Proteomic analysis of young and old mouse hematopoietic stem cells and their progenitors reveals post-transcriptional regulation in stem cells.

Balyn W Zaro,Gunsagar S Gulati,Monika Zukowska,Allison Banuelos,Benson George,Janos Demeter,Irving L Weissman,Rahul Sinha,Adam C Wilkinson,Peter Jackson,Ryan A Flynn,Joseph J Noh,Victoria L Mascetti,Allison Zhang

doi:10.7554/elife.62210

Balyn W Zaro, Gunsagar S Gulati + Show 12 more

Open Access

https://doi.org/10.7554/elife.62210

Copy DOI

Abstract

The balance of hematopoietic stem cell (HSC) self-renewal and differentiation is critical for a healthy blood supply; imbalances underlie hematological diseases. The importance of HSCs and their progenitors have led to their extensive characterization at genomic and transcriptomic levels. However, the proteomics of hematopoiesis remains incompletely understood. Here we report a proteomics resource from mass spectrometry of mouse young adult and old adult mouse HSCs, multipotent progenitors and oligopotent progenitors; 12 cell types in total. We validated differential protein levels, including confirmation that Dnmt3a protein levels are undetected in young adult mouse HSCs until forced into cycle. Additionally, through integrating proteomics and RNA-sequencing datasets, we identified a subset of genes with apparent post-transcriptional repression in young adult mouse HSCs. In summary, we report proteomic coverage of young and old mouse HSCs and progenitors, with broader implications for understanding mechanisms for stem cell maintenance, niche interactions and fate determination.

Highlights

Hematopoietic stem cells (HSCs) are responsible for persistent renewal of blood and immune cells throughout a lifetime
We created a workflow whereby cells were purified by FACS using three sorting panels allowing for the isolation of 12 cells types: young adult and old adult HSC (Lin, cKit+, Sca1+, CD34, CD150+, Flt3-), MPPa (Lin, cKit+, Sca1+, CD34+, CD150+, Flt3-), young adult and old adult MPPb (Lin, cKit+, Sca1+, CD34+, CD150, Flt3-), young adult and old adult MPPc (Lin, cKit+, Sca1+, CD34+, CD150, Flt3+), young adult CLP (Lin, cKitlo, Sca1lo, Flt3+, IL7Ra+), young adult CMP (Lin, cKit+, Sca1lo/, CD34med/hi, CD16/32-/lo), young adult MEP (Lin, cKit+, Sca1lo/, CD34, CD16/32-/lo, CD150+) and young adult GMP (Lin, cKit+, Sca1lo/, CD34hi, CD16/32hi) (Figure 1A and Figure 1—figure supplement 1)
For all young and old adult mouse HSCs, at least three biological cohorts of mice were utilized for each cell type, and the sample was run in technical duplicate with 200 ng (~7000 cells) of loading material per replicate

Summary

Introduction

Hematopoietic stem cells (HSCs) are responsible for persistent renewal of blood and immune cells throughout a lifetime. They have the ability to self-renew, and differentiate into effector cells in response to physiological demands such as infection or bleeding (Figure 1A; Spangrude et al, 1988; Baum et al, 1992; Seita and Weissman, 2010). HSCs are the locus of disease-causative mutations in a number of relatively common blood diseases and leukemias. HSC clones sustaining several driver mutations inhibit differentiation, drive proliferation, block programmed cell death and phagocytosis, and outcompete

Methods

Results

Conclusion