2004 – A SINGLE-CELL PROTEOMICS BY MASS SPECTROMETRY BASED MAP OF THE HUMAN CD34+ HEMATOPOIETIC STEM AND PROGENITOR CELL COMPARTMENT

Our ability to characterize hematopoietic differentiation has been revolutionized by novel single cell technologies of which scRNAseq is undoubtedly the most influential. While this has led to novel insights into early hematopoietic decision events, e.g. how stem cells decide on their future fates, it is important to remember that mRNA levels are only proxies for the levels of the true cellular workhorses, i.e. the proteins. Since protein levels are regulated by additional cellular events such as translational initiation, elongation and protein decay, there is not necessarily a one-to-one relationship between mRNA and protein levels. Therefore, relying only on mRNA levels for the characterization of complex biological systems comes at a risk of missing important biological information.

Here, we present the first single-cell proteomics by Mass Spectrometry (scp-MS) based map of the human CD34+ hematopoietic stem and progenitor cells (HSPCs) compartment (>2,500 cells averaging approximately 1,000 proteins/cell). We used the GLUE autoencoder to integrate the scp-MS data with corresponding scRNAseq data to generate a common embedding, allowing us to compare mRNA and protein levels from similar computationally inferred cells. Trajectory analysis demonstrated high concordance between mRNA and protein levels along the granulocytic/monocytic and erythroid trajectories, whereas early HSC differentiation events were associated with significant lower concordance levels, highlighting the importance of protein-level data. We leveraged these findings to identify and validate novel regulators of early hematopoietic differentiation. This work demonstrates the feasibility and potential of scp-MS to gain novel insights into normal and, in the future, malignant hematopoiesis.