Biology subjects

Kong, X.

Publications and source records attributed to Kong, X..

At least 109 records · Page 6Linked to original sources

bioRxiv2019

Circulating bacterial DNA as a tool towards non-invasive biomarkers for colorectal cancer and adenoma

ObjectiveThe gut microbiota is closely associated with colorectal neoplasia. While most metagenomics studies utilized fecal samples, circulating microbial DNA in colorectal neoplasia patients remained unexplored. This study aimed to characterize microbial DNA in plasma samples and build a machine learning model for colorectal neoplasia early detection.\n\nDesignWe performed whole genome sequencing of plasma samples from 25 colorectal cancer (CRC) patients, 10 colorectal adenoma (CRA) patients and 22 healthy controls (HC). Microbial DNA was obtained by removing the host genome and relative abundance was measured by mapping reads into microbial genomes. Significant biomarker species were identified in the discovery cohort and built into a random forest model, which was tested in the validation cohort.\n\nResultsIn the discovery cohort, there were 127 significant species between CRC patients and HC. Based on the random forest model, 28 species were selected from the discovery cohort (AUC=0.944) and yielded an AUC of 1 in the validation cohort. Interestingly, relative abundance of most biomarker species in CRA patients were between CRC patients and HC with a trend towards CRC patients. Furthermore, pathway enrichment analysis also showed similar pattern where CRA patients had intermediate relative abundance of significant pathways compared to CRC patients and HC. Finally, species network analysis revealed that CRC and HC displayed distinct patterns of species association.\n\nConclusionsWe demonstrated characteristic alteration of circulating bacterial DNA in colorectal neoplasia patients. The predictive model accurately distinguished CRC and CRA from HC, suggesting the utility of circulating bacterial biomarkers as a non-invasive tool for colorectal neoplasia screening and early diagnosis.

cancer biology↗

bioRxiv2019

Multi-disease associated risk locus in IL6 represses the anti-inflammatory gene GPNMB through chromatin looping and recruiting MEF2-HDAC complex

We have previously revealed a genetic association between Takayasu arteritis and a non-coding genetic variant in an enhancer region within IL6 (rs2069837 A/G). The risk allele in this variant (allele A) has a protective effect against chronic viral infection and cancer. Using a combination of experimental and bioinformatics tools, we identified the monocyte/macrophage anti-inflammatory gene GPNMB, [~]520kb away, as a target gene regulated by rs2069837. We revealed preferential recruitment of myocyte enhancer factor 2-histone deacetylase (MEF2-HDAC) repressive complex to the Takayasu arteritis risk allele. Further, we demonstrated suppression of GPNMB expression in monocyte-derived macrophages from healthy individuals with the AA compared to AG genotype, which was reversed by histone deacetylase inhibition. Our data suggest that the A allele in rs2069837 represses the expression of GPNMB by recruiting MEF2-HDAC complex, enabled through a long-range intra-chromatin looping mediated by CTCF. Suppression of this anti-inflammatory gene might mediate increased susceptibility in Takayasu arteritis and enhance protective immune responses in chronic infection and cancer. Our data highlight long-range chromatin interactions in functional genomic studies.

genetics↗

bioRxiv2019

Risk of multiple primary cancer diagnosis over age in families of Li-Fraumeni syndrome: a single institution perspective

Li-Fraumeni syndrome (LFS) is a rare autosomal dominant disorder associated with TP53 germline mutations and an increased lifetime risk of multiple primary cancers (MPC). Penetrance estimation of time to first and second primary cancer within LFS remains challenging due to limited data availability and the difficulty of accounting for the characteristic effects of a primary cancer on the penetrance of a second primary cancer. Using a recurrent events survival modeling approach, we estimated penetrance for both first and second primary cancer diagnosis from a pediatric sarcoma cohort (number families=189; Single primary cancer, or SPC=771; MPC=87). We then validated the risk prediction performance using an independent clinical cohort of TP53 tested individuals from MD Anderson (SPC=102; MPC=58). Our penetrance estimates are dependent on TP53 genotype, sex, and, importantly, age of diagnosis (AoD) for the first PC. We observed the later the AoD is, the shorter the gap time is for this person to be diagnosed with a second PC. We achieved an Area under the ROC curve (AUC) of 0.77 for predicting individual outcomes of MPC vs. SPC. In summary, we have provided the first set of penetrance estimates for SPC and MPC for TP53 mutation carriers, and demonstrated its accuracy for cancer risk assessment. Our open-source R package, LFSPRO, provides future risk estimates for SPC and MPC among TP53 germline mutation carriers.\n\nSignificanceOur tool can be used to support clinical counseling of LFS cancer survivors for better health management.

genetics↗

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↗

bioRxiv2019

Identification of functional variants for platelet CD36 expression by Massively Parallel Reporter Assay.

CD36 is a platelet membrane glycoprotein whose engagement with oxidized low-density lipoprotein (oxLDL) results in platelet activation. The CD36 gene has been associated with platelet count, platelet volume, as well as lipid levels and CVD risk by genome-wide association studies. Platelet CD36 expression levels have been shown to be associated with both the platelet oxLDL response and an elevated risk of thrombo-embolism. Several genomic variants have been identified as associated with platelet CD36 levels, however none have been conclusively demonstrated to be causative. We screened 81 expression quantitative trait loci (eQTL) single nucleotide polymorphisms (SNPs) associated with platelet CD36 expression by a Massively Parallel Reporter Assay (MPRA) and analyzed the results with a novel Bayesian statistical method. Ten eQTLs located in a 35kb region upstream of the CD36 transcriptional start site demonstrated significant transcription shifts between their minor and major allele in the MPRA assay. Of these, rs2366739 and rs1194196, separated by only 20bp, were confirmed by luciferase assay to alter transcriptional regulation. In addition, electromobility shift assays demonstrated differential DNA:protein complex formation between the two alleles of this locus. Furthermore, deletion of the genomic locus by CRISPR/Cas9 in K562 cells results in upregulation of CD36 transcription. These data indicate that we have identified a variant that regulates expression of CD36, which in turn affects platelet function. To assess the clinical relevance of our findings we used the PhenoScanner tool, which aggregates large scale GWAS findings; the results reinforce the clinical relevance of our variants and the utility of the MPRA assay. The study demonstrates a generalizable paradigm for functional testing of genetic variants to inform mechanistic studies, support patient management and develop precision therapies. Author SummaryPlatelets are anucleate cells that are best known as regulators of vascular hemostasis and thrombosis but also play important roles in cancer, angiogenesis, and inflammation. CD36 is a platelet surface marker that can activate platelet in response to oxidized low density lipoprotein (oxLDL). CD36 has been associated with numerous cardiovascular traits in human including blood lipid levels, platelet count, and cardiovascular disease prevalence in human genetic studies. Human variability in platelet CD36 levels are associated with the platelet response to oxLDL. However, the genetic mechanisms responsible for the variability of CD36 levels are unknown. We examined 81 genetic variants associated with CD36 levels for functionality using a high-throughput assay. Of the ten variants that were identified in that assay, one doublet, rs2366739 and rs1194196, were confirmed using additional molecular and cellular assays. Deletion of the genomic region containing rs2366739 and rs1194196 resulted in overexpression of CD36 in a cell culture system. This finding indicates a control locus which can serve as a potential target in modulating CD36 expression and altering platelet function in cardiovascular disease.

genomics↗

bioRxiv2019

A chromosome-level genome of black rockfish, Sebastes schlegelii, provides insights into the evolution of live birth

Black rockfish (Sebastes schlegelii) is a teleost species where eggs are fertilized internally and retained in the maternal reproductive system, where they undergo development until live birth (termed viviparity). In the present study, we report a chromosome-level black rockfish genome assembly. High-throughput transcriptome analysis (RNA-seq and ATAC-seq), coupled with in situ hybridization (ISH) and immunofluorescence, identify several candidate genes for maternal preparation, sperm storage and release, and hatching. We propose that zona pellucida (ZP) genes retain sperm at the oocyte envelope, while genes in two distinct astacin metalloproteinase subfamilies serve to release sperm from the ZP and free the embryo from chorion at pre-hatching stage. Finally, we present a model of black rockfish reproduction, and propose that the rockfish ovarian wall has a similar function to uterus of mammals. Taken together, these genomic data reveal unprecedented insights into the evolution of an unusual teleost life history strategy, and provide a sound foundation for studying viviparity in non-mammalian vertebrates and an invaluable resource for rockfish ecology and evolution research.

genomics↗

bioRxiv2019

Comparative analysis of mitochondrial genome between UG93A and UG93B reveal common feature of 5’-end heterogeneity in mitochondrial genes of kenaf

Plant cytoplasmic male sterility (CMS) being maternal phenomenon trait that result from pollen abortion and closely linked with mitochondrial DNA rearrangement in many crops including kenaf. However, the molecular mechanism in kenaf is poorly known. In present work, we described the mitochondrial genome in isonuclear CMS line UG93A and its maintainer line UG93B. Findings of the current study revealed that a total of 398 SNPs and 230 InDels were identified in UG93A mtDNA. Total of 26 SNPs variations and three InDels were identified in the coding region of atp6, indicating its active role in mitochondrial genome re-arrangement. Northern blot analysis showed that the transcripts of atp1, atp4, atp6, cox3 and sdh4 in F1 were consistent with UG93A but different for UG93B. The transcript of atp9 was found similar between UG93B and F1 while different for UG93A, which depict that atp9 may be regulated by nuclear genes in F1 hybrid. The expression of atp9 in UG93A was substantially lower compared with UG93B, suggesting its key role for energy supplying in microspore development of kenaf. Circularized RNA (CR)-RT-PCR revealed that mitochondrial RNAs with heterogeneous 5-ends but uniform 3 - ends are common feature in kenaf mitochondrial genes, and the promoter architecture analysis showed that the promoter sequences in kenaf mitochondrial genome are highly diverged in comparison to those in other plants. Our data highlight that the translation of mitochondrial genes in kenaf is closely associated with heterogeneity of the 5-end of plant mRNA. The present result provides the basic information in understanding the transcription of kenaf mitochondrial genome and can be used as reference in other plants.

molecular biology↗

bioRxiv2019

LncRNA Dnmt3aos regulates Dnmt3a expression leading to aberrant DNA methylation in macrophage polarization

Long non-coding RNAs (lncRNAs) play key roles in various biological processes. However, the roles of lncRNAs in macrophage polarization remain largely unexplored. In this study, thousands of lncRNAs were identified that are differentially expressed in distinct polarized bone marrow-derived macrophages (BMDMs). Among them, Dnmt3aos (DNA methyltransferase 3A, opposite strand), as a known lncRNA, locates on the antisense strand of Dnmt3a. Functional experiments further confirmed that Dnmt3aos were highly expressed in M(IL-4) macrophages and participated in the regulation of Dnmt3a expression, and played a key role in macrophage polarization. The DNA methylation profiles between the Dnmt3aos knockdown group and the control group in M(IL-4) macrophages were determined by MeDIP-seq technique for the first time, and the Dnmt3aos-Dnmt3a axis-mediated DNA methylation modification-regulated macrophage polarization related gene IFN-{gamma} was identified. Our study will help to enrich our knowledge of the mechanism of macrophage polarization and will provide new insights for immunotherapy in macrophage-associated diseases.

cell biology↗

bioRxiv2018

‘Chromatic’ Neuronal Jamming in a Primitive Brain

Jamming models developed in inanimate matter have been widely used to describe cell packing in tissues1-7, but predominantly neglect cell diversity, despite its prevalence in biology. Most tissues, animal brains in particular, comprise a mix of many cell types, with mounting evidence suggesting that neurons can recognize their respective types as they organize in space8-11. How cell diversity revises the current jamming paradigm is unknown. Here, in the brain of the flatworm planarian Schmidtea mediterranea, which exhibits remarkable tissue plasticity within a simple, quantifiable nervous system12-16, we identify a distinct packing state, chromatic jamming. Combining experiments with computational modeling, we show that neurons of distinct types form independent percolating networks barring any physical contact. This jammed state emerges as cell packing configurations become constrained by cell type-specific interactions and therefore may extend to describe cell packing in similarly complex tissues composed of multiple cell types.

biophysics↗

bioRxiv2018

Ulinastatin prevents hyperoxia-induced lung injury on newborn rats by downregulating TNF-α and inhibiting macrophage infiltration

Ulinastatin (UTI) can support protection for several organs through inhibition of the release of inflammatory factors, absorbing oxygen radicals, and inhibition the progression of fibrosis. However, whether UTI has effect on hyperoxia-induced lung injury is still unclear. In this study, the effects of UTI treatment on newborn rats suffering hyperoxia-induced lung injury were examined. The results demonstrated that UTI treatment significantly attenuated the wet/dry weight ratio, downregulated the levels of tumor necrosis factor- (TNF-), inhibited macrophage infiltration, and improved the average weight of rats. The most significant changes were observed in high-dose UTI treatment group. However, UTI had no effect on pulmonary superoxide dismutase (SOD) and malondialdehyde (MDA) levels. Our results suggested that Ulinastatin prevents hyperoxia-induced lung injury on newborn rats by downregulating TNF- and inhibiting macrophage infiltration.

pathology↗

bioRxiv2018

Single-nucleus RNA-seq identifies divergent populations of FSHD2 myotube nuclei

FSHD is characterized by the misexpression of DUX4 in skeletal muscle. However, DUX4 is lowly expressed in patient samples and analysis of the consequences of DUX4 expression has largely relied on artificial overexpression. To better understand the native expression profile of DUX4 and its targets, we performed pooled RNA-seq differentiation time-course in FSHD2 patient-derived primary myoblasts and identified early-and late-induced sets of FSHD-associated genes. Using single-cell and single-nucleus RNA-seq on FSHD2 myoblasts and myotubes respectively, we captured DUX4 expression in single-nuclei and found that only some DUX4 targets are coexpressed. We identified two populations of FSHD myotube nuclei with distinct transcriptional profiles. One population is highly enriched with DUX4 and FSHD related genes, including the DUX4 paralog DUXA ("FSHD-Hi"). The other population has no expression of DUX4 and expresses low amounts of FSHD related genes ("FSHD-Lo"), but is marked by the expression of CYTL1 and CHI3L1. "FSHD-Hi" myotube nuclei upregulated a set of transcription factors (TFs) that may form a self-sustaining network of gene dysregulation, which perpetuates this disease after DUX4 is no longer expressed.

genomics↗

bioRxiv2018

Miniaturized Permeability Assay for CRISPR-Mediated Genome-Wide Screens

The impermeability of the luminal endothelial cell monolayer is crucial for the normal performance of the vascular and lymphatic systems. A key to this function is the integrity of the monolayers intercellular junctions. The known repertoire of junction-regulating genes is incomplete. Current permeability assays are incompatible with high-throughput genome-wide screens that could identify these genes. To overcome these limitations, we designed a new permeability assay that consists of cell monolayers grown on [~]150 m microcarriers. Each microcarrier functions as a miniature individual assay of permeability (MAP). We demonstrate that false-positive results can be minimized, and that MAP sensitivity to thrombin-induced increase in monolayer permeability is similar to the sensitivity of the measurement of impedance. We validate the assay by showing that the expression of single guide RNAs (sgRNAs) that target genes encoding known thrombin signaling proteins blocks effectively thrombin-induced junction disassembly, and that MAPs carrying such cells can be separated effectively by a fluorescent probe from those that carry cells expressing non-targeting sgRNAs. These results indicate that MAPs are suitable for high-throughput experimentation and for genome-wide screens for genes that mediate the disruptive effect of thrombin on endothelial cell junctions.

cell biology↗

bioRxiv2018

Spatiotemporal expression of FRIGIDA modulate flowering time in Arabidopsis thaliana

FRIGIDA (FRI) as the major regulator of flowering time in Arabidopsis accessions can activate its target FLOWERING LOCUS C (FLC) to delay flowering before vernalization. Besides FLC, other FRI targets also exist in Arabidopsis. Although leaves sense environmental cues to modulate flowering time, it is not known if roots also regulate the floral transition. In this study, we investigated the spatiotemporal effect of FRI on flowering time. Local expression of FRI in the phloem and leaves activated FLC to delay flowering. Furthermore, we found that local expression of FRI in the roots also delayed flowering by activating other targets MADS AFFECTING FLOWERING4 (MAF4) and MAF5 in the roots. Graft and genetic experiments revealed that the spatial expression of FRI in the root might generate a mobile signal, which is transmitted from roots to shoot and antagonizes the FT signal to delay flowering. Specifically expressing FRI in the embryo efficiently delayed flowering, even expressing FRI as early as pro-embryo stage is enough to upregulate FLC expression to delay flowering. Together, our findings confirm the spatiotemporal effect of FRI on delaying flowering, and propose that root tissue also perceives the flowering signal to fine-tune the flowering time through MAF4/5 as novel targets of FRI.\n\nHighlightRoot FRIDIGA activated the novel targets MAF4/5 to delay flowering; Temporal expressing FRIGIDA at as early as pro-embryo stage is efficient to delay flowering.

plant biology↗

bioRxiv2018

Dynamic mode decomposition of resting-state and task fMRI

Component analysis is a powerful tool to identify dominant patterns of interactions in multivariate datasets. In the context of fMRI data, methods such as principal component analysis or independent component analysis have been used to identify the brain networks shaping functional connectivity (FC). Importantly, these approaches are static in the sense that they ignore the temporal information contained in fMRI time series. Therefore, the corresponding components provide a static characterization of FC. Building upon recent findings suggesting that FC dynamics encode richer information about brain functional organization, we use a dynamic extension of component analysis to identify dynamic modes (DMs) of fMRI time series. We demonstrate the feasibility and relevance of this approach using resting-state and motor-task fMRI data of 730 healthy subjects of the Human Connectome Project (HCP). In resting-state, dominant DMs have strong resemblance with classical resting-state networks, with an additional temporal characterization of the networks in terms of oscillatory periods and damping times. In motor-task conditions, dominant DMs reveal interactions between several brain areas, including but not limited to the posterior parietal cortex and primary motor areas, that are not found with classical activation maps. Finally, we identify two canonical components linking the temporal properties of the resting-state DMs with 158 behavioral and demographic HCP measures. Altogether, these findings illustrate the benefits of the proposed dynamic component analysis framework, making it a promising tool to characterize the spatio-temporal organization of brain activity.

neuroscience↗

bioRxiv2018

Mate-pair Library Construction with Controlled Polymerization Enables Comprehensive Structural Rearrangement Detection

The diversity of disease presentations warrants one single assay for detection and delineation of various genomic disorders. Herein, we describe a gel-free and biotin-capture-free mate-pair method through coupling Controlled Polymerizations by Adapter-Ligation (CP-AL). We first demonstrated the feasibility and ease-of-use in monitoring DNA nick-translation and primer extension by limiting the nucleotide input. By coupling these two controlled polymerizations by a reported non-conventional adapter ligation reaction 3 branch ligation, we evidenced that CP-AL significantly increased DNA-circularization efficiency (by 4-fold) and was applicable for different sequencing methods but at a faction of current cost. Its advantages were further demonstrated by fully elimination of small-insert-contaminated (by 39.3-fold) with a ~50% increment of physical coverage, and producing uniform genome/exome coverage and the lowest chimeric rate. It achieved single-nucleotide variants detection with sensitivity and specificity up to 97.3 and 99.7%, respectively, compared with data from small-insert libraries. In addition, this method can provide a comprehensive delineation of structural rearrangements, evidenced by a potential diagnosis in a patient with oligo-atheno-terato-spermia. Moreover, it enables accurate mutation identification by integration of genomic variants from different aberration types. Overall, it provides a potential single-integrated solution for detecting various genomic variants, facilitating a genetic diagnosis in human diseases.

genomics↗

bioRxiv2018

Natural selection on TMPRSS6 associated with the blunted erythropoiesis and improved blood viscosity in Tibetan Pigs

Tibetan pigs, indigenous to Tibetan plateau, are well adapted to hypoxia. So far, there have been not any definitively described genes and functional sites responsible for hypoxia adaptation for the Tibetan pig. Here we conducted resequencing of the nearly entire genomic region (40.1 kb) of the candidate gene TMPRSS6 (Transmembrane protease, serine 6) associated with hemoglobin concentration (HGB) and red blood cell count (RBC) in 40 domestic pigs and 40 wild boars from five altitudes along the Tea-horse ancient road and identified 708 SNPs, in addition to an indel (CGTG/----) in the intron 10. Both the CGTG deletion frequency and the pairwise r2 linkage disequilibrium showed an increase with elevated altitudes in 838 domestic pigs from five altitudes, suggesting that TMPRSS6 has been under Darwinian positive selection. As the conserved core sequence of hypoxia-response elements (HREs), the deletion of CGTG in Tibetan pigs decreased the expression levels of TMPRSS6 mRNA and protein in the liver revealed by real-time quantitative PCR and western blot, respectively. To explore whether reduced TMPRSS6 expression level could improve blood viscosity, the relationship between CGTG indel and hematologic and hemorheologic parameters in 482 domestic pigs from continuous altitudes was detected and dissected a genetic effect on reducing HGB by 13.25g/L in Gongbogyamda Tibetan pigs and decreasing MCV by 4.79 fl in Diqing Tibetan pigs. In conclusion, the CGTG deletion of TMPRSS6 resulted in lower HGB and smaller MCV, thereby blunting erythropoiesis and improving blood viscosity as well as erythrocyte deformability.

physiology↗

bioRxiv2018

NAD consumption by PARP1 in response to DNA damage triggers metabolic shift critical for damaged cell survival

DNA damage signaling is critical for the maintenance of genome integrity and cell fate decision. Poly(ADP-ribose) polymerase 1 (PARP1) is a DNA damage sensor rapidly activated in a damage dose and complexity-dependent manner playing a critical role in the initial chromatin organization and DNA repair pathway choice at damage sites. However, the cell-wide consequence of its activation in damaged cells is not well delineated. Using the phasor approach to fluorescence lifetime imaging microscopy (FLIM) and fluorescence-based biosensors in combination with laser microirradiation, we found a rapid cell-wide increase of the bound/free NADH ratio in response to nuclear DNA damage, which is triggered by NAD+ depletion by PARP activation. This change is linked to the metabolic balance shift to oxidative phosphorylation (oxphos) over glycolysis. Inhibition of the respiratory chain resulted in rapid PARP-dependent ATP reduction and intracellular acidification, and eventually, PARP-dependent, AIF-independent, apoptosis indicating that oxphos becomes critical for damaged cell survival. The results reveal the novel pro-survival effect of PARP activation through a change in cellular metabolism, and demonstrate how unique applications of advanced fluorescence imaging technologies in combination with laser microirradiation can be a powerful tool to interrogate damage-induced metabolic changes at high spatiotemporal resolution in a live cell.

cell biology↗

bioRxiv2018

Negative Cooperativity between Gemin2 and RNA Determines RNA Selection and Release of the SMN Complex in snRNP Assembly

The assembly of snRNP cores, in which seven Sm proteins, D1/D2/F/E/G/D3/B, form a ring around the nonameric Sm site of snRNAs, is the early step of spliceosome formation and essential to eukaryotes. It is mediated by the PMRT5 and SMN complexes sequentially in vivo. SMN deficiency causes neurodegenerative disease spinal muscular atrophy (SMA). How the SMN complex assembles snRNP cores is largely unknown, especially how the SMN complex achieves high RNA assembly specificity and how it is released. Here we show, using crystallographic and biochemical approaches, that Gemin2 of the SMN complex enhances RNA specificity of SmD1/D2/F/E/G via a negative cooperativity between Gemin2 and RNA in binding SmD1/D2/F/E/G. Gemin2, independent of its N-tail, constrains the horseshoe-shaped SmD1/D2/F/E/G from outside in a physiologically relevant, narrow state, enabling high RNA specificity. Moreover, the assembly of RNAs inside widens SmD1/D2/F/E/G, causes the release of Gemin2/SMN allosterically and allows SmD3/B to join. The assembly of SmD3/B further facilitates the release of Gemin2/SMN. This is the first to show negative cooperativity in snRNP assembly, which provides insights into RNA selection and the SMN complexs release. These findings reveal a basic mechanism of snRNP core assembly and facilitate pathogenesis studies of SMA.

molecular biology↗