Biology subjects

Yu, L.

Publications and source records attributed to Yu, L..

At least 217 records · Page 12Linked to original sources

bioRxiv2022

CDK4/6 inhibition and dsRNA sensor agonism co-operate to enhance anti-cancer effects through ER stress and immune modulation of tumour cells

Cytoplasmic pattern recognition receptors (PRRs) for double-stranded RNA (RIG-I/MDA5) are key mediators of anti-viral responses. PRR agonists, such as dsRNA oncolytic Reovirus type 3 Dearing (Rt3D), potently activate RNA sensors. We used an unbiased cytotoxicity screen to reveal synergistic drug-virotherapy combinations and found potent effects of Rt3D combined with the CDK4/6 inhibitor, palbociclib. The combination augmented oncolytic virus-induced endoplasmic reticulum (ER) stress/unfolded protein response (UPR) and the expression and activation/signaling of RNA sensors. Combined Rt3D-palbociclib treatment potently increased interferon production and signaling, and knockdown studies implicated key UPR proteins and the RNA sensor, RIG-I, as essential to the phenotype observed. Further experiments, using canonical RIG-I agonists and an ER stress inducer, thapsigargin, confirmed cross-talk between RNA sensing and ER stress pathways that augmented cancer cell death and interferon production. Combined Rt3D-palbociclib also increased innate immune activation within tumour cells and IFN-induced HLA expression. Analysis of the immunopeptidome revealed changes to HLA-captured peptides with Rt3D-palbociclib, including altered expression of peptides from cancer/testis antigens (CTA) and endogenous retroviral elements (ERVs). Our findings highlight cross-talk between UPR signaling and RNA-mediated PRR activation as a means of enhancing anti-cancer efficacy with potential pro-immunogenic consequences. This has implications for future clinical development of PRR agonists and oncolytic viruses, and broadens the therapeutic remit of CDK4/6 inhibitors to include roles as both ER stress and dsRNA PRR sensitizers.

cancer biology↗

bioRxiv2022

Large scale production of human blastoids amenable to modeling blastocyst development and maternal-fetal crosstalk

Recent advances in human blastoids generated from naive pluripotent stem cells have opened a new avenue for modelling early human development and implantation. Despite the success, however, existing protocols have several limitations, e.g., the use of custom-built microwell arrays impedes wide adoption by the research community, and mass production of human blastoids is hampered by low-output or low-efficiency methods. To address these issues, here we developed an optimized protocol based on commercially available microwell plates, which enabled efficient generation of high-fidelity human blastoids at a large scale. Leveraging on the improved protocol, we identified MAPK. PI3K/AKT and mTOR signaling pathways were activated in both blastoids and blastocyst, and discovered endometrial stromal effects in promoting trophoblast cell survival, proliferation and syncytialization during extended co-culture with blastoids. Our optimized protocol will facilitate broader use of human blastoids as an accessible, perturbable, scalable, tractable, and ethical model for human blastocysts.

developmental biology↗

bioRxiv2022

Imprinted SARS-CoV-2 humoral immunity induces converging Omicron RBD evolution

Continuous evolution of Omicron has led to a rapid and simultaneous emergence of numerous variants that display growth advantages over BA. 5. Despite their divergent evolutionary courses, mutations on their receptor-binding domain (RBD) converge on several hotspots. The driving force and destination of such convergent evolution and its impact on humoral immunity remain unclear. Here, we demonstrate that these convergent mutations can cause striking evasion of neutralizing antibody (NAb) drugs and convalescent plasma, including those from BA.5 breakthrough infection, while maintaining sufficient ACE2 binding capability. BQ.1.1.10, BA.4.6.3, XBB, and CH. 1.1 are the most antibody-evasive strain tested, even exceeding SARS-CoV-1 level. To delineate the origin of the convergent evolution, we determined the escape mutation profiles and neutralization activity of monoclonal antibodies (mAbs) isolated from BA.2 and BA.5 breakthrough-infection convalescents. Importantly, due to humoral immune imprinting, BA.2 and especially BA.5 breakthrough infection caused significant reductions in the epitope diversity of NAbs and increased proportion of non-neutralizing mAbs, which in turn concentrated humoral immune pressure and promoted convergent evolution. Moreover, we showed that the convergent RBD mutations could be accurately inferred by integrated deep mutational scanning (DMS) profiles, and the evolution trends of BA.2.75/BA.5 subvariants could be well-simulated through constructed convergent pseudovirus mutants. Together, our results suggest current herd immunity and BA.5 vaccine boosters may not provide good protection against infection. Broad-spectrum SARS-CoV-2 vaccines and NAb drugs development should be highly prioritized, and the constructed mutants could help to examine their effectiveness in advance.

immunology↗

bioRxiv2022

A Corollary Discharge Circuit in Human Speech

When we vocalize, our brain distinguishes self-generated sounds from external ones. A corollary discharge signal supports this function in animals, however, in humans its exact origin and temporal dynamics remain unknown. We report Electrocorticographic (ECoG) recordings in neurosurgical patients and a novel connectivity approach based on Granger-causality that reveals major neural communications. We find a reproducible source for corollary discharge across multiple speech production paradigms localized to ventral speech motor cortex before speech articulation. The uncovered discharge predicts the degree of auditory cortex suppression during speech, its well-documented consequence. These results reveal the human corollary discharge source and timing with far-reaching implication for speech motor-control as well as auditory hallucinations in human psychosis. Significance statementHow do organisms dissociate self-generated sounds from external ones? A fundamental brain circuit across animals addresses this question by transmitting a blueprint of the motor signal to sensory cortices, referred to as a corollary discharge. However, in humans and non-human primates auditory system, the evidence supporting this circuit has been limited to its direct consequence, auditory suppression. Furthermore, an impaired corollary discharge circuit in humans can lead to auditory hallucinations. While hypothesized to originate in the frontal cortex, direct evidence localizing the source and timing of an auditory corollary discharge is lacking in humans. Leveraging rare human neurosurgical recordings combined with connectivity techniques, we elucidate the exact source and dynamics of the corollary discharge signal in human speech. One-sentence summaryWe reveal the source and timing of a corollary discharge from speech motor cortex onto auditory cortex in human speech.

neuroscience↗

bioRxiv2022

Effects of HSP70 chaperones Ssa1 and Ssa2 on Ste5 scaffold and the mating mitogen-activated protein kinase (MAPK) Pathway in Saccharomyces cerevisiae

Ste5 is a prototype of scaffold proteins that regulate activation of mitogen-activated protein kinase (MAPK) cascades in all eukaryotes. Ste5 associates with many proteins including G{beta}{gamma} (Ste4), Ste11 MAPKKK, Ste7 MAPKK, Fus3 and Kss1 MAPKs, Bem1, Cdc24. Here we show that Ste5 also associates with heat shock protein 70 chaperone (Hsp70) Ssa1 and that Ssa1 and its ortholog Ssa2 are together important for Ste5 function and efficient mating responses. The majority of purified overexpressed Ste5 associates with Ssa1. Loss of Ssa1 and Ssa2 has deleterious effects on Ste5 abundance, integrity, and localization particularly when Ste5 is expressed at native levels. The status of Ssa1 and Ssa2 influences Ste5 electrophoresis mobility and formation of high molecular weight species thought to be phosphorylated, ubiquitinylated and aggregated and lower molecular weight fragments. A Ste5 VWA domain mutant with greater propensity to form punctate foci has reduced predicted propensity to bind Ssa1 near the mutation sites and forms more punctate foci when Ssa1 Is overexpressed, supporting a dynamic protein quality control relationship between Ste5 and Ssa1. Loss of Ssa1 and Ssa2 reduces activation of Fus3 and Kss1 MAPKs and FUS1 gene expression and impairs mating shmoo morphogenesis. Surprisingly, ssa1, ssa2, ssa3 and ssa4 single, double and triple mutants can still mate, suggesting compensatory mechanisms exist for folding. Additional analysis suggests Ssa1 is the major Hsp70 chaperone for the mating and invasive growth pathways and reveals several chaperone-network proteins required for mating morphogenesis.

molecular biology↗

bioRxiv2022

Further humoral immunity evasion of emerging SARS-CoV-2 BA.4 and BA.5 subvariants

Multiple BA.4 and BA.5 subvariants with R346 mutations on the spike glycoprotein have been identified in various countries, such as BA.4.6/BF.7 harboring R346T, BA.4.7 harboring R346S, and BA.5.9 harboring R346I. These subvariants, especially BA.4.6, exhibit substantial growth advantages compared to BA.4/BA.5. In this study, we showed that BA.4.6, BA.4.7, and BA.5.9 displayed higher humoral immunity evasion capability than BA.4/BA.5, causing 1.5 to 1.9-fold decrease in NT50 of the plasma from BA.1 and BA.2 breakthrough-infection convalescents compared to BA.4/BA.5. Importantly, plasma from BA.5 breakthrough-infection convalescents also exhibits significant neutralization activity decrease against BA.4.6, BA.4.7, and BA.5.9 than BA.4/BA.5, showing on average 2.4 to 2.6-fold decrease in NT50. For neutralizing antibody drugs, Bebtelovimab remains potent, while Evusheld is completely escaped by these subvariants. Together, our results rationalize the prevailing advantages of the R346 mutated BA.4/BA.5 subvariants and urge the close monitoring of these mutants, which could lead to the next wave of the pandemic.

immunology↗

bioRxiv2022

The response mechanism of oreorchis patens (Lindl.) Lindl. to cold stress

Cold stress, a major environmental factor, has an important impact on the production of landscape plants and crop yield, and its stress and resistance mechanisms have always been hot research issues. Oreorchis patens (Lindl.) Lindl., an important germplasm resource, has strong frost resistance and can resist low temperatures of -40 {degrees}C. However, the mechanism by which O. patens responds to cold stress remains poorly understood. Here, we examined the adaptation to the low-temperature environment of O. patens by changing the leaf tissue structure, while the synergistic effect of osmotic regulation, reactive oxygen scavenging and protein improved the resistance to cold stress. In addition, analysis of the photosynthetic system showed that cold stress changes the photosynthetic capacity of O. patens leaves to affect cold resistance. Analysis by nonparametric transcriptome sequencing revealed 2402 genes that were differentially expressed, most of which were related to resistance. Simultaneously, quantitative real-time polymerase chain reaction (qRT-PCR) analysis obtained results consistent with the transcriptome. These results indicated that O. patens could alter leaf structure and physiological and biochemical metabolic processes by initiating resistance-related molecular regulatory networks to improve the ability to resist cold stress. This study was the first to discuss the physiological, biochemical and molecular regulatory mechanisms of O. patens resistance to cold stress, which laid a foundation for revealing the biological and molecular mechanisms of overwintering of O. patens and breeding cold-resistant horticultural crops in northern China.

plant biology↗

bioRxiv2022

Neutralizing antibody evasion and receptor binding features of SARS-CoV-2 Omicron BA.2.75

Recently emerged SARS-CoV-2 Omicron subvariant, BA.2.75, displayed a local growth advantage over BA.2.38, BA.2.76 and BA.5 in India. The underlying mechanism of BA.2.75s enhanced infectivity, especially compared to BA.5, remains unclear. Here, we show that BA.2.75 exhibits substantially higher ACE2-binding affinity than BA.5. Also, BA.2.75 spike shows decreased thermostability and increased "up" RBD conformation in acidic conditions, suggesting enhanced low-pH-endosomal cell-entry pathway utilization. BA.2.75 is less humoral immune evasive than BA.4/BA.5 in BA.1/BA.2 breakthrough-infection convalescents; however, BA.2.75 shows heavier neutralization evasion in Delta breakthrough-infection convalescents. Importantly, plasma from BA.5 breakthrough infection exhibit significantly weaker neutralization against BA.2.75 than BA.5, mainly due to BA.2.75s distinct RBD and NTD-targeting antibody escaping pattern from BA.4/BA.5. Additionally, Evusheld and Bebtelovimab remain effective against BA.2.75, and Sotrovimab recovered RBD-binding affinity. Together, our results suggest BA.2.75 may prevail after the global BA.4/BA.5 wave, and its increased receptor-binding capability could allow further incorporation of immune-evasive mutations.

immunology↗

bioRxiv2022

Tetraspanin 4 mediates migrasome formation via a two-stage mechanism

Migrasomes are recently discovered signalling organelles, enriched with tetraspanin proteins (TSPAN)1. They form by local swelling of retraction fibers, the cylindrical protrusions of cell membranes that form as a result of cell migration along external substrates. Migrasomes can grow up to several micrometers in diameter2, and allow cells to release contents such as chemokines at specific locations, hence, transmitting signals to surrounding cells through the relevant chemokine receptors. Recently, evidence emerged showing that migrasomes play essential roles in fundamental cellular processes such transfer of mRNA and proteins3, organ morphogenesis4, and mitochondria quality control5. Thus, understanding the mechanism of migrasome biogenesis is of outstanding importance. Previously, it was established that the molecules crucial for migrasome formation are tetraspanin proteins and cholesterol forming macrodomains in the migrasome membrane, while the physical forces driving local swelling of the retraction fibers originate from membrane tension and bending rigidity1. Yet, it remained unknown how and in which time sequence these factors are involved in migrasome nucleation, growth, and stabilization, and what are the possible intermediate stages of migrasome biogenesis.

biophysics↗

bioRxiv2022

Lignin Composites with Sustained Oxygenation and Reactive Oxygen Species-Scavenging Improve Neovascularization and Healing of Diabetic Wounds

Although delayed wound healing is an important clinical complication in diabetic patients, few targeted treatments are available, and it remains a challenge to promote diabetic wound healing. Impaired neovascularization is one of the prime characteristics of the diabetic phenotype of delayed wound healing. Additionally, increased levels of reactive oxygen species (ROS) and chronic low-grade inflammation and hypoxia are associated with diabetes, which disrupts mechanisms of wound healing. We developed lignosulfonate composites with several wound healing properties, including sustained oxygen release through calcium peroxide nanoparticles and reactive oxygen species (ROS) and free radical scavenging by thiolated lignosulfonate nanoparticles. Sustained release of oxygen and ROS-scavenging by these composites promoted endothelial cell branching and characteristic capillary-like network formation under high glucose conditions in vitro. Gene co-expression network analysis of RNA-sequencing results from ECs cultured on lignin composites showed regulation of inflammatory pathways, alongside the regulation of angiogenic hypoxia-inducible factor-1 (HIF-1a) and vascular endothelial growth factor (VEGF) pathways. In vivo, lignosulfonate composite treatment promoted angiogenic growth factor expression and angiogenesis in full thickness skin wounds in diabetic (db/db) mice, a model of delayed wound healing. Treatment of diabetic wounds with lignosulfonate composites also promoted faster epithelial gap closure and increased granulation tissue deposition by day 7 post-wounding, with a higher presence of pro-healing type macrophages. These effects significantly improved tissue repair outcomes by day 14. Our findings demonstrate that lignosulfonate composites promote diabetic wound healing without requiring additional drugs. This highlights the potential of functionalized lignosulfonate for wound healing applications that requires balanced antioxidation and controlled oxygen release. Statement of SignificanceThe lignosulfonate composites developed in this study offer a promising solution for delayed wound healing in diabetic patients. By effectively addressing key factors contributing to the multifaceted pathophysiology of the diabetic wounds, including impaired neovascularization, increased ROS levels, and chronic inflammation and wound proteolysis, these composites demonstrate significant potential for promoting wound repair and reducing the complications associated with diabetic wounds. The unique combination of pro-angiogenic, oxygen-releasing, ECM remodeling and antioxidant properties in these lignosulfonate-based materials highlights their potential as a valuable therapeutic option, providing a novel approach to diabetic wound healing without the need for additional drugs.

bioengineering↗

bioRxiv2022

Comparative multi-omic analyses reveal gene regulatory networks controlling physiological and metabolic responses to drought in upland cotton

Drought stress substantially impacts crop physiology resulting in alteration of growth and productivity. Understanding the genetic and molecular crosstalk between stress responses and agronomically important traits such as fiber yield is particularly complicated in the allopolyploid species, upland cotton (Gossypium hirsutum), due to reduced sequence variability between A and D subgenomes. To better understand how drought stress impacts yield, the transcriptomes of 22 genetically and phenotypically diverse upland cotton accessions grown under well-watered and water-limited conditions in the Arizona low desert were sequenced. Gene co-expression analyses were performed, uncovering a group of stress response genes, in particular transcription factors GhDREB2A-A and GhHSFA6B-D, associated with improved yield under water-limited conditions in an ABA-independent manner. DNA affinity purification sequencing (DAP-seq), as well as public cistrome data from Arabidopsis, were used to identify targets of these two TFs. Among these targets were two lint-yield associated genes previously identified through genome-wide association studies (GWAS) -based approaches, GhABP-D and GhIPS1-A. Biochemical and phylogenetic approaches were used to determine that GhIPS1-A is positively regulated by GhHSFA6B-D, and that this regulatory mechanism is specific to Gossypium spp. containing the A (old-world) genome. Finally, a SNP was identified within the GhHSFA6B-D binding site in GhIPS1-A that is positively associated with yield under water limiting conditions. These data lay out a regulatory connection between abiotic stress and fiber yield in cotton that appears conserved in other systems such as Arabidopsis. This regulatory mechanism highlights how sub-genome dynamics contribute to phenotypic stress-response plasticity in cotton.

plant biology↗

bioRxiv2022

Transmembrane proteins tetraspanin 4 and CD9 sense membrane curvature

Multiple membrane shaping and remodelling processes are associated with tetraspanin proteins by yet unknown mechanisms. Tetraspanins constitute a family of proteins with four transmembrane domains present in high copy numbers in every cell type. Prominent examples are tetraspanin4 and CD9 that are required for the fundamental cellular processes of migrasome formation and fertilization, respectively. These proteins are enriched in curved membrane structures, such as cellular retraction fibers and oocyte microvilli. The factors driving this enrichment are, however, unknown. Here we revealed that tetrasapnin4 and CD9 are curvature sensors with a preference for positive membrane curvature. To this end we used a biomimetic system emulating membranes of cell retraction fibers and oocyte microvilli by membrane tubes pulled out of giant plasma membrane vesicles with controllable membrane tension and curvature. We developed a simple thermodynamic model for the partitioning of curvature sensors between flat and tubular membranes, which allowed us to estimate the individual intrinsic curvatures of the two proteins. Overall, our findings illuminate the process of migrasome formation and oocyte microvilli shaping and provide insight into the role of tetraspanin proteins in membrane remodelling processes.

biophysics↗

bioRxiv2022

IDPConformerGenerator: A Flexible Software Suite for Sampling Conformational Space of Disordered Protein States

The power of structural information for informing biological mechanism is clear for stable folded macromolecules, but similar structure-function insight is more difficult to obtain for highly dynamic systems such as intrinsically disordered proteins (IDPs) which must be described as structural ensembles. Here we present IDPConformerGenerator, a flexible, modular open source software platform for generating large and diverse ensembles of disordered protein states that builds conformers that obey geometric, steric and other physical restraints on the input sequence. IDPConformerGenerator samples backbone phi ({varphi}), psi ({psi}), and omega ({omega}) torsion angles of relevant sequence fragments from loops and secondary structure elements extracted from folded protein structures in the RCSB Protein Data Bank, and builds side chains from robust Monte Carlo algorithms using expanded rotamer libraries. IDPConformerGenerator has many user-defined options enabling variable fractional sampling of secondary structures, supports Bayesian models for assessing agreement of IDP ensembles for consistency with experimental data, and introduces a machine learning approach to transform between internal to Cartesian coordinates with reduced error. IDPConformerGenerator will facilitate the characterization of disordered proteins to ultimately provide structural insights into these states that have key biological functions.

biophysics↗

bioRxiv2022

Eudicot primary cell wall glucomannan is related in synthesis, structure and function to xyloglucan

The functional differences between plant cell wall hemicelluloses such as glucomannan, xylan and xyloglucan (XyG) remain unclear. These polysaccharides influence assembly and properties of the wall, perhaps by interacting with cellulose to affect the deposition and bundling of the fibrils. As the most abundant hemicellulose, XyG is considered important in eudicot primary cell walls (PCWs), but plants devoid of XyG show relatively mild phenotypes. We report here that a patterned {beta}-galactoglucomannan ({beta}-GGM) is widespread in PCW of eudicots and shows remarkable similarities to XyG. The sugar linkages forming the backbone and side chains of {beta}-GGM are analogous to those that make up XyG, and moreover, these linkages are formed by glycosyltransferases from the same CAZy families. Solid-state NMR indicated that {beta}-GGM shows low mobility in the cell wall, consistent with interaction with cellulose. Although Arabidopsis {beta}-GGM synthesis mutants show no obvious growth defects, genetic crosses between {beta}-GGM and XyG mutants produce exacerbated phenotypes compared to XyG mutants. These findings demonstrate a related role of these two similar but distinct classes of hemicelluloses in PCWs. This work will provide new avenues to study the roles of both {beta}-GGM and XyG in PCWs. One sentence summaryPatterned {beta}-GGM resembles xyloglucan in structure, biosynthesis and function.

plant biology↗

bioRxiv2022

BA.2.12.1, BA.4 and BA.5 escape antibodies elicited by Omicron infection

SARS-CoV-2 Omicron sublineages BA.2.12.1, BA.4 and BA.5 exhibit higher transmissibility over BA.21. The new variants receptor binding and immune evasion capability require immediate investigation. Here, coupled with Spike structural comparisons, we show that BA.2.12.1 and BA.4/BA.5 exhibit comparable ACE2-binding affinities to BA.2. Importantly, BA.2.12.1 and BA.4/BA.5 display stronger neutralization evasion than BA.2 against the plasma from 3-dose vaccination and, most strikingly, from post-vaccination BA.1 infections. To delineate the underlying antibody evasion mechanism, we determined the escaping mutation profiles2, epitope distribution3 and Omicron neutralization efficacy of 1640 RBD-directed neutralizing antibodies (NAbs), including 614 isolated from BA.1 convalescents. Interestingly, post-vaccination BA.1 infection mainly recalls wildtype-induced humoral memory. The resulting elicited antibodies could neutralize both wildtype and BA.1 and are enriched on non-ACE2-competing epitopes. However, most of these cross-reactive NAbs are heavily escaped by L452Q, L452R and F486V. BA.1 infection can also induce new clones of BA.1-specific antibodies that potently neutralize BA.1; nevertheless, these NAbs are largely escaped by BA.2/BA.4/BA.5 due to D405N and F486V, and react weakly to pre-Omicron variants, exhibiting poor neutralization breadths. As for therapeutic NAbs, Bebtelovimab4 and Cilgavimab5 can effectively neutralize BA.2.12.1 and BA.4/BA.5, while the S371F, D405N and R408S mutations would undermine most broad sarbecovirus NAbs. Together, our results indicate that Omicron may evolve mutations to evade the humoral immunity elicited by BA.1 infection, suggesting that BA.1-derived vaccine boosters may not achieve broad-spectrum protection against new Omicron variants.

immunology↗

bioRxiv2022

Repositioning linifanib as a potent anti-necroptosis agent for sepsis

BackgroundSepsis is a systemic inflammatory syndrome (SIRS) caused by acute microbial infection with high mortality rate. The role of tumour necrosis factor (TNF-)-induced necroptosis in promoting the pathophysiology of sepsis has been identified. Effective prevention of necroptosis is expected to improve the prognosis of sepsis patients. MethodsWe conducted bioinformatics prediction of candidate drugs by analyzing differentially expressed genes of sepsis patients extracted from GEO database, combining library of integrated network-based cellular signatures (LINCS) L1000 perturbation database. Biological experiments based on TNF--induced necroptosis in cellular and mouse model were performed to verify the protection of candidate drugs from SIRS. Cell viability was measured by CellTiter-Glo luminescent ATP assay. Effects of linifanib on necroptosis were investigated by western blotting, immunoprecipitation, and in vitro RIPK1 kinase assay. Survival curve analysis of SIRS mice treated by linifanib was performed. ResultsA total of 16 candidate drugs was screened out through bioinformatics analysis. Our experiments demonstrated that linifanib effectively protected cells from necroptosis and rescued the death of SIRS mice from shock induced by TNF-. In vitro, linifanib directly suppressed RIPK1 kinase activity. In vivo, linifanib effectively reduced the overexpressed level of IL-6, a good marker of severity during severe sepsis, in the lung of SIRS mice. ConclusionWe provide preclinical evidence for the potential clinical utility of linifanib in sepsis. Study of drug repositioning using bioinformatical predictions combined with experimental validations provides novel strategies for the development of sepsis drug.

bioinformatics↗

bioRxiv2022

Multifactorial remodeling of the cancer immunopeptidome by interferon gamma

IFN{gamma} alters the immunopeptidome presented on HLA class I (HLA-I), and its activity on cancer cells is known to be important for effective immunotherapy responses. We performed proteomic analyses of untreated and IFN{gamma}-treated colorectal cancer patient-derived organoids (PDOs) and combined this with transcriptomic and HLA-I immunopeptidomics data to dissect mechanisms that lead to remodeling of the immunopeptidome through IFN{gamma}. IFN{gamma}-induced changes in the abundance of source proteins, switching from the constitutive- to the immunoproteasome, and differential upregulation of different HLA alleles explained some, but not all, observed peptide abundance changes. By selecting for peptides which increased or decreased the most in abundance, but originated from proteins with limited abundance changes, we discovered that the amino acid composition of presented peptides also influences whether a peptide is up- or downregulated on HLA-I through IFN{gamma}. The presence of proline within the peptide core was most strongly associated with peptide downregulation. This was validated in an independent dataset. Proline substitution in relevant core positions did not influence the predicted HLA-I binding affinity or stability, indicating that proline effects on peptide processing may be most relevant. Understanding the multiple factors that influence the abundance of peptides presented on HLA-I in the absence or presence of IFN{gamma} is important to identify the best targets for antigen-specific cancer immunotherapies such as vaccines or T-cell receptor engineered therapeutics.

cancer biology↗

bioRxiv2022

Quantitative prediction of ensemble dynamics, shapes and contact propensities of intrinsically disordered proteins

Intrinsically disordered proteins (IDPs) are highly dynamic systems that play an important role in cell signaling processes and their misfunction often causes human disease. Proper understanding of IDP function not only requires the realistic characterization of their three-dimensional conformational ensembles at atomic-level resolution but also of the time scales of interconversion between their conformational substates. Large sets of experimental data are often used in combination with molecular modeling to restrain or bias models to improve agreement with experiment. It is shown here for the N-terminal transactivation domain of p53 (p53TAD) and Pup how the latest advancements in molecular dynamics (MD) simulations methodology produces native conformational ensembles by combining replica exchange with series of microsecond MD simulations. They closely reproduce experimental data at the global conformational ensemble level, in terms of the distribution properties of the radius of gyration tensor, and at the local level, in terms of NMR properties including 15N spin relaxation, without the need for reweighting. The IDP ensembles were analyzed by graph theory to identify dominant inter-residue contact clusters and characteristic amino-acid contact propensities. These findings indicate that modern MD force fields with residue-specific backbone potentials can produce highly realistic IDP ensembles sampling a hierarchy of nano- and picosecond time scales providing new insights into their biological function. AUTHOR SUMMARYAccurate prediction of the conformational ensemble dynamics sans bias is shown for intrinsically disordered proteins including the transactivation domain of p53.

biophysics↗