Biology subjects

Lin, L.

Publications and source records attributed to Lin, L..

At least 235 records · Page 13Linked to original sources

bioRxiv2019

A bacterial endosymbiont enables fungal immune evasion during fatal mucormycete infection

Opportunistic infections by environmental fungi are a growing clinical problem, driven by an increasing population of people with immunocompromising conditions. Spores of the Mucorales order are ubiquitious in the environment but can also cause acute invasive infections in humans through germination and evasion of the mammalian host immune system. How they achieve this, and the evolutionary drivers underlying the acquisition of virulence mechanisms, are poorly understood. Here we show that a clinical isolate of Rhizopus microsporus contains a Ralstonia pickettii bacterial endosymbiont required for virulence in both zebrafish and mice, and that this endosymbiosis enables secretion of factors that potently suppress growth of the soil amoeba Dictyostelium discoideum, as well as their ability to engulf and kill other microbes. As amoebae are natural environmental predators of both bacteria and fungi, we propose this tri-kingdom interaction contributes to establishing the endosymbiosis, and acquisition of anti-phagocyte activity. Importantly, we show this activity also protects fungal spores from phagocytosis and clearance by human macrophages, and endosymbiont removal renders the fungal spores avirulent in vivo. Together, these findings describe a novel role for a bacterial endosymbiont in Rhizopus microsporus pathogenesis in animals, and suggest a mechanism of virulence acquisition through environmental interactions with amoebae. In briefHow environmental fungi evolved the mechanisms that enable them to cause opportunistic infections in humans is unclear. Here, we identify a novel tri-kingdom interaction, whereby a bacterial endosymbiont, living within a clinical isolate of the ubiquitous environmental fungus Rhizopus microsporus, causes the generation of a secreted activity that blocks the growth and predatory activity of amoebae. We suggest this provides a new evolutionary driver for the establishment of bacterial/fungal endosymbiosis and demonstrate this is critical for fungal pathogenicity in vivo.

microbiology↗

bioRxiv2019

On triangular inequalities of correlation-based distances for gene expression profiles

Various distance functions for evaluating the differences be- tween gene expression profiles have been proposed in the past. Such a function would output a low value if the profiles are strongly correlated--either negatively or positively--and vice versa. One popular distance function is the absolute correlation distance, da = 1 - |{rho}|, where{rho} is some similarity measures, such as Pearson or Spearman correlation. How- ever, absolute correlation distance fails to fulfill the triangular inequality, which would have guaranteed better performance at vector quantization, allowed fast data localization, as well as sped up data clustering. In this work, we propose [Formula] as an alternative. We prove that dr satisfies the triangular equality when{rho} represents Pearson correlation, Spearman correlation, or Cosine similarity. We empirically compared dr with da in gene clustering and sample clustering experiment, using real biological data. The two distances performed similarly in both gene cluster and sample cluster in hierarchical cluster and PAM cluster. However, dr demonstrated more robust clustering. According to bootstrap experiment, the number of times where dr generated more robust sample pair partition is significantly (p-value < 0.05) larger. This advantage in robustness is also supported by the class \"dissolved\" event.

bioinformatics↗

bioRxiv2019

Retrovirus insertion site analysis of LGL leukemia patient genomes

BackgroundLarge granular lymphocyte (LGL) leukemia is an uncommon cancer characterized by a sustained clonal proliferation of LGL cells. Antibodies reactive to retroviruses have been documented in the serum of patients with LGL leukemia. Culture or molecular approaches have to date not been successful in identifying a retrovirus. MethodsBecause a retrovirus must integrate into the genome of an infected cell, we focused our efforts on detecting a novel retrovirus integration site in the clonally expanded LGL cells. We present a new computational tool that uses long-insert mate pair sequence data to search the genome of LGL leukemia cells for retrovirus integration sites. We also utilize recently published methods to interrogate the status of polymorphic human endogenous retrovirus type K (HERV-K) provirus in patient genomes. ResultsWhile our analysis did not reveal any new retrovirus insertions in LGL genomes from LGL leukemia patients, we did identify four HERV-K provirus integration sites that are polymorphic in the human population and absent from the human reference genome, hg19. To determine if the prevalence of these or other polymorphic proviral HERV-Ks differed between LGL leukemia patients and the general population, we applied a recently developed approach that reports all sites in the human genome occupied by a proviral HERV-K. Using the 1000 genomes project (KGP) data as a reference database for HERV-K proviral prevalence at each polymorphic site, we show that there are significant differences in the number of polymorphic HERV-Ks in the genomes of LGL leukemia patients of European origin compared to individuals with European ancestry in the KGP data. ConclusionsOur study confirms that the integration of a new infectious or endogenous retrovirus does not cause the clonal expansion of LGL cells in LGL leukemia, although we do not rule out that these cells could be responding to retroviral antigens produced in other cell types. However, it is of interest that the burden of polymorphic proviral HERV-K is elevated in LGL leukemia patient genomes. Our research emphasizes the merits of comprehensive genomic assessment of HERV-K in cancer samples and suggests that further analyses to determine contributions of HERV-K to LGL leukemia are warranted.

genomics↗

bioRxiv2019

A plant protein NbP3IP induces autophagy and mediates the autophagic degradation of RSV p3 to inhibit viral infection

In plants, autophagy is involved in responses to viral infection. However, understanding of new host factors mediating autophagic clearance of plant viruses is very limited. We here identified a new host factor NbP3IP participating in autophagy-mediated plant defense against viral infection. NbP3IP interacted with p3, a RNA silencing suppressor encoded by Rice stripe virus (RSV), a negative-strand RNA virus, and mediated its autophagic degradation. NbP3IP could also interact with NbATG8f, which was required for NbP3IP-miediated p3 degradation. Overexpression of NbP3IP induced autophagy and down-regulation of NbP3IP reduced autophagy. Both overexpression of NbP3IP and silencing of GAPC, which also induces autophagy, inhibited RSV infection. In contrast, silencing of ATG7 promoted RSV infection. Thus, through identification of a new potential selective autophagy receptor P3IP, we revealed a new mechanism of autophagy-mediated plant defense against plant viruses and provided the first evidence that plant autophagy can also play an antiviral role against negative-strand RNA viruses.

plant biology↗

bioRxiv2019

Cross-talk of cellulose and mannan perception pathways leads to inhibition of cellulase production in several filamentous fungi

It is essential for microbes to acquire information about their environment. Fungi use soluble degradation products of plant cell wall components to understand the substrate composition they grow on. Individual signaling pathways have been well described. However, the interconnections between pathways remain poorly understood. In the present work, we provide evidence of \"confusion\" due to cross-talk between the perception pathways for cellulose and the hemicellulose mannan in several filamentous fungi, leading to the inhibition of cellulase expression. We used the functional genomics tools available for Neurospora crassa to investigate this signaling overlap at the molecular level. Cross-talk and competitive inhibition could be identified both during uptake by cellodextrin transporters and intracellularly. Importantly, the overlap is independent of CRE-1-mediated catabolite repression. These results provide novel insights into the regulatory networks of lignocellulolytic fungi and will contribute to the rational optimization of fungal enzyme production for efficient plant biomass depolymerization and utilization.

genetics↗

bioRxiv2019

A brainstem-central amygdala circuit underlies defensive responses to learned threats

Norepinephrine (NE) plays a central role in the acquisition of aversive learning via actions in the lateral nucleus of the amygdala (LA)1,2. However, the function of NE in expression of aversively-conditioned responses has not been established. Given the role of the central nucleus of the amygdala (CeA) in the expression of such behaviors3, and the presence of NE projections in this brain nucleus, we assessed the effects of NE activity in the CeA on behavioral expression using receptor-specific pharmacology and cell-and projection-specific chemogenetic manipulations. We found that inhibition and activation of locus coeruleus (LC) neurons decreases and increases freezing to aversively conditioned cues, respectively. We then show that locally inhibiting or activating LC terminals in CeA is sufficient to achieve this bidirectional modulation of defensive reactions. These findings support the hypothesis that LC projections to CeA are required for the expression of defensive responses elicited by conditioned threats.

neuroscience↗

bioRxiv2018

A Computational Framework To Assess Genome-Wide Distribution Of Polymorphic Human Endogenous Retrovirus-K In Human Populations

Human Endogenous Retrovirus type K (HERV-K) is the only HERV known to be insertionally polymorphic. It is possible that HERV-Ks contribute to human disease because people differ in both number and genomic location of these retroviruses. Indeed viral transcripts, proteins, and antibody against HERV-K are detected in cancers, auto-immune, and neurodegenerative diseases. However, attempts to link a polymorphic HERV-K with any disease have been frustrated in part because population frequency of HERV-K provirus at each site is lacking and it is challenging to identify closely related elements such as HERV-K from short read sequence data. We present an integrated and computationally robust approach that uses whole genome short read data to determine the occupation status at all sites reported to contain a HERV-K provirus. Our method estimates the proportion of fixed length genomic sequence (k-mers) from whole genome sequence data matching a reference set of k-mers unique to each HERV-K loci and applies mixture model-based clustering to account for low depth sequence data. Our analysis of 1000 Genomes Project Data (KGP) reveals numerous differences among the five KGP super-populations in the frequency of individual and co-occurring HERV-K proviruses; we provide a visualization tool to easily depict the prevalence of any combination of HERV-K among KGP populations. Further, the genome burden of polymorphic HERV-K is variable in humans, with East Asian (EAS) individuals having the fewest integration sites. Our study identifies population-specific sequence variation for several HERV-K proviruses. We expect these resources will advance research on HERV-K contributions to human diseases.\n\nAuthor summaryHuman Endogenous Retrovirus type K (HERV-K) is the youngest of retrovirus families in the human genome and is the only group that is polymorphic; a HERV-K can be present in one individual but absent from others. HERV-Ks could contribute to disease risk but establishing a link of a polymorphic HERV-K to a specific disease has been difficult. We develop an easy to use method that reveals the considerable variation existing among global populations in the frequency of individual and co-occurring polymorphic HERV-K, and in the total number of HERV-K that any individual has in their genome. Our study provides a global reference set of HERV-K genomic diversity and tools needed to determine the genomic landscape of HERV-K in any patient population.

genomics↗

bioRxiv2018

Synthetic essentiality of metabolic regulator PDHK1 in PTEN-deficient cells and cancers

PTEN is a tumor suppressor that is often inactivated in cancer and possesses both lipid and protein phosphatase activities. We report the metabolic regulator PDHK1 (pyruvate dehydrogenase kinase1) is a synthetic-essential gene in PTEN-deficient cancer and normal cells. The predominant mechanism of PDHK1 regulation and dependency is the PTEN protein phosphatase dephosphorylates NF{kappa};B activating protein (NKAP) and limits NF{kappa}B activation to suppress expression of PDHK1, a NF{kappa}B target gene. Loss of the PTEN protein phosphatase upregulates PDHK1 to drive aerobic glycolysis and induce PDHK1 cellular dependence. PTEN-deficient human tumors harbor increased PDHK1, which is a biomarker of decreased patient survival, establishing clinical relevance. This study uncovers a PTEN-regulated signaling pathway and reveals PDHK1 as a potential target in PTEN-deficient cancers.\n\nSIGNIFICANCEThe tumor suppressor PTEN is widely inactivated in cancers and tumor syndromes. PTEN antagonizes PI3K/AKT signaling via its lipid phosphatase activity. The modest success of PI3K/AKT inhibition in PTEN-deficient cancer patients provides rationale for identifying other vulnerabilities in PTEN-deficient cancers to improve clinical outcomes. We show that PTEN-deficient cells are uniquely sensitive to PDHK1 inhibition. PTEN and PDHK1 co-suppression reduced colony formation and induced cell death in vitro and tumor regression in vivo. PDHK1 levels were high in PTEN-deficient patient tumors and associated with inferior patient survival, establishing clinical relevance. Our study identifies a PTEN-regulated signaling pathway linking the PTEN protein phosphatase to the metabolic regulator PDHK1 and provides a mechanistic basis for PDHK1 targeting in PTEN-deficient cancers.

cancer biology↗

bioRxiv2018

Haplotype Analysis of the TRB Locus by TCRB Repertoire Sequencing

Polymorphism within the T cell receptor beta variable gene (TRBV) has been implicated in autoimmune disease and immuneCrelated adverse events (IRAEs) during immunotherapy. Previous efforts to evaluate TRBV polymorphism by whole genome sequencing (WGS) have been hampered by the repetitive nature of the TCRB locus. We present a novel longCamplicon TCRB repertoire sequencing approach to evaluate TRBV polymorphism from peripheral blood, which we use to identify TRBV allele haplotypes in 81 Caucasians.

immunology↗

bioRxiv2018

PIRD: Pan immune repertoire database

MotivationT and B cell receptors (TCRs and BCRs) play a pivotal role in the adaptive immune system by recognizing an enormous variety of external and internal antigens. Understanding these receptors is critical for exploring the process of immunoreaction and exploiting potential applications in immunotherapy and antibody drug design. Although a large number of samples have had their TCR and BCR repertoires sequenced using high-throughput sequencing in recent years, very few databases have been constructed to store these kinds of data. To resolve this issue, we developed a database.\n\nResultsWe developed a database, the Pan Immune Repertoire Database (PIRD), located in China National GeneBank (CNGBdb), to collect and store annotated TCR and BCR sequencing data, including from Homo sapiens and other species. In addition to data storage, PIRD also provides functions of data visualisation and interactive online analysis. Additionally, a manually curated database of TCRs and BCRs targeting known antigens (TBAdb) was also deposited in PIRD.\n\nAvailability and ImplementationPIRD can be freely accessed at https://db.cngb.org/pird.

immunology↗

bioRxiv2018

Differential T cell reactivity to hypocretin and flu antigen epitopes provides clues to the pathophysiology of Type 1 narcolepsy

Type 1 narcolepsy (T1N) is caused by hypocretin (HCRT) neuronal loss. Association with the Human Leukocyte Antigen (HLA)-DQB1*06:02/DQA1*01:02 (98% vs 25%) heterodimer (DQ0602), T cell receptor (TCR) and other immune loci suggest autoimmunity but autoantigen(s) are unknown. Onset is seasonal and associated with influenza A, notably pandemic 2009 H1N1 (pH1N1). An extensive unbiased DQ0602 binding peptide screen was performed encompassing peptides derived from Pandemrix(R) X-179-A pH1N1 influenza-A vaccine, a known T1N trigger, other H1N1 strains, and potential human autoantigens HCRT and RFX4, identifying 109 binders. The presence of cognate tetramer-peptide specific CD4+ T cells was studied in 35 narcolepsy cases and 22 DQ0602 controls after expansion of antigen-specific cells in Peripheral Blood Monocytes Cell (PBMC) cultures. Higher reactivity to influenza epitopes pHA273-287 (pH1N1 specific) and PR8 (H1N1 pre 2009)-specific NP17-31 were observed in T1N. Extensive reactivity to C-amidated but not native version of HCRT54-66 and HCRT86-97, which are two highly homologous peptides (HCRTNH2) was observed with higher frequencies of specific T cells in T1N. TCR/{beta} CDR3 sequences found in pHA273-287, NP17-31 and HCRTNH2 tetramer positive CD4+ cells were also retrieved in single INF{gamma}-secreting CD4+ sorted cells stimulated with Pandemrix(R), confirming immunodominance and functional significance in DQ0602-mediated responses and molecular mimicry. TCR/{beta} CDR3 motifs of HCRT54-66 and HCRT86-97 tetramers were extensively shared. Particularly notable was sharing across subjects of an CDR3, CAVETDSWGKLQF (in association with various CDR3{beta} that used TRAJ24, a chain modulated by Single Nucleotide Polymorphism (SNPs) rs1154155 and rs1483979 associated with T1N. Sharing of CDR3{beta} CASSQETQGRNYGYTF (in association with various CDR3 was also observed with HCRTNH2 and pHA273-287-tetramers across subjects. This segment uses TRBV4-2, a segment modulated by narcolepsy-associated SNP rs1008599. Higher HCRTNH2 positive CD4+ T cell numbers in T1N together with sharing of J24 CAVETDSWGKLQF in HCRTNH2 autoimmune responses, indicates causal DQ0602-mediated CD4+ autoreactivity to HCRT in T1N. Our results provide evidence for autoimmunity and molecular mimicry with flu antigens modulated by genetic components in the pathophysiology of T1N.

immunology↗

bioRxiv2018

Narcolepsy risk loci are enriched in immune cells and suggest autoimmune modulation of the T cell receptor repertoire

Type 1 narcolepsy (T1N) is a neurological condition, in which the death of hypocretin-producing neurons in the lateral hypothalamus leads to excessive daytime sleepiness and symptoms of abnormal Rapid Eye Movement (REM) sleep. Known triggers for narcolepsy are influenza-A infection and associated immunization during the 2009 H1N1 influenza pandemic. Here, we genotyped all remaining consented narcolepsy cases worldwide and assembled this with the existing genotyped individuals. We used this multi-ethnic sample in genome wide association study (GWAS) to dissect disease mechanisms and interactions with environmental triggers (5,339 cases and 20,518 controls). Overall, we found significant associations with HLA (2 GWA significant subloci) and 11 other loci. Six of these other loci have been previously reported (TRA, TRB, CTSH, IFNAR1, ZNF365 and P2RY11) and five are new (PRF1, CD207, SIRPG, IL27 and ZFAND2A). Strikingly, in vaccination-related cases GWA significant effects were found in HLA, TRA, and in a novel variant near SIRPB1. Furthermore, IFNAR1 associated polymorphisms regulated dendritic cell response to influenza-A infection in vitro (p-value =1.92*10-25). A partitioned heritability analysis indicated specific enrichment of functional elements active in cytotoxic and helper T cells. Furthermore, functional analysis showed the genetic variants in TRA and TRB loci act as remarkable strong chain usage QTLs for TRAJ*24 (p-value = 0.0017), TRAJ*28 (p-value = 1.36*10-10) and TRBV*4-2 (p-value = 3.71*10-117). This was further validated in TCR sequencing of 60 narcolepsy cases and 60 DQB1*06:02 positive controls, where chain usage effects were further accentuated. Together these findings show that the autoimmune component in narcolepsy is defined by antigen presentation, mediated through specific T cell receptor chains, and modulated by influenza-A as a critical trigger.

genetics↗

bioRxiv2018

Comparative genomic analysis revealed rapid differentiation in the pathogenicity-related gene repertoires between Pyricularia oryzae and Pyricularia penniseti isolated from a Pennisetum grass

BackgroundsPyricularia is a multispecies complex that could infect and cause severe blast disease on diverse hosts, including rice, wheat and many other grasses. Although the genome size of this fungal complex is small [~40 Mbp for Pyricularia oryzae (syn. Magnaporthe oryzae), and ~45 Mbp for P. grisea], the genome plasticity allows the fungus to jump and adapt to new hosts. Therefore, deciphering the genome basis of individual species could facilitate the evolutionary and genetic study of this fungus. However, except for the P. oryzae subgroup, many other species isolated from diverse hosts, such as the Pennisetum grasses, remain largely uncovered genetically.\n\nResultsHere, we report the genome sequence of a pyriform-shaped fungal strain P. penniseti P1609 isolated from a Pennisetum grass (JUJUNCAO) using PacBio SMRT sequencing technology. We performed a phylogenomic analysis of 28 Magnaporthales species and 5 non-Magnaporthales species and addressed P1609 into a Pyricularia subclade that is distant from P. oryzae. Comparative genomic analysis revealed that the pathogenicity-related gene repertoires were fairly different between P1609 and the P. oryzae strain 70-15, including the cloned avirulence genes, other putative secreted proteins, as well as some other predicted Pathogen-Host Interaction (PHI) genes. Genomic sequence comparison also identified many genomic rearrangements.\n\nConclusionTaken together, our results suggested that the genomic sequence of the P. penniseti P1609 could be a useful resource for the genetic study of the Pennisetum-infecting Pyricularia species.

genomics↗

bioRxiv2018

The landscape and diagnostic potential of T and B cell repertoire in Immunoglobulin A Nephropathy

Immunoglobulin A Nephropathy (IgAN) is the most common glomerulonephritis worldwide. In IgAN, immune complex deposite in glomerular mesangium, which induce inflammation and affect the kidneys normal functions. However, the exact pathogenesis of IgAN is still incompletely understood. Further, in current practice the clinical diagnosis relies on needle biopsy on renal tissue. Therefore, a non-invasive method for clinical diagnosis and prognosis surveillance of the disease is in high demand. In this paper, we investigated both the T cell receptor bata chain (TCRB) and immunoglobulin heavy chain (IGH) repertoire of kidney infiltrating and circulating lymphocytes of IgAN patients by immune repertoire high throughput sequencing. We found that the features of TCRB and IGH in the renal tissues were remarkably different from that in blood, including a decreased repertoire diversity and increased IgA and IgG frequency, and more activated B cells. The CDR3 length of PBMC TCRB and IGH in patients is significantly shorter than that in healthy controls, which is the result of both VDJ rearrangement and clone selection. We also found that the IgA1 frequency in the PBMC of IgAN is significant higher than that in other Nephropathy (NIgAN) and healthy control, which is consistent with the previous reports on the level of IgA1 producing B cells and serum IgA1. Significantly, we identified a set of IgAN disease related TCRB and IGH CDR3s, which can be used to distinguish IgAN from NIgAN and healthy controls from the blood with high accuracy. These results indicated that TCRB and IGH repertoire can potentially serve as non-invasive biomarkers for IgAN diagnosis. The characteristics of kidney infiltrating and circulating lymphocytes repertoire shed light on IgAN detection, treatment and surveillance.

immunology↗

bioRxiv2018

Illuminating microbial metabolic activities in the dark deep ocean with metaproteomics

The deep ocean is the largest habitat on earth and holds diverse microbial life forms. Significant advances have been made in microbial diversity and their genomic potential in the deep ocean, however, little is known about microbial metabolic activity that is crucial to regulate the bathypelagic carbon sequestration. Here, we characterized proteomes covering large particulate (>0.7 m), small particulate (0.2-0.7 m) and dissolved (10 kDa-0.2 m) fractions collected at a depth of 3000 m in the South China Sea. The Rhodospirillales, SAR324, SAR11, Nitrosinae/Tectomicrobia were the major contributors in the particulate fraction whereas Alteromonadales and viruses dominated the dissolved counterpart. Frequent detection of transcription or translation proteins in the particulate fractions indicated active metabolism of SAR324, Archaea, SAR11, and possible viable surface microbes, e.g. Prochlorococcus. Transporters for diverse substrates were the most abundant functional groups, and numerous spectra of formate dehydrogenases and glycine betaine transporters unveiled the importance of methylated compounds for the survival of deep-sea microbes. Notably, abundant non-viral proteins, especially transporters and cytoplasmic proteins, were detected in the dissolved fraction, indicating their potential roles in nutrient scavenging and the stress response. Our size-based proteomic study implied the holistic microbial activity mostly acting on the labile dissolved organic matter as well as the potential activities of surface microbes and dissolved non-viral proteins in the deep ocean.\n\nImportanceThe deep ocean produces one third of the biological CO2 in the ocean. However, little is known about metabolic activity of the bathypelagic microbial community which is crucial for understanding the biogeochemical cycling of organic matter, especially the formation of bulk refractory dissolved organic matter (DOM), one of the largest reservoirs of reduced carbon on Earth. This study provided the protein evidence firstly including both particulate and dissolved fractions to comprehensively decipher the active microbes and metabolic processes involved in the DOM recycling in the deep ocean. Our data supported the hypothesis of the carbon and energy supply from the labile DOM after the solution of sinking particles to the bathypelagic microbial community.

microbiology↗

bioRxiv2018

Circularization of genes and chromosome by CRISPR in human cells

Extrachromosomal circular DNA (eccDNA) and ring chromosomes are genetic alterations found in humans with genetic disorders and diseases such as cancer. However, there is a lack of genetic engineering tool to recapitulate these features. Here, we report the discovery that delivery of pairs of CRISPR/Cas9 guide RNAs into human cells generate functional eccDNAs and ring chromosomes. We generated a dual-fluorescence eccDNA biosensor system, which allows us to study CRISPR deletion, inversion, and circularization of genes inside cells. Analysis after CRISPR editing at intergenic and genic loci in human embryonic kidney 293T cells and human mammary fibroblasts reveal that CRISPR deleted DNA readily form eccDNA in human cells. DNA in sizes from a few hundred base pairs up to a 47.4 megabase-sized ring chromosome (chr18) can be circularized. Our discoveries advance and expand CRISPR-Cas9 technology applications for genetic engineering, modeling of human diseases, and chromosome engineering.\n\nOne Sentence Summary: CRISPR circularization of DNA offers new tools for studying eccDNA biogenesis, function, chromosome engineering, and synthetic biology.

genomics↗

bioRxiv2018

Identification of Variable and Joining germline genes and alleles for Rhesus macaque from B-cell receptor repertoires

The Rhesus macaque is a valuable preclinical animal model to estimate vaccine effectiveness, and is also important for understanding antibody maturation and B-cell repertoire evolution responding to vaccination; however, incomplete mapping of rhesus immunoglobulin germline genes hinders the research efforts. To address this deficiency, we sequenced B-cell receptor (BCR) repertoires of 75 India Rhesus macaques. Using a bioinformatic method that has been validated with BCR repertoire analysis of three human donors, we were able to infer rhesus Variable (V) and Joint(J) germline alleles, identifying a total of 122 V and 20 J germline alleles. Importantly, 91 V and 13 J alleles were novel, and 40 V and 13 J genes were found at a novel genome region that has not been previously recorded. The novelty of these newly identified alleles was supported by two observations. Firstly, 50 V and 5 J novel alleles were observed in whole genome sequencing data of 10 Rhesus macaques. Secondly, using alignment reference including the novel alleles, the mutation rate of rearranged repertoires was significant declined in 9 other irrelevant samples, and all our identified novel V and J alleles were 100% identity mapped by rearranged repertoire data. These newly identified novel alleles, along with previous reported alleles, provide an important reference for future investigations of rhesus immune repertoire evolution, in response to vaccination or infection. In addition, the method outlined in our study offered an example to future efforts in identifying novel immunoglobulin alleles.

immunology↗

bioRxiv2017

Resolution of Reprogramming Transition States by Single Cell RNA-Sequencing

The Yamanaka factors convert mouse embryonic fibroblasts (MEFs) into induced pluripotent stem cells (iPSCs) through a highly heterogeneous process. Here we profile single cells undergoing an optimized 7-day reprogramming process and show that cells start reprogramming relatively in sync, but diverge into two branches around day 2. The first branch of cells expressing Cd34/Fxyd5/Psca become nonpluripotent. The second one contains cells that are first Oct4+, then Dppa5a+ and pluripotent. We show that IFN-{gamma} blocks this late transition. Our results reveal the heterogeneous nature of somatic cell reprogramming, identify Dppa5a as a marker for pluripotent and innate immunity as a potential barrier for reprogramming.\n\nOne Sentence SummarySingle cell RNA sequencing reveals a continuum of cell fates from somatic to pluripotent and Dppa5a as a marker for chimera-competent iPSCs.

cell biology↗