Biology subjects

Bieberich, F.

Publications and source records attributed to Bieberich, F..

5 recordsLinked to original sources

bioRxiv2021

A single-cell atlas of lymphocyte adaptive immune repertoires and transcriptomes reveals age-related differences in convalescent COVID-19 patients

COVID-19 disease outcome is highly dependent on adaptive immunity from T and B lymphocytes, which play a critical role in the control, clearance and long-term protection against SARS-CoV-2. To date, there is limited knowledge on the composition of the T and B cell immune receptor repertoires [T cell receptors (TCRs) and B cell receptors (BCRs)] and transcriptomes in convalescent COVID-19 patients of different age groups. Here, we utilize single-cell sequencing (scSeq) of lymphocyte immune repertoires and transcriptomes to quantitatively profile the adaptive immune response in COVID-19 patients of varying age. We discovered highly expanded T and B cells in multiple patients, with the most expanded clonotypes coming from the effector CD8+ T cell population. Highly expanded CD8+ and CD4+ T cell clones show elevated markers of cytotoxicity (CD8: PRF1, GZMH, GNLY; CD4: GZMA), whereas clonally expanded B cells show markers of transition into the plasma cell state and activation across patients. By comparing young and old convalescent COVID-19 patients (mean ages = 31 and 66.8 years, respectively), we found that clonally expanded B cells in young patients were predominantly of the IgA isotype and their BCRs had incurred higher levels of somatic hypermutation than elderly patients. In conclusion, our scSeq analysis defines the adaptive immune repertoire and transcriptome in convalescent COVID-19 patients and shows important age-related differences implicated in immunity against SARS-CoV-2.

immunology↗

bioRxiv2021

Single-cell sequencing of plasma cells from COVID-19 patients reveals highly expanded clonal lineages produce specific and neutralizing antibodies to SARS-CoV-2

Isolation and characterization of antibodies in COVID-19 patients has largely focused on memory B cells, however it is the antibody-secreting plasma cells that are directly responsible for the production of serum antibodies, which play a critical role in controlling and resolving SARS-CoV-2 infection. To date there is little known about the specificity of plasma cells in COVID-19 patients. This is largely because plasma cells lack surface antibody expression, which complicates their screening. Here, we describe a technology pipeline that integrates single-cell antibody repertoire sequencing and high-throughput mammalian display screening to interrogate the specificity of plasma cells from 16 convalescent COVID-19 patients. Single-cell sequencing allows us to profile antibody repertoire features in these patients and identify highly expanded clonal lineages. Mammalian display screening is employed to reveal that 37 antibodies (out of 132 candidates) derived from expanded plasma cell clonal lineages are specific for SARS-CoV-2 antigens, including antibodies that target the receptor binding domain (RBD) with high affinity and exhibit potent neutralization of SARS-CoV-2. One Sentence SummarySingle-cell antibody repertoire sequencing and high-throughput screening identifies highly expanded plasma cells from convalescent COVID-19 patients that produce SARS-CoV-2-specific antibodies capable of potent neutralization.

immunology↗

bioRxiv2020

Platypus: an open-access software for integrating lymphocyte single-cell immune repertoires with transcriptomes

High-throughput single-cell sequencing (scSeq) technologies are revolutionizing the ability to molecularly profile B and T lymphocytes by offering the opportunity to simultaneously obtain information on adaptive immune receptor repertoires (VDJ repertoires) and transcriptomes. An integrated quantification of immune repertoire parameters such as germline gene usage, clonal expansion, somatic hypermutation and transcriptional states opens up new possibilities for the high-resolution analysis of lymphocytes and the inference of antigen-specificity. While multiple tools now exist to investigate gene expression profiles from scSeq of transcriptomes, there is a lack of software dedicated to single-cell immune repertoires. Here, we present Platypus, an open-source software platform providing a user-friendly interface to investigate B cell receptor (BCR) and T cell receptor (TCR) repertoires from single-cell sequencing experiments. Platypus provides a framework to automate and ease the analysis of single-cell immune repertoires while also incorporating transcriptional information involving unsupervised clustering, gene expression, and gene ontology. To showcase the capabilities of Platypus, we use it to analyze and visualize single-cell immune repertoires and transcriptomes from B and T cells from convalescent COVID-19 patients, revealing unique insight into the repertoire features and transcriptional profiles of clonally expanded lymphocytes. Platypus will expedite progress by increasing accessibility to the broader immunology community by facilitating the analysis of single-cell immune repertoire and transcriptome sequencing.

immunology↗

bioRxiv2020

Transposable element expression at unique loci in single cells with CELLO-seq

The role of Transposable Elements (TEs) in regulating diverse biological processes, from early development to cancer, is becoming increasing appreciated. However, unlike other biological processes, next generation single-cell sequencing technologies are ill-suited for assaying TE expression: in particular, their highly repetitive nature means that short cDNA reads cannot be unambiguously mapped to a specific locus. Consequently, it is extremely challenging to understand the mechanisms by which TE expression is regulated and how they might themselves regulate other protein coding genes. To resolve this, we introduce CELLO-seq, a novel method and computational framework for performing long-read RNA sequencing at single cell resolution. CELLO-seq allows for full-length RNA sequencing and enables measurement of allelic, isoform and TE expression at unique loci. We use CELLO-seq to assess the widespread expression of TEs in 2-cell mouse blastomeres as well as human induced pluripotent stem cells (hiPSCs). Across both species, old and young TEs showed evidence of locus-specific expression, with simulations demonstrating that only a small number of very young elements in the mouse could not be mapped back to with high confidence. Exploring the relationship between the expression of individual elements and putative regulators revealed surprising heterogeneity, with TEs within a class showing different patterns of correlation, suggesting distinct regulatory mechanisms.

molecular biology↗

bioRxiv2020

CRISPR-targeted display of functional T cell receptors enables engineering of enhanced specificity and prediction of cross-reactivity

T cell receptor (TCR) gene therapy is a promising cell therapy approach for the treatment of cancer. However, most naturally occurring TCRs display low affinities to their peptide-MHC targets, and engineering of TCRs for enhanced affinity is complicated by the risk of introducing cross-reactivity and the poor correlation between affinity and function. Here we report the establishment of the TCR-accepting T cell (TnT) platform through five sequential CRISPR-Cas9 genome editing steps of a human T cell line, and demonstrate its application for functional engineering of TCRs and prediction of cross-reactivity. Using the TnT platform, we profile the mutational landscapes of tumor-specific TCRs at high-throughput to reveal a substantial discordance between antigen binding and antigen-induced signaling. Furthermore, we combine CRISPR-targeting, functional selection and deep sequencing to screen TCR mutagenesis libraries and identify variants with enhanced recognition of the cancer-testis antigen MAGE-A3. Finally, functional cross-reactivity profiling using TnT cells was able to accurately predict off-targets and identify engineered TCRs with exquisite specificity to MAGE-A3. Thus, the TnT platform represents a valuable technology for the engineering of TCRs with enhanced functional and safety profiles.

immunology↗