Preprint: researchers suppressed 520 genes to map hematopoietic stem cell function
Preprint: researchers suppressed 520 genes to map hematopoietic stem cell function
On August 29, a preprint appeared on bioRxiv. The authors partially suppressed 520 genes in primary cultures of mouse hematopoietic stem cells and their immediate progenitors. They measured the response of individual cells and identified 19 programs, defined as recurring changes in gene activity caused by different perturbations. They then compared these programs with data on the number of progeny produced by stem cells after transplantation and with existing datasets on aging and acute myeloid leukemia.
Hematopoietic stem cells continually replenish the blood and parts of the immune system. Their fate depends on the balance among quiescence, division, and differentiation into various blood cell types. Conventional gene activity maps show which genes are active together. In this study, the authors tested how cellular processes change when individual genes are suppressed.
The authors used CRISPRi-Perturb-seq, a method that selectively reduces the activity of one gene and then measures the activity of the remaining genes in each cell. They used three guide RNAs for each of 520 genes and added 120 controls. After quality control, the analysis included transcriptomes from 111 003 cells.
Cells in this type of culture already occupy different states, and the descendants of one original cell may expand faster than others. The researchers therefore compared each perturbed cell with the mean expression profile of its 20 nearest control cells in the same state. This approach allowed them to isolate the local response to gene suppression and retain responses that were reproduced by multiple guide RNAs targeting the same gene.
From these consistent responses, the authors derived 19 co-regulatory programs. Each program contains genes that change in similar ways when several regulators are suppressed. The analysis also identifies which regulators shift each program. The programs include DNA replication, cell division, the inflammatory response, and early specification toward lymphocyte lineages.
The authors then applied this set of programs to previously published mouse datasets. In a transplantation experiment, high activity of one program, GRP18, was associated with the number of progeny cells produced by a stem cell clone. In another dataset, GRP18 was more active in the daughter cell that retained stem cell identity after asymmetric division. The resulting programs allowed the researchers to compare responses to gene suppression with independent data on stem cell function.
Using the same approach, the authors analyzed hematopoietic cell profiles from 148 people aged 23 to 91 years. The inflammatory program was associated with age, while the early lymphoid specification program declined. In existing data from patients with acute myeloid leukemia, a blood cancer, individual programs were also associated with survival and response to venetoclax.
A June analysis described a cell atlas from people of different ages that showed coordinated shifts in gene activity and DNA regions. This preprint provides a way to test the relationships in such an atlas: suppress a specific regulator, measure the resulting local shift in a program, and compare that program with data on hematopoietic stem cell function.