Biology subjects

Zuo, T.

Publications and source records attributed to Zuo, T..

14 recordsLinked to original sources

bioRxiv2025

Breakthrough infection elicits synergistic broadly neutralizing antibodies targeting non-dominant epitopes on RBD and NTD in an individual of inactivated SARS-CoV-2 vaccine

Background/ObjectivesIdentification and characterization of broadly neutralizing monoclonal antibodies from individuals exposed to SARS-CoV-2, either by infection or vaccination, can inform the development of next-generation vaccines and antibody therapeutics with pan-SARS-CoV-2 protection. MethodsThrough single B cell sorting and RT-PCR, monoclonal antibodies (mAbs) were isolated from a donor who experienced BA.5 or BF.7 breakthrough infection after three doses of inactivated vaccines. Their binding and neutralizing capacities were respectively measured with ELISA and pseudovirus-based neutralization assay. Their epitopes were mapped by competition ELISA and site- directed mutation. ResultsAmong a total of 67 spike-specific mAbs cloned from the donor, four mAbs (KXD643, KXD652, KXD681, and KXD686) can neutralize all tested SARS-CoV-2 variants from wild-type to KP.3. Moreover, KXD643, KXD652, and KXD681 belong to a clonotype encoded by IGHV5-51 and IGKV1-13, and recognize the cryptic and conserved RBD-8 epitope on the receptor-binding domain (RBD). In contrast, KXD686 is encoded by IGHV1-69 and IGKV3-20, and targets a conserved epitope (NTD Site iv) outside the antigenic supersite (NTD Site i) of the N-terminal domain (NTD). Notably, antibody cocktails containing these two groups of mAbs can neutralize SARS-CoV-2 more potently due to synergistic effects. In addition, bispecific antibodies derived from KXD643 and KXD686 demonstrate further improved neutralizing potency compared to antibody cocktails. ConclusionsThese four mAbs can be developed as candidates of pan-SARS-CoV-2 antibody therapeutics through further antibody engineering. On the other hand, vaccines designed to simultaneously elicit neutralizing antibodies towards RBD-8 and NTD Site iv have the potential to provide pan-SARS-CoV-2 protection.

immunology↗

bioRxiv2024

Longitudinal profiling of B cells primed by mRNA vaccine and recalled by Omicron variants uncovers antibodies broadly neutralizing sarbecoviruses

A key question on ancestral SARS-CoV-2 immune imprinting is to what extent imprinted B cells can develop neutralizing breadth and potency in immune recalls. Here, we longitudinally tracked B cells recognizing wild-type spike in two individuals, who were sequentially infected by Omicron variants after receiving mRNA vaccines. Functional and genetic analysis of 632 monoclonal antibodies (mAbs) from those B cells reveals that mAbs cloned after second infection have dramatically enhanced neutralizing breadth and potency, which is attributed to recall and maturation of pre-existing memory B cells. Among the 11 mAbs that potently neutralize SARS-CoV-2 variants from wild-type to KP.3, 5 mAbs are classified into public clonotypes encoded by IGHV3-53 or IGHV3-66, whereas the rest belong to a rarely reported clonotype encoded by IGHV3-74. Notably, IGHV3-74 mAbs can also broadly neutralize other sarbecoviruses by targeting a novel epitope on receptor-binding domain of spike. These results support that ancestral SARS-CoV-2 immune imprinting can be harnessed in developing pan-SARS-CoV-2 and even pan-sarbecovirus vaccines. SummaryChen et al. demonstrate that B cells imprinted by ancestral SARS-CoV-2 have tremendous potential to develop neutralizing breadth and potency in repeated immune recalls driven by Omicron variants, implicating that ancestral SARS-CoV-2 immune imprinting can be harnessed in developing pan-SARS-CoV-2 and even pan-sarbecovirus vaccines.

immunology↗

bioRxiv2024

Structural Characterization of mRNA Lipid Nanoparticles in the Presence of Intrinsic Drug-free Lipid Nanoparticles

Lipid nanoparticles (LNPs) have emerged as a versatile platform for mRNA delivery across a range of applications, including disease prevention, cancer immunotherapy, and gene editing. Structural models of mRNA-containing lipid nanoparticles (mRNA-LNPs) have also been proposed based on characterization of samples by using various advanced techniques. Among these, small angle neutron scattering (SANS) has proven essential for elucidating the lipid distribution within mRNA-LNPs, a factor crucial to both their preparation and efficacy. However, recent findings suggest that the mRNA-LNP samples prepared via commercial microfluidic techniques may contain a substantial fraction of drug-free LNPs, casting doubt on the validity of earlier structural models. In this study, we employed contrast variation SANS to characterize both drug-free LNPs and our mRNA-LNP sample, and quantified the proportion of drug-free LNPs present to be [~]30% in our mRNA-LNP sample using nano flow cytometry. By removing the contributions of drug-free LNPs from the SANS data of our mRNA-LNP sample, we were able to precisely characterize the structure of mRNA-LNPs. Consequently, we proposed structural models for both drug-free LNPs and mRNA-LNPs. Notably, our analysis revealed similar lipid distributions and shell thicknesses between the two particle types, while the solvent content in mRNA-LNPs was significantly higher, leading to a larger core size. This work not only offers a method for accurately characterizing the structure of mRNA-LNPs, but also establishes criteria for selecting appropriate analytical techniques based on the structural parameters of interest. Therefore, our findings hold significant implications for the mechanistic understanding and quality control of mRNA-based vaccines. SignificancePrecise structural determination of mRNA-containing lipid nanoparticles (mRNA-LNPs) is vital for mechanistic insights into their preparation, delivery, immunogenicity, and storage, which are critical to the development of mRNA-based vaccines. However, most previous studies overlooked the substantial presence of drug-free LNPs within these samples. Here, we identified that approximately 30% of the nanoparticles in our mRNA-LNP sample were drug-free. By integrating contrast variation small angle neutron scattering (SANS) data from both drug-free LNPs and mRNA-LNPs, we developed structural models for both particle types, and provided a guidance for characterization technique selection based on concerned structural features. Beyond mechanistic insight on structure, our approach offers a robust method for quality assessment and process monitoring in mRNA-based vaccine production.

biophysics↗

bioRxiv2024

Ancestral SARS-CoV-2 immune imprinting persists on RBD but not NTD after sequential Omicron infections

Whether Omicron exposures could overcome ancestral SARS-CoV-2 immune imprinting remains controversial. Here we analyzed B cell responses evoked by sequential Omicron infections in vaccinated and unvaccinated individuals. Plasma neutralizing antibody titers against ancestral SARS-CoV-2 and variants indicate that immune imprinting is not consistently induced by inactivated or recombinant protein vaccines. However, once induced, immune imprinting is not countered by successive Omicron challenges. We compared binding specificities, neutralizing capacities, developing origins and targeting epitopes of monoclonal antibodies from individuals with or without immune imprinting. Although receptor-binding domain (RBD) and N-terminal domain (NTD) of spike are both primary targets for neutralizing antibodies, immune imprinting only shapes antibody responses to RBD by impeding the production of Omicron-specific neutralizing antibodies while facilitating the development of broadly neutralizing antibodies. We propose that immune imprinting can be either neglected by NTD-based vaccines to induce variant-specific antibodies or leveraged by RBD-based vaccines to induce broadly neutralizing antibodies.

immunology↗

bioRxiv2023

Breakthrough infection elicits hypermutated IGHV3-53/3-66 public antibodies with broad and potent neutralizing activity against SARS-CoV-2 variants including BQ and XBB lineages

The rapid emergence of SARS-CoV-2 variants of concern (VOCs) calls for efforts to study broadly neutralizing antibodies elicited by infection or vaccination so as to inform the development of vaccines and antibody therapeutics with broad protection. Here, we identified two convalescents of breakthrough infection with relatively high neutralizing titers against all tested viruses including BQ and XBB lineages. Among 50 spike-specific monoclonal antibodies (mAbs) cloned from their B cells, the top 6 neutralizing mAbs (KXD01-06) belong to previously defined IGHV3-53/3-66 public antibodies. Although most antibodies in this class are dramatically escaped by VOCs, KXD01-06 exhibit broad neutralizing capacity with the IC50s of KXD01 ranging from 0.011[~]0.059g/ml. Deep mutational scanning reveals that KXD01-06 target highly conserved sites on RBD including D420, Y421, L455, F456, A475 and N487. Genetic and functional analysis further indicates that the extent of somatic hypermutation is critical for the breadth of IGHV3-53/3-66 public antibodies. Overall, we discovered and characterized IGHV3-53/3-66 public antibodies with broad and potent neutralizing activity against SARS-CoV-2, which provides rationale for novel vaccines and antibody therapeutics based on this class of antibodies.

immunology↗

bioRxiv2022

Treg cells drive MYCN-mediated immunosuppression and tumor aggressiveness in high-risk neuroblastoma

Solid tumors, especially those with aberrant MYCN activation, harbor an immunosuppressive microenvironment to fuel malignant growth and trigger treatment resistance1,2, yet the underlying mechanisms are elusive and effective strategies to tackle this challenge are lacking. Here we demonstrated the crucial role of T regulatory (Treg) cells in MYCN-mediated immune repression and tumor aggression using high-risk neuroblastoma (NB) as a model system. Human MYCN-activated NB attracts CD4+ Treg cells, which are also found enriched in MYCN-high primary patient samples. Zebrafish MYCN-overexpressing neural crests recruit Cd4+ cells before tumor formation and induce an immunosuppressive microenvironment, thereby promoting tumor onset and progression. Strikingly, disruption of Treg cells through depletion of forkhead box protein 3a restores anti-tumor immunity and impairs NB development. Together, our studies establish Treg cells as the key driver of MYCN-mediated immunosuppression and tumor aggressiveness, providing mechanistic insights and therapeutic implications.

cancer biology↗

bioRxiv2022

Using NAMs to characterize chemical bioactivity at the transcriptomic, proteomic and phosphoproteomic levels

Omic-based technologies are of particular interest and importance for non-animal chemical hazard and risk characterization based on the premise that any apical endpoint change must be underpinned by some alterations measured at the omic levels. In this work we studied cellular responses to caffeine and coumarin by generating and integrating multi-omic data from transcriptomic, proteomic and phosphoproteomic experiments. We have shown that the methodology presented here is able to capture the complete chain of events from the first compound-induced changes at the phosphoproteome level to changes in gene expression induced by transcription factors and lastly to changes in protein abundance that further influence changes at the cellular level. In HepG2 cells we found the metabolism of lipids and general cellular stress to be dominant biological processes in response to caffeine and coumarin exposure, respectively. The phosphoproteomic changes were detected early in time, at very low concentrations and provided a fast adaptive cellular response to chemical exposure. Changes in protein abundance were found much less frequently than the transcriptomic changes and can be used, together with the transcriptomic changes, to facilitate a more complete understanding of pathway responses to chemical exposure. GRAPHIC ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=169 SRC="FIGDIR/small/492410v1_ufig1.gif" ALT="Figure 1"> View larger version (48K): org.highwire.dtl.DTLVardef@1e348c8org.highwire.dtl.DTLVardef@bf50c5org.highwire.dtl.DTLVardef@4fea36org.highwire.dtl.DTLVardef@998cb8_HPS_FORMAT_FIGEXP M_FIG C_FIG

pharmacology and toxicology↗

bioRxiv2022

The phosphoproteome is a first responder in tiered cellular adaptation to chemical stress followed by proteomics and transcriptomics alteration

Next-generation risk assessment for environmental chemicals and ingredients in consumer products involves a weight of evidence (WoE) framework integrating a suite of new approach methodologies (NAMs) based on points of departure (PoD) obtained from in vitro assays. Omics techniques provide broad coverages of the molecular toxicity pathway space. Transcriptomics assays especially play a leading role by providing relatively conservative PoDs in comparison with apical endpoints. However, it is unclear whether and how parameters measured using other omics technicquesparticipate in the cellular response to chemical perturbations, especially at exposure levels below the transcriptomically defined PoD. Multi-omics coverage may provide additional sensitive or confirmative biomarkers to complement and reduce the uncertainty in safety decisions made using targeted and transcriptomics assays. In the present study, we compared changes in transcriptomics, proteomics and phosphoproteomics with two prototype compounds, coumarin, as a main study and doxorubicin, as a complementary study to understand the sensitivity of the three omics techniques in response to chemically-induced changes in HepG2 and AC16 cells. With measurements obtained for multiple chemical concentrations and time points, we demonstrated that, compared with proteomics and transcriptomics, phosphoproteomics alterations occur not only earlier in time as expected, but also at much lower chemical concentrations and hence are proximal to the very early effects induced by chemical exposure. The phosphoproteomics changes appear to approach maximum when the transcriptomics alterations begin to be initiated. The results are consistent with a tiered framework of cellular response to chemical insults, where posttranslational modification of preexisting proteins is first seen before transcriptomics induction is engaged to launch a more energy-expensive defense that defines a useful PoD. We conclude that as the cost becomes more affordable, proteomics covering posttranslational modifications can be utilized to provide a more complete coverage of chemical-induced cellular alteration and supplement transcriptomics-based health safety decision making.

pharmacology and toxicology↗

bioRxiv2021

Memory B cell repertoire for recognition of evolving SARS-CoV-2 spike

Memory B cell reserves can generate protective antibodies against repeated SARS-CoV-2 infections, but with an unknown reach from original infection to antigenically drifted variants. We charted memory B cell receptor-encoded monoclonal antibodies (mAbs) from 19 COVID-19 convalescent subjects against SARS-CoV-2 spike (S) and found 7 major mAb competition groups against epitopes recurrently targeted across individuals. Inclusion of published and newly determined structures of mAb-S complexes identified corresponding epitopic regions. Group assignment correlated with cross-CoV-reactivity breadth, neutralization potency, and convergent antibody signatures. mAbs that competed for binding the original S isolate bound differentially to S variants, suggesting the protective importance of otherwise-redundant recognition. The results furnish a global atlas of the S-specific memory B cell repertoire and illustrate properties conferring robustness against emerging SARS-CoV-2 variants.

immunology↗

bioRxiv2021

Transposon-induced inversions activate gene expression in Maize pericarp

Chromosomal inversions can have considerable biological and agronomic impacts including disrupted gene function, change in gene expression and inhibited recombination. Here we describe the molecular structure and functional impact of six inversions caused by Alternative Transpositions between p1 and p2 genes responsible for floral pigmentation in maize. In maize line p1-wwB54, the p2 gene is expressed in anther and silk but not in pericarp, making the kernels white. We identified inversions in this region caused by transposition of Ac and fractured Ac (fAc) transposable elements. These inversions change the position of a p1 enhancer and activate the expression of p2 in the kernel pericarp, resulting in red kernel color. We hypothesize that these inversions place the p2 gene promoter near a p1 gene enhancer, thereby activating p2 expression in kernel pericarp.

genetics↗

bioRxiv2020

Ectopic expression of a maize gene is induced by Composite Insertions generated by Alternative Transposition

Transposable elements (TEs) are DNA sequences that can mobilize and proliferate throughout eukaryotic genomes. Previous studies have shown that in plant genomes, TEs can influence gene expression in various ways such as inserting in introns or exons to alter transcript structure and content, and providing novel promoters and regulatory elements to generate new regulatory patterns. Furthermore, TEs can also regulate gene expression at the epigenetic level by modifying chromatin structure, changing DNA methylation status and generating small RNAs. In this study, we demonstrated that Ac/fAc transposable elements are able to induce ectopic gene expression by duplicating and shuffling enhancer elements. Ac/fAc elements belong to the hAT family of Class II TEs. They can undergo standard transposition events, which involve the two termini of a single transposon, or alternative transposition events which involve the termini of two different, nearby elements. Our previous studies have shown that alternative transposition can generate various genome rearrangements such as deletions, duplications, inversions, translocations and Composite Insertions (CIs). We identified over 50 independent cases of CIs generated by Ac/fAc alternative transposition and analyzed 10 of them in detail. We show that these CIs induced ectopic expression of the maize pericarp color 2 (p2) gene, which encodes a Myb-related protein. All the CIs analyzed contain sequences including a transcriptional enhancer derived from the nearby p1 gene, suggesting that the CI-induced activation of p2 is effected by mobilization of the p1 enhancer. This is further supported by analysis of a mutant in which the CI is excised and p2 expression is lost. These results show that alternative transposition events are not only able to induce genome rearrangements, but also generate Composite Insertions that can control gene expression. SummaryWhen Barbara McClintock originally identified and characterized Transposable Elements (TEs) in maize, she termed them "Controlling Elements" due to their effects on gene expression. Here we show that maize Ac/Ds TEs can acquire a genomic enhancer and generate Composite Insertions (CIs) that activate expression of a nearby gene. CIs are structurally variable elements that include TE termini enclosing sequences from an original donor locus, and are formed when the termini of two nearby TEs transpose during S phase from a replicated to unreplicated site. In this way, TEs may acquire genomic enhancers to generate Controlling Elements as described by McClintock.

genetics↗

bioRxiv2020

siRNA-mediated de novo silencing of Ac/Ds transposons is initiated by alternative transposition in maize

Although Transposable Elements (TEs) comprise a major fraction of many higher eukaryotic genomes, most TEs are silenced by host defense mechanisms. The means by which otherwise active TEs are recognized and silenced remains poorly understood. Here we analyzed two independent cases of spontaneous silencing of the active maize Ac/Ds transposon system. This silencing was initiated by Alternative Transposition (AT), a type of aberrant transposition event that engages the termini of two nearby separate TEs. AT during DNA replication can generate Composite Insertions (CIs) that contain inverted duplications of the transposon sequences. We show that the inverted duplications of two CIs are transcribed to produce dsRNAs that trigger the production of two distinct classes of siRNAs: a 24-nt class complementary to the TE terminal inverted repeats (TIRs) and non-coding sub-terminal regions, and a 21-22 nt class corresponding to the TE transcribed regions. Plants containing these siRNA-generating CIs exhibit decreased levels of Ac transcript and heritable repression of Ac/Ds transposition. This study documents the first case of TE silencing attributable to transposon self-initiated AT and may represent a general initiating mechanism for silencing of DNA transposons. Article summaryTransposable Elements (TEs) are often silenced by their hosts, but how TEs are initially recognized for silencing remains unclear. Here we describe two independent loci that induce de novo heritable silencing of maize Ac/Ds transposons. Plants containing these loci produce dsRNA and Ac-homologous small interfering RNAs, and exhibit decreased levels of Ac transcript and heritable repression of Ac/Ds transposition. We show that these loci comprise inverted duplications of TE sequences generated by Alternative Transposition coupled with DNA re-replication. This study documents the first case of transposon silencing induced by AT and may represent a general initiating mechanism for TE silencing.

genetics↗

bioRxiv2019

BR-bodies provide selectively permeable condensates that stimulate mRNA decay and prevent release of decay intermediates

Biomolecular condensates play a key role in organizing RNAs and proteins into membraneless organelles. Bacterial RNP-bodies (BR-bodies) are a biomolecular condensate containing the RNA degradosome mRNA decay machinery, but the biochemical function of such organization remains poorly defined. Here we define the RNA substrates of BR-bodies through enrichment of the bodies followed by RNA-seq. We find that long, poorly translated mRNAs, small RNAs, and antisense RNAs are the main substrates, while rRNA, tRNA, and other conserved ncRNAs are excluded from these bodies. BR-bodies stimulate the mRNA decay rate of enriched mRNAs, helping to reshape the cellular mRNA pool. We also observe that BR-body formation promotes complete mRNA decay, avoiding the build-up of toxic endo-cleaved mRNA decay intermediates. The combined selective permeability of BR-bodies for both, enzymes and substrates together with the stimulation of the sub-steps of mRNA decay provide an effective organization strategy for bacterial mRNA decay.

biochemistry↗

bioRxiv2018

Leveraging mutational burden for complex trait prediction in sorghum

Sorghum (Sorghum bicolor (L.) Moench) is a major staple food cereal for millions of people worldwide. The sorghum genome, like other species, accumulates deleterious mutations, likely impacting its fitness. Though selection keeps deleterious mutations rare, their complete removal from the genome is impeded due to lack of recombination, drift, and their coupling with favorable loci. To study how deleterious mutations impact agronomic phenotypes, we identified putative deleterious mutations among ~5.5M segregating variants of 229 diverse sorghum lines. We provide the whole-genome estimate of the deleterious burden in sorghum, showing that about 33% of nonsynonymous substitutions are putatively deleterious. The pattern of mutation burden varies appreciably among racial groups; the caudatum shows higher mutation burden while the guinea has lower burden. Across racial groups, the mutation burden correlated negatively with biomass, plant height, Specific Leaf Area (SLA), and tissue starch content, suggesting deleterious burden decreases trait fitness. Putatively deleterious variants explain roughly half of the genetic variance. However, there is only moderate improvement in total heritable variance explained for biomass (7.6%) and plant height (5.2%). There is no advantage in total heritable variance for SLA and starch. The contribution of putatively deleterious variants to phenotypic diversity therefore appears to be dependent on the genetic architecture of traits. Overall, our results suggest that including putatively deleterious variants in models do not significantly improve breeding accuracy because of extensive linkage. However, knowledge of deleterious variants could be leveraged for sorghum breeding through genome editing.

genetics↗