The transcriptomic landscape of normal and ineffective erythropoiesis at single cell resolution

Ineffective erythropoiesis, the death of maturing erythroid cells, is a
common cause of anemia. To better understand why this occurs, we studied
the fates and adaptations of single erythroid marrow cells from
individuals with Diamond Blackfan anemia (DBA), del(5q) myelodysplastic
syndrome (del(5q) MDS), and normal controls, and defined an unhealthy (vs.
healthy) differentiation trajectory, using velocity pseudotime and cell
surface protein assessment. The pseudotime trajectories diverge
immediately after the cells upregulate transferrin receptor (CD71), import
iron, and initiate heme synthesis, although cell death occurs much later.
Cells destined to die highly express heme-responsive genes, including
ribosomal protein and globin genes. In contrast, surviving cells
downregulate heme synthesis, while upregulating DNA damage response,
hypoxia, and HIF1 pathways. Surprisingly, 24±12% of cells from controls
follow the unhealthy trajectory, implying that heme also regulates cell
fate decisions during normal red cell production. Del(5q) MDS (unlike DBA)
results from somatic mutations, so many normal (unmutated) erythroid cells
persist. By independently tracking their trajectory, we gained insight
into why they cannot expand to prevent anemia. In addition, we show that
intron retention is especially prominent during red cell differentiation.
The additional information provided by messages with retained introns also
allowed us to align data from multiple independent experiments and thus
accurately query the transcriptomic changes that occur as single erythroid
cells mature.