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bioRxiv2018

Immunization by replication-competent controlled herpesvirus vectors

Replication-competent controlled virus vectors were derived from virulent HSV-1 wildtype strain 17syn+ by placing one or two replication-essential genes under the stringent control of a gene switch that is co-activated by heat and an antiprogestin. Upon activation of the gene switch, the vectors replicate in infected cells with an efficacy that approaches that of the wildtype virus from which they were derived. Essentially no replication occurs in the absence of activation. When administered to mice, localized application of a transient heat treatment in the presence of systemic antiprogestin results in efficient but limited virus replication at the site of administration. The immunogenicity of these viral vectors was tested in a mouse footpad lethal challenge model. Unactivated viral vectors - which may be regarded as equivalents of inactivated vaccines - induced detectable protection against lethality caused by wildtype virus challenge. Single activation of the viral vectors at the site of administration (rear footpads) greatly enhanced protective immune responses, and second immunization resulted in complete protection. Once activated vectors also induced far better neutralizing antibody and HSV-1-specific T cells responses than unactivated vectors. To find out whether the immunogenicity of a heterologous antigen was also enhanced in the context of efficient transient vector replication, a virus vector constitutively expressing an equine influenza virus hemagglutinin was constructed. Immunization of mice with this recombinant induced detectable antibody-mediated neutralization of equine influenza virus as well as a hemagglutinin-specific T cell response. Single activation of viral replication resulted in a several-fold enhancement of this immune response.\n\nIMPORTANCEWe hypothesized that vigorous replication of a pathogen may be critical for eliciting the most potent and balanced immune response against it. Hence, attenuation/inactivation (as in conventional vaccines) should be avoided. Instead, necessary safety should be provided by placing replication of the pathogen under stringent control and of activating time-limited replication of the pathogen strictly in an administration region in which pathology cannot develop. Immunization will then occur in the context of highly efficient pathogen replication and uncompromised safety. We found that localized activation in mice of efficient but limited replication of a replication-competent controlled herpesvirus vector resulted in a greatly enhanced immune response to the virus or an expressed heterologous antigen. This finding supports the above hypothesis as well as suggests that the vectors may be promising novel agents worth exploring for the prevention/mitigation of infectious diseases for which efficient vaccination is lacking, in particular in immunocompromised patients.

immunology↗

bioRxiv2018

Brain scans from 21297 individuals reveal the genetic architecture of hippocampal subfield volumes

The hippocampus is a heterogeneous structure, comprising histologically distinguishable subfields. These subfields are differentially involved in memory consolidation, spatial navigation and pattern separation, complex functions often impaired in individuals with brain disorders characterized by reduced hippocampal volume, including Alzheimers disease (AD) and schizophrenia. Given the structural and functional heterogeneity of the hippocampal formation, we sought to characterize the subfields genetic architecture. T1-weighted brain scans (n=21297, 16 cohorts) were processed with the hippocampal subfields algorithm in FreeSurfer v6.0. We ran a genome-wide association analysis on each subfield, covarying for total hippocampal volume. We further calculated the single nucleotide polymorphism (SNP)-based heritability of twelve subfields, as well as their genetic correlation with each other, with other structural brain features, and with AD and schizophrenia. All outcome measures were corrected for age, sex, and intracranial volume. We found 15 unique genome-wide significant loci across six subfields, of which eight had not been previously linked to the hippocampus. Top SNPs were mapped to genes associated with neuronal differentiation, locomotor behaviour, schizophrenia and AD. The volumes of all the subfields were estimated to be heritable (h2 from .14 to .27, all p< 1x10-16) and clustered together based on their genetic correlations compared to other structural brain features. There was also evidence of genetic overlap of subicular subfield volumes with schizophrenia. We conclude that hippocampal subfields have partly distinct genetic determinants associated with specific biological processes and traits. Taking into account this specificity may increase our understanding of hippocampal neurobiology and associated pathologies.

genetics↗

bioRxiv2018

Mice harboring a SCA28 patient mutation in AFG3L2 develop late-onset ataxia associated with enhanced mitochondrial proteotoxicity.

Spinocerebellar ataxia 28 is an autosomal dominant neurodegenerative disorder caused by missense mutations affecting the proteolytic domain of AFG3L2, a major component of the mitochondrial m-AAA protease. However, little is known of the underlying pathogenetic mechanisms or how to treat patients with SCA28. Currently available Afg3l2 mutant mice harbour deletions that lead to severe, early-onset neurological phenotypes that do not faithfully reproduce the late-onset and slowly progressing SCA28 phenotype. Here we describe production and detailed analysis of a new knock-in murine model harbouring an Afg3l2 allele carrying the p.Met665Arg patient-derived mutation. Heterozygous mutant mice developed normally but signs of ataxia were detectable by beam test at 18 months. Cerebellar pathology was negative; electrophysiological analysis showed increased spontaneous firing in Purkinje cells from heterozygous mutants with respect to wild-type controls, although not statistically significant. As homozygous mutants died perinatally with evidence of cardiac atrophy, for each genotype we generated mouse embryonic fibroblasts (MEFs) to investigate mitochondrial function. MEFs from mutant mice showed altered mitochondrial bioenergetics, with decreased basal oxygen consumption rate, ATP synthesis and mitochondrial membrane potential. Mitochondrial network formation and morphology was also altered, in line with greatly reduced expression of Opa1 fusogenic protein L-isoforms. The mitochondrial alterations observed in MEFs were also detected in cerebella of 18-month-old heterozygous mutants, suggesting they may be a hallmark of disease. Pharmacological inhibition of de novo mitochondrial protein translation with chloramphenicol caused reversal of mitochondrial morphology in homozygous mutant MEFs, supporting the relevance of mitochondrial proteotoxicity for SCA28 pathogenesis and therapy development.

genetics↗

bioRxiv2018

Precise detection of S phase onset reveals decoupled G1/S transition events

The eukaryotic cell division cycle is the process by which cells duplicate their genomes and proliferate. Transitions between sequential cell cycle phases are tightly orchestrated to ensure precise and efficient cell cycle progression. Interrogating molecular events at these transitions is important for understanding normal and pathological cell proliferation and mechanisms that ensure genome stability. A popular fluorescent reporter system known as \"FUCCI\" has been widely adopted for identifying cell cycle phases. Using time-lapse fluorescence microscopy, we quantitatively analyzed the dynamics of the FUCCI reporters relative to the transitions into and out of S phase. Although the original reporters reflect the E3 ubiquitin ligase activities for which they were designed, SCFSkp2 and APCCdh1, their dynamics are significantly and variably offset from actual S phase boundaries. To precisely mark these transitions, we generated and thoroughly validated a new reporter containing a PCNA-interacting protein degron whose oscillations are directly coupled to the process of DNA replication itself. We combined this reporter with the geminin-based APCCdh1 reporter to create \"PIP-FUCCI.\" PIP degron reporter dynamics closely correlate with S phase transitions irrespective of reporter expression levels. Using PIP-FUCCI, we made the unexpected observation that the apparent timing of APCCdh1 inactivation frequently varies relative to the onset of S phase. We demonstrate that APCCdh1 inactivation is not a strict pre-requisite for S phase entry, though delayed APCCdh1 inactivation correlates with longer S phase. Our results illustrate the benefits of precise delineation of cell cycle phase boundaries for uncovering the sequences of molecular events at critical cell cycle transitions.

cell biology↗

bioRxiv2018

Pervasive chromatin remodeling at X-inactivation escape genes in schizophrenic males

Reanalyzing a large methylome dataset of 225 schizophrenic and 450 control samples derived from the prefrontal cortex revealed that 6 male patients have predominantly hypomethylated probes mostly on chromosome X, affecting the same genes in all six. Network analysis of the differentially methylated genes revealed a dense network of transcription factors, histone and chromatin remodeling proteins, with 15 of the X-located genes expressed at the synapse, including NLGN4X, SYN1 and MECP2. Mapping a recent experimental dataset of G-quadruplexes (G4s) onto the differentially methylated probes revealed that the probes in the group of six overlapping with G4s on chromosome X are significantly more hypomethylated than non-overlapping and non-X probes whereas in the rest of the patients G4-overlapping probes are more methylated than non-overlapping ones, revealing a distinct pathology, involving chromatin remodeling for the six patients. Unexpectedly, the hypomethylated genes in them significantly overlapped with gene locations where X-inactivation escapism was observed in women.

neuroscience↗

bioRxiv2018

MicroRNA-205 affects mouse granulosa cell apoptosis and estradiol synthesis by targeting CREB1

MicroRNAs-205 (miR-205), were reportedly to be involved in various physiological and pathological processes, but its biological function in follicular atresia remain unknown. In this study, we investigated the expression of miR-205 in mouse granulosa cells (mGCs), and explored its functions in primary mGCs using a serial of in vitro experiments. The result of qRT-PCR demonstrated that miR-205 expression was significantly increased in early atretic follicles (EAF), and progressively atretic follicles (PAF) compared to healthy follicles (HF). Our results also revealed that overexpression of miR-205 in mGCs significantly promoted apoptosis, caspas-3/9 activities, and inhibited estrogen E2 release, and cytochrome P450 family 19 subfamily A polypeptide 1 (CYP19A1, a key gene in E2 production) expression. Bioinformatics and luciferase reporter assays revealed that the gene of cyclic AMP response element (CRE)-binding protein 1 (CREB1) was a potential target of miR-205. qRT-PCR and western blot assays revealed that overexpression of miR-205 inhibited the expression of CREB1 in mGCs. Importantly, CREB1 upregulation partially rescued the effects of miR-205 on apoptosis, caspase-3/9 activities, E2 production and CYP19A1 expression in mGCs. Our results indicate that miR-205 may play an important role in ovarian follicular development and provide new insights into follicular atresia.

evolutionary biology↗

bioRxiv2018

The mechanical impact of col11a2 loss on joints; col11a2 mutant zebrafish show changes to joint development and function which lead to early onset osteoarthritis

Abstract (max 200 words)Collagen is the major structural component of cartilage and mutations in the genes encoding Type XI collagen are associated with severe skeletal dysplasias (Fibrochondrogenesis and Stickler syndrome) and early onset osteoarthritis. The impact of the lack of Type XI collagen on cell behaviour and mechanical performance during skeleton development is unknown. We studied a zebrafish mutant for col11a2 and evaluated cartilage, bone development and mechanical properties to address this. We show that in col11a2 mutants Type II collagen is made but is prematurely degraded in maturing cartilage and ectopically expressed in the joint. These changes are correlated with increased stiffness of both bone and cartilage; quantified using Atomic Force Microscopy. In the mutants, the skeletal rudiment terminal region in the jaw joint are broader and the interzone smaller. These differences in shape and material properties impact on joint function and mechanical performance, which we modelled using Finite Element Analyses. Finally, we show that col11a2 heterozygous carriers reach adulthood but show signs of severe early onset osteoarthritis. Taken together our data demonstrate a key role for Type XI collagen in maintaining the properties of cartilage matrix; which when lost leads to alterations to cell behaviour that give rise to joint pathologies.

developmental biology↗

bioRxiv2018

PRESTO, a new tool for integrating large-scale -omics data and discovering disease-specific signatures

BackgroundCohesive visualization and interpretation of hyperdimensional, large-scale -omics data is an ongoing challenge, particularly for biologists and clinicians involved in current highly complex sequencing studies. Multivariate studies are often better suited towards non-linear network analysis than differential expression testing. Here, we present PRESTO, a PREdictive Stochastic neighbor embedding Tool for Omics, which allows unsupervised dimensionality reduction of multivariate data matrices with thousands of subjects or conditions. PRESTO is intuitively integrated into an interactive user interface that helps to visualize the multidimensional patterns in genome-wide transcriptomic data from basic science and clinical studies.\n\nResultsPRESTO was tested with multiple input omics platforms, including microarray and proteomics from both mouse and human clinical datasets. PRESTO can analyze up to tens of thousands of genes and shows no increase in processing time with a large number of samples or patients. In complex datasets, such as those with multiple time points, several patient groups, or diverse mouse strains, PRESTO outperformed conventional methods. Core co-expressed gene networks were intuitively grouped in clusters, or gates, after dimensionality reduction and remained consistent across users. Networks were identified and assigned to physiological and pathological functions that cannot be gleaned from conventional bioinformatics analyses. PRESTO detected gene networks from the natural variations among mouse macrophages and human blood leukocytes. We applied PRESTO to clinical transcriptomic and proteomic data from large patient cohorts and detected disease-defining signatures in antibody-mediated kidney transplant rejection, renal cell carcinoma, and relapsing acute myeloid leukemia (AML). In AML, PRESTO confirmed a previously described gene signature and found a new signature of 10 genes that is highly predictive of patient outcome.\n\nConclusionsPRESTO offers an important integration of powerful bioinformatics tools with an interactive user interface that increases data analysis accessibility beyond bioinformaticians and coders. Here, we show that PRESTO out performs conventional methods, such as DE analysis, in multi-dimensional datasets and can identify biologically relevant co-expression gene networks. In paired samples or time points, co-expression networks could be compared for insight into longitudinal regulatory mechanisms. Additionally, PRESTO identified disease-specific signatures in clinical datasets with highly significant diagnostic and prognostic potential.

bioinformatics↗

bioRxiv2018

integRATE: a desirability-based data integration framework for the prioritization of candidate genes across heterogeneous omics and its application to preterm birth

BackgroundThe integration of high-quality, genome-wide analyses offers a robust approach to elucidating genetic factors involved in complex human diseases. Even though several methods exist to integrate heterogeneous omics data, most biologists still manually select candidate genes by examining the intersection of lists of candidates stemming from analyses of different types of omics data that have been generated by imposing hard (strict) thresholds on quantitative variables, such as P-values and fold changes, increasing the chance of missing potentially important candidates.\n\nMethodsTo better facilitate the unbiased integration of heterogeneous omics data collected from diverse platforms and samples, we propose a desirability function framework for identifying candidate genes with strong evidence across data types as targets for follow-up functional analysis. Our approach is targeted towards disease systems with sparse, heterogeneous omics data, so we tested it on one such pathology: spontaneous preterm birth (sPTB).\n\nResultsWe developed the software integRATE, which uses desirability functions to rank genes both within and across studies, identifying well-supported candidate genes according to the cumulative weight of biological evidence rather than based on imposition of hard thresholds of key variables. Integrating 10 sPTB omics studies identified both genes in pathways previously suspected to be involved in sPTB as well as novel genes never before linked to this syndrome. integRATE is available as an R package on GitHub (https://github.com/haleyeidem/integRATE).\n\nConclusionsDesirability-based data integration is a solution most applicable in biological research areas where omics data is especially heterogeneous and sparse, allowing for the prioritization of candidate genes that can be used to inform more targeted downstream functional analyses.

genomics↗

bioRxiv2018

Machine learning based classification of cells into chronological stages using single-cell transcriptomics.

Age-associated deterioration of cellular physiology leads to pathological conditions. The ability to detect premature aging could provide a window for preventive therapies against age-related diseases. However, the techniques for determining cellular age are limited, as they rely on a limited set of histological markers and lack predictive power. Here, we implement GERAS (GEnetic Reference for Age of Single-cell), a machine learning based framework capable of assigning individual cells to chronological stages based on their trans criptomes. GERAS displays greater than 90% accuracy in classifying the chronological stage of zebrafish and human pancreatic cells. The framework demonstrates robustness against biological and technical noise, as evaluated by its performance on independent samplings of single-cells. Additionally, GERAS determines the impact of differences in calorie intake and BMI on the aging of zebrafish and human pancreatic cells, respectively. We further harness the predictive power of GERAS to identify genome-wide molecular factors that correlate with aging. We show that one of these factors, junb, is necessary to maintain the proliferative state of juvenile beta-cells. Our results showcase the applicability of a machine learning framework to classify the chronological stage of heterogeneous cell populations, while enabling to detect pro-aging factors and candidate genes associated with aging.

bioinformatics↗

bioRxiv2018

CD161 mediates prenatal immune suppression of IFNγ-producing PLZF+ T cells

While the fetal immune system defaults to a program of tolerance, there is concurrent need for protective immunity to meet the antigenic challenges after birth. Activation of fetal T cells is associated with fetal inflammation and the termination of pregnancy, yet which fetal T cells contribute to this process is poorly understood. Here we show a transcriptionally distinct population of pro-inflammatory T cells that predominates in the human fetal intestine. Activation of PLZF+ T cells results in rapid production of Th1 cytokines and is inhibited upon ligation of surface CD161. This mechanism of fetal immune suppression may inform how immune dysregulation could result in fetal and neonatal inflammatory pathologies such as preterm birth. Our data support that human development of protective adaptive immunity originates in utero within the specialized microenvironment of the fetal intestine.

immunology↗

bioRxiv2018

In silico analysis of piRNAs in retina reveals potential targets in intracellular transport and retinal degeneration

Long considered to be active only in germline, PIWI/piRNA pathway is now known to play significant role in somatic cells, especially neurons. Nonetheless, so far there is no evidence for the presence of piRNAs in the neurosensory retina. In this study, we have uncovered 102 piRNAs in human retina and retinal pigment epithelium (RPE) by analysing RNA-seq data. The identified piRNAs were enriched with three motifs predicted to be involved in rRNA processing and sensory perception. Further, expression of piRNAs in donor eyes were assessed by qRT-PCR. Loss of piRNAs in HIWI2 knockdown ARPE19 cells downregulated targets implicated in intracellular transport (SNAREs and Rabs), circadian clock (TIMELESS) and retinal degeneration (LRPAP1 and RPGRIP1). Moreover, piRNAs were dysregulated under oxidative stress indicating their potential role in retinal pathology. Intriguingly, computational analysis revealed complete and partial seed sequence similarity between piR-62011 and sensory organ specific miR-183/96/182 cluster. Furthermore, the expression of retina enriched piR-62011 positively correlated with miR-182 in HIWI2 silenced Y79 cells. Thus, our data provides an evidence for the expression of piRNAs in human retina and RPE. Collectively, our work demonstrates that piRNAs dynamically regulate distinct molecular events in the maintenance of retinal homeostasis.

cell biology↗

bioRxiv2018

Elevated O-GlcNAcylation Enhances Pro-Inflammatory Th17 Function by Altering the Lipid Microenvironment

Chronic, low-grade inflammation increases the risk of atherosclerosis, cancer, and autoimmunity in diseases like obesity and diabetes. Here, we show that increased levels of the nutrient-responsive, post-translational protein modification, O-GlcNAc (O-linked {beta}-N-acetylglucosamine) are present in naive CD4+ T cells from a diet-induced obesity murine model, and elevation in O-GlcNAc leads to increased pro-inflammatory IL-17A production. Importantly, CD4+ T helper 17 (Th17) cells, which secrete IL-17A, are increased in obesity and contribute to the inflammatory milieu. We found increased binding of the Th17 master transcription factor, ROR{gamma}t, at the IL-17 locus and significant alterations in the lipid microenvironment, leading to increased ligands capable of increasing ROR{gamma}t transcriptional activity. Importantly, the rate-limiting enzyme of fatty acid biosynthesis, acetyl CoA carboxylase 1 (ACC1), is necessary for production of these ROR{gamma}t activating ligands and is O-GlcNAcylated. Thus, we have identified O-GlcNAc as a critical link between excess nutrients and pathological inflammation.

immunology↗

bioRxiv2018

Inter-institutional variation in predictive value of the ThyroSeq v2 genomic classifier for cytologically indeterminate thyroid nodules

BackgroundThe ThyroSeq v2 next-generation sequencing assay (ThyroSeq) estimates the probability of malignancy in indeterminate thyroid nodules (ITN). Its diagnostic accuracy in different practice settings and patient populations is not well understood.\n\nMethodsWe analyzed 273 Bethesda III/IV ITN evaluated with ThyroSeq at 4 institutions: 2 comprehensive cancer centers (n=98 and 102), a multicenter healthcare system (n=60), and an academic medical center (n=13). The positive (PPV) and negative predictive values (NPV) of ThyroSeq, and distribution of final pathology were analyzed and compared to values predicted by Bayes Theorem.\n\nResultsAcross 4 institutions, the PPV was 35% (22-43%), and NPV was 93% (88-100%). Predictive values correlated closely with Bayes Theorem estimates (r2=.84), although PPVs were lower than expected. RAS mutations were the most frequent molecular alteration. Among 84 RAS-mutated nodules, malignancy risk was variable (25%, range 10-37%), and distribution of benign diagnoses differed across institutions (adenoma/hyperplasia 12-85%, NIFTP 5-46%).\n\nConclusionsIn a multi-institutional analysis, ThyroSeq PPVs were variable and lower than expected. This is attributable to differences in the prevalence of malignancy, and variability in pathologist interpretations of non-invasive tumors. It is important that clinicians understand ThyroSeq performance in their practice setting when evaluating these results.

cancer biology↗

bioRxiv2018

Glial cells in the heart? Replicating the diversity of the myocardium with low-cost 3D models

Excitation-contraction (EC) coupling in the heart has, until recently, been solely accredited to cardiomyocytes. The inherent complexities of the heart make it difficult to examine nonmuscle contributions to contraction in vivo, and conventional in vitro models fail to capture multiple features and cellular heterogeneity of the myocardium. Here, we report on the development of a 3D cardiac Tissue towards recapitulating the architecture and composition of native myocardium in vitro. Cells are encapsulated within micropatterned gelatin-based hydrogels formed via visible light photocrosslinking. This system enables spatial control of cardiac microarchitecture, perturbation of the cellular composition, and functional measures of EC coupling via video microscopy and a custom algorithm to quantify beat frequency and degree of coordination. To demonstrate the robustness of these tools and evaluate the impact of altered cell population densities on cardiac Tissues, contractility and cell morphology were assessed with the inclusion of exogenous non-myelinating Schwann cells (SCs). Results demonstrate that the addition of exogenous SCs alter cardiomyocyte EC, profoundly inhibiting the response to electrical pacing. Computational modeling of connexin-mediated coupling suggests that SCs impact cardiomyocyte resting potential and rectification following depolarization. Cardiac Tissues hold potential for examining the role of cellular heterogeneity in heart health, pathologies, and cellular therapies.

bioengineering↗

bioRxiv2018

Myelinated inhibitory axons in human neocortex

Numerous myelinated axons traverse the human neocortex. In a previous paper (Micheva et al., 2016) we showed that in mouse many of these axons belong to local inhibitory neurons, the parvalbumin-positive basket cells. Here, using samples of neurosurgically-excised cortex, we confirm the presence of myelinated inhibitory axons in all layers of human neocortex. As in mouse, these axons have distinctive features, including high neurofilament content, short nodes of Ranvier, and high content of myelin basic protein in their myelin sheath. We further show that, consistent with the known high-energy demands of parvalbumin interneurons, the inhibitory myelinated axons have more mitochondria, as well more 2,3-cyclic nucleotide 3-phosphodiesterase (a protein enriched in the myelin cytoplasmic channels thought to provide access for trophic support from oligodendrocytes). The distinctive features of myelinated inhibitory axons in human cortical grey matter may have important implications for neurological disorders that involve pathologies of myelinated axons.

neuroscience↗

bioRxiv2018

Enrichment of rare protein truncating variants in amyotrophic lateral sclerosis patients

To discover novel genetic risk factors underlying amyotrophic lateral sclerosis (ALS), we aggregated exomes from 3,864 cases and 7,839 ancestry matched controls. We observed a significant excess of ultra-rare and rare protein-truncating variants (PTV) among ALS cases, which was primarily concentrated in constrained genes; however, a significant enrichment in PTVs does persist in the remaining exome. Through gene level analyses, known ALS genes, SOD1, NEK1, and FUS, were the most strongly associated with disease status. We also observed suggestive statistical evidence for multiple novel genes including DNAJC7, which is a highly constrained gene and a member of the heat shock protein family (HSP40). HSP40 proteins, along with HSP70 proteins, facilitate protein homeostasis, such as folding of newly synthesized polypeptides, and clearance of degraded proteins. When these processes are not regulated, misfolding and accumulation of degraded proteins can occur leading to aberrant protein aggregation, one of the pathological hallmarks of neurodegeneration.

genomics↗

bioRxiv2018

Rationally designed mimotope library for profiling of the human IgM repertoire

Specific antibody reactivities are routinely used as biomarkers but the use of antibody repertoire profiles is still awaiting recognition. Here we suggest to expedite the adoption of this class of system level biomarkers by rationally designing a peptide array as an efficient probe for an appropriately chosen repertoire compartment. Most IgM antibodies are characterized by few somatic mutations, polyspecificity and physiological autoreactivity with housekeeping function. Previously, probing this repertoire with a set of immunodominant self-proteins provided only coarse information on repertoire profiles. In contrast, here we describe the rational selection of a peptide mimotope set, appropriately sized as a potential diagnostic, that also represents optimally the diversity of the human public IgM reactivities. A 7-mer random peptide phage display library was panned on pooled human IgM. Next generation sequencing of the selected phage yielded a non-exhaustive set of 224087 mimotopes which clustered in 790 sequence clusters. A set of 594 mimotopes, representative of the most significant clusters, was used to demonstrate that this approach samples symmetrically the space of IgM reactivities. When probed with diverse patients sera in an oriented peptide array, this set produced a higher and more dynamic signal as compared to 1) random peptides, 2) random peptides purged of mimotope-like sequences and 3) mimotopes from a small subset of clusters. In this respect, the representative library is an optimized probe of the human IgM diversity. Proof of principle predictors for randomly selected diagnoses based on the optimized library demonstrated that it contains more than 1070 different profiles with the capacity to correlate with diverse pathologies. Thus, an optimized small library of IgM mimotopes is found to address very efficiently the dynamic diversity of the human IgM repertoire providing informationally dense and structurally interpretable IgM reactivity profiles. Author SummaryThe presence in the blood of antibodies specific for a particular infectious agent is used routinely as a diagnostic tool. The overall profile of available antibody reactivities (or their repertoire) in an individual has been studied much less. As an omics approach to immunity it can be a rich source of information about the system beyond just the individual history of antigenic exposure. Using a subset of antibodies - IgM, which are involved also in housekeeping functions like removing dead cells, and bacteriophage based techniques for selection of specific peptides, we managed to define a non-exhaustive set of 224087 peptides recognized by IgM antibodies present in most individuals. They were found to group naturally in 790 structural groups. Limiting these to the most outstanding 594 groups, we used one representative from each group to assemble a reasonably small set of peptides that extracts the maximum information from the antibody repertoire at a minimum cost per test. We demonstrate, that this representative peptide library is a better probe of the human IgM diversity than comparably sized libraries constructed on other principles. The optimized library contains more than 1070 different potentially profiles useful for the diagnosis, prognosis or monitoring of inflammatory and infectious conditions, tumors, neurodegenerative diseases, etc.

immunology↗