specific epigenetic marks and transcription factors, shapes the chromatin structure in different cell types and renders them distinct genomic properties. The mosaicity of the genome manifested in terms of alternative forests and prairies of a species could be related to its biological processes such as differentiation, aging and body temperature control.

biophysics↗