CRUK Career Development Fellow

Email: as889@cam.ac.uk

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Research themes:

Regulation of Haematopoiesis in Homeostasis and Disease

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Description of research

Blood stem cells need to both perpetuate (self-renew) themselves and differentiate into all mature blood cells to maintain blood formation throughout life. However, it is unclear how the underlying gene regulatory network maintains this population of self-renewing and differentiating stem cells, and how it accommodates the transition from a stem cell to a mature blood cell. Clarifying how HSCs differentiate into diverse cell types is important for understanding how this process is subverted in the generation of blood pathologies. The aim of my group is to bridge this knowledge gap by providing a method in a relevant model organism (zebrafish, Danio rerio) that will allow us to dissect the role of novel blood genes and to determine their hierarchical position in regulatory networks that underlie haematopoiesis.

Our current program is divided into two research themes:
1. Functional genomics
2. Single cell transcriptomics

1. Functional genomics

In the last few years, DNA sequencing technologies have been developed that allow the identification of every genetic change in a given cancer sample, promising a new harvest of cancer genes. However, many of the identified genes had not previously been implicated in blood formation and there is a real need to investigate the biology and potential therapeutic aspects of these genes. This will be achieved by pursuing two main objectives:

First, a high-throughput screen in zebrafish to dissect the functional role of novel cancer genes, implicated in myelodysplasia and myeloproliferative neoplasms, in haematopoiesis. This effort will define the function of novel cancer genes in blood cell formation by a morpholino and CRISPR/Cas knock-down approach in zebrafish.

This early objective will be followed by a far more ambitious long-term one. This objective will be based on in-depth functional characterisation of a subset of genes.

2. Single cell transcriptomics

Our current knowledge of transcriptomes of various blood cell types has mainly been advanced by population-level analysis. However, the population of seemingly homogenous blood cells may include many distinct cell types with substantially different transcriptomes and abilities to make diverse fate decisions. To overcome these limitations, we will use single-cell transcriptome sequencing of zebrafish blood cells. We will apply an integrative strategy, combining genetic perturbation with computational sequence and network analysis methods, to reconstruct the regulatory networks that maintain the dynamic balance between different blood cell types.

1) We will create a comprehensive atlas of single cell gene expression in adult zebrafish blood cells and computationally reconstruct the blood lineage tree. We will order cells according to their most likely developmental chronology and identify genes and gene regulatory networks that define distinct cell types.

2) In the next step we will generate a number of loss-of-function and transgenic zebrafish lines.

By sequencing thousands of single cells, this study is poised to go beyond traditional approaches in examining the complex relationships between the continuous spectra of blood cells, and will provide unprecedented insight into the regulation of blood cell formation.

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Key Publications:

Botthof JG, Bielczyk-Maczyńska E, Ferreira L, Cvejic A (2017). Loss of the homologous recombination gene rad51 leads to Fanconi anemia-like symptoms in zebrafish. PNAS, Accepted. Available in biorxiv: http://biorxiv.org/content/early/2016/12/20/095646.

Svensson V, Natarajan KN, Ly LH, Miragaia RJ, Labalette C, Macaulay IC, Cvejic A, Teichmann SA (2017) Power Analysis of Single Cell RNA‐Sequencing Experiments. Nature Methods, 14(4):381-387.

Carmona SJ, Teichmann SA*^,♯, Ferreira L, Macaulay IC, Stubbington MJT, Cvejic A*^,♯, Gfeller D*^,♯ (2017). Single-cell transcriptome analysis of fish immune cells provides insight into the evolution of vertebrate immune cell types. Genome Research, 2017 Jan 13. pii: gr.207704.116. (^♯corresponding author, *joint senior author)

Dee CT, Nagaraju TR, Athanasiadis EI, Gray C, del Ama LF, Johnston SA, Secombes CJ*^,♯, Cvejic A*^,♯, Hurlstone AFL*^,♯ (2016). CD4-Transgenic Zebrafish Reveals Tissue Resident T_H2- and T_reg-like T cell Populations and Diverse Mononuclear Phagocytes. The Journal of Immunology, 197(9):3520-30 (^♯corresponding author, *joint senior authors)

Macaulay IC, Svensson V, Labalette C, Ferreira L, Hamey F, Voet T, Teichmann S, Cvejic A (2016). Single cell RNA-sequencing reveals a continuous spectrum of differentiation in haematopoietic cells. Cell Reports, 14(4):966-7.

Cvejic A (2016). Mechanisms of fate decision and lineage commitment during haematopoiesis. Immunology and Cell Biology, doi: 10.1038/icb.2015.96, Invited Review.

Bielczyk-Maczyńska E, Lam Hung L, Ferreira L, Fleischmann T, Weis F, Fernández-Pevida A, Harvey SA, Wali N, Warren AJ, Barroso I, Stemple DL, Cvejic A (2015). The ribosome biogenesis protein Nol9 is essential for definitive hematopoiesis and pancreas morphogenesis in zebrafish. PLOS Genetics, 11(12):e1005677.

Chen L, Kostadima M, [59 co-authors], Cvejic A, Soranzo N, Ouwehand WH, Stunnenberg HG, Frontini M, Rendon A (2014). Transcriptional diversity during lineage commitment of human blood progenitors. Science, 345(6204).

Bielczyk-Maczyńska E, Serbanovic-Canic J, Ferreira L, Soranzo N, Stemple D, Ouwehand WH, Cvejic A (2014). A loss of function screen of identified genome-wide association study loci reveals new genes controlling haematopoiesis. PLOS Genetics, 10(7):e1004450.

Cvejic A*^,♯, Haer-Wigman L*, Stephens JC*, Kostadima M, Smethurst PA, Frontini M, Sipos B, Akker Evd, Bertone P, Bielczyk E, Farrow S, Fehrmann RSN, Gray A, Haas M, Haver VG, Jordan G, Karjalainen J, Kerstens HHD, Kiddle G, Loyd-Jones H, Needs M, Poole J, Soussan A, Rendon A, Rieneck K, Sambrook JG, Schepers H, Siljer HHW, Swinkels D, Tamuri AU, Verweij N, Watkins NA, Westra HJ, Stemple D, Franke L, Soranzo N, Stunnenberg HG, Goldman N, Harst Pvd, C Schoot Evd, Ouwehand WH^♯, Albers C^♯ (2013). The red blood cell GWAS gene SMIM1 underlies the Vel blood group and is a novel regulator of red blood cell formation. Nature Genetics, 45(5):542-5. (^♯corresponding author, *joint first authors)