Linking phenotype with transcriptomic and clonotype of the adaptive immune response in human
Simone Rizzettoab, A. Elthalaab, M. Singhc, Mehdi R. Pirozyanab, J. Samirab, E. Keoshkerianab, C. H. Caiab, V. Venturib, A. R. Lloydab, R. A. Bullab and F. Lucianiab
aSchool of Medical Science, UNSW, Australia; bKirby Institute, UNSW, Australia; cGarvan Institute of Medical Research, Australia
The diversity of the B and T cells receptors (BCR and TCR) constitutes an important feature of a successful immune response as it define the specificity and breadth of the adaptive immune response against pathogens. The full length of these heterodimers receptors has been difficult to study given the complex genetic rearrangements and the highly diverse repertoire present in any subject. During an immune response, both B and T cells undergo a series of phenotypic changes, usually associated to the expression of specific surface markers, however the full extent of heterogeneity remains still unknown. Recent studies revealed a high level of transcriptomic variability even in cells with the same clonotype and surface phenotype, suggesting that further subsets may be identified along with distinct functions.
With single cell RNA-sequencing (scRNA-seq) is now possible to measure the gene expression of individual cells. By combining scRNA-seq with flow cytometry index sorting we have developed a framework (called VDJPuzzle) to simultaneously measure gene expression profile and surface protein expression as well as reconstructing the full BCR and TCR sequence.
Our software requires output data from scRNA-seq and from flow cytometry machine (FCS file). This method has been successfully applied to both human T and B cell data in HCV infection and in vaccine samples against influenza virus, identifying new cellular sub-populations belonging to the same clonotype, but characterized by distinct transcriptomic signature. VDJPuzzle is also provided with a graphical user interface that help users through the execution of the pipeline and show summary statistics.
Credits: None available.
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