SEC-seq: Association of molecular signatures with antibody secretion in thousands of single human plasma cells

Abstract

Protein secretion drives many functions in vivo; however, methods to link secretions with surface markers and transcriptomes have been lacking. By accumulating secretions close to secreting cells held within cavity-containing hydrogel nanovials, we demonstrate workflows to analyze the amount of IgG secreted from single human antibody-secreting cells and link this information to surface marker expression and transcriptional profiles from the same cells. Measurements using flow cytometry and imaging flow cytometry corroborated an association between levels of IgG secretion and CD138 expression. Using oligonucleotide-labeled antibodies and droplet-based sequencing, we show that pathways encoding protein localization to the endoplasmic reticulum, NADH complex assembly, and mitochondrial respiration were most associated with high IgG secretion. Altogether, this method links secretion information to cell surface and single-cell sequencing information (SEC-seq) and enables exploration of links between genome and secretory function, laying the foundation for numerous discoveries in immunology, stem cell biology, and beyond.

Secretion encoded single-cell sequencing (SEC-seq) uncovers gene expression signatures associated with high VEGF-A secretion in mesenchymal stromal cells

“…Often the expression of a secretory protein is equated to secretion, however, our results suggest this assumption may not be correct across different cell types 41 . We found that VEGFA transcripts and VEGF-A secretion have very low correlation in normoxic and hypoxic MSCs.…”

“…Cell isolation schemes such as FACS sorting are often used prior to single-cell sequencing to enrich sub-populations, but many isolation or adherent cell suspension methods affect data quality. Although this work addressed secretion in adherent MSCs, suspension cells, such as B cells, are also compatible with SEC-seq through antibody capture on nanovials 41 .…”

Advancing Cell Therapies: Single-Cell Profiling, Generation, Expansion, and Gene Delivery in Rhesus Macaque Plasma B Cells

“…In contrast to previous findings in human cells 13 , we observed an upregulation of several MHC-class I molecules in NHP PB/PCs. We previously reported that CD59 is highly expressed in human IgG PCs and correlated with immunoglobulin secretion 2 . Because of its robust expression in NHP PCs, and its correlation with PC function in human cells, we propose that CD59, and possibly other genes in this dataset, are likely to be useful surrogate markers for functional NHP PCs.…”

“…Recently, we developed a cell-based method to deliver protein drugs, which we have successfully tested in immune-deficient mice. To do this, we generated ex vivo differentiated human plasma cells (PCs) and engineered them to produce protein drugs (including bispecific antibodies), and in immuno-deficient mice, we observed long-lasting antibody secretion for a year and potent tumor killing [1][2][3][4] .…”

Defining T cell receptor repertoires using nanovial-based affinity and functional screening

“…Recent work has emphasized the importance of functional characterization of TCRs, such as through assaying Ca 2+ flux upon mechanical engagement of TCRs with pMHC-coated hydrogel beads, a platform that could be synergistic with nanovials to more fully functionally screen TCRs ( 28 ). Separately, we have shown that oligonucleotide-barcoded antibodies can be used as labels in the nanovial assay format (see secretion-encoded single cell (SEC)-seq workflows) ( 29 ). Using barcoded antibodies to label secreted cytokines would allow encoding of this cellular function into the single-cell sequencing data set and ranking of TCR sequences based on the amount of cytokine(s) secreted.…”