PF453 TRANSCRIPTOME‐PROTEOME CORRELATION IN HUMAN HEMATOPOIETIC STEM AND PROGENITOR CELLS
Authors: Meier‐Abt F. , Amon S. , Gillet L. , Wolski W. , Dimitrieva S. , Theocharides A. , Manz M. , Aebersold R. 
Published: December 2023

Background:

Hematopoietic stem cells (HSCs) can self‐renew and/or differentiate into various functionally divergent progenitor cell types, such as common myeloid progenitors (CMPs), megakaryocyte‐erythrocyte progenitors (MEPs) or granulocyte‐macrophage progenitors (GMPs). When the process of self‐renewal and differentiation is altered, e.g. upon genetic or epigenetic changes in HSCs, abnormal (pre)leukemic stem cell subpopulations may form, eventually resulting in the onset of hematopoietic malignancies. To gain insight into the physiology and patho‐physiology of self‐renewal and differentiation, highly refined analyses of HSCs and downstream progenitor cells are needed. Whereas such small cell subpopulations have been routinely analyzed by genomics and transcriptomics, corresponding proteomic analyses have so far not been possible due to methodological limitations.

Aims:

To develop a new integrated workflow for the high‐coverage, quantitative proteome profiling of minute amounts of sorted cells, its application to human hematopoietic stem and progenitor cell subpopulations, and the comparison of proteomic and transcriptomic profiles in human HSCs and downstream progenitor cells.

Methods:

We developed a new ultra‐sensitive and robust quantitative proteomics technique based on data‐independent acquisition mass spectrometry. We quantified the proteome of sets of 25,000 human hematopoietic stem/multipotent progenitor (HSC) and three committed progenitor cell subpopulations of the myeloid differentiation pathway (CMPs, MEPs, GMPs), isolated by fluorescence‐activated cell sorting from five healthy donors. On average, 5,851 protein groups were identified per sample, and a subset of 4,131 stringently filtered protein groups was quantitatively compared across the 20 samples. The proteomic analyses were complemented by transcriptomic analyses.

Results:

A comparison of proteomic and transcriptomic profiles of the respective cell types indicated hematopoietic stem/multipotent progenitor cell‐specific divergent regulation of biochemical processes essential for maintaining stemness at the proteome rather than transcriptome level. Specifically, several telomerase maintenance proteins and quiescence‐inducing isocitrate dehydrogenase proteins, both assumed to be essential for long‐lived stem cells, were found to be upregulated in HSCs on the protein but not on the mRNA level when compared to myeloid progenitor cell subpopulations (CMPs, MEPs, GMPs).

Summary/Conclusion:

The divergent mRNA/protein regulation of telomerase maintenance and quiescence‐inducing isocitrate dehydrogenase proteins in HSCs illustrates the relevance of generating high quality proteomic data for well‐defined cell subpopulations with the goal to identify biological processes that are insufficiently determined by genomic or transcriptomic analyses. The presented methodology is equally applicable to almost any rare cell type, including healthy and cancer stem cells or physiologically and pathologically infiltrating cell populations. It thus paves the way for proteomic profiling of relevant disease sample sub‐fractions and, ultimately, could allow to find new therapeutic targets in (pre)leukemic/cancer stem cells.

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