Molecular dissection of blood cell fate determination.
Our aims are two-fold: (i) to understand the molecular mechanisms underlying specification of the blood lineage from mesodermal cells and (ii) to model blood stem cell development in vitro.
(i) Molecular mechanisms underlying lineage specification
During development, mesodermal cells differentiate into several lineages such as the blood, cardiac and paraxial tissues. Despite an established understanding of cell movements during gastrulation, the exact molecular mechanisms driving lineage fate decisions and the developmental relationship between lineages are still unclear. To study these questions, we take two complementary approaches. First, we examine the function and mechanisms of action of the basic helix-loop-helix transcription factor and oncoprotein SCL/Tal-1. SCL, originally identified by virtue of its involvement in T-cell Acute Lymphoblastic Leukaemia (T-ALL), is essential to confer blood fate to mesodermal cells. It is also required to suppress development of alternative mesodermal lineages, therefore offering an excellent entry point for mechanistic studies of cell fate determination. Second, we identify and characterise the earliest lineage-committed progenitors in developing embryos to investigate the intricate developmental relationships between blood and alternative mesodermal lineages.
To address these questions, we use state-of-the-art technologies: lineage tracing and single cell approaches, single molecule mRNA imaging, high-throughput genomic and transcriptomic assays (such as ATAC-, ChIP- and RNA-seq to document chromatin accessibility, gene expression and protein:DNA interactions genome-wide), proteomics studies and CRISPR/Cas9-based technologies to engineer new molecular and cellular tools. A large part of our work uses in vitro differentiation assays of embryonic stem (ES) cells. Altogether, this will allow characterisation of gene regulatory networks at the heart of blood development and provide principles to a recurring question in developmental biology: at what stage do lineage-fated cells arise and what is the extent of multi-potentiality of common progenitors?
(ii) Modelling blood stem cell development in vitro
Over the past decade, numerous efforts have been deployed to produce the elusive blood (haematopoietic) stem cell (HSC) in vitro from pluripotent stem cells (PSCs) for mechanistic and therapeutic purposes. However, none of the differentiation cultures developed so far have been able to produce long-term repopulating HSCs, the cells with self-renewal and multilineage potentialities that give rise to the entire blood system when transplanted into a host organism. This is likely to reflect the lack of critical signalling molecules supporting HSC development in the current differentiation cultures. We aim to carefully recapitulate haematopoietic development in serum-free mouse and human PSC differentiation cultures, as it happens during embryonic development. To do this, we apply our knowledge of the signalling pathways controlling HSC development into in vitro differentiation protocols for a tight control of the stepwise development of PSCs towards HSCs. Cell fate replating studies combined with single cell transcriptomics, epigenetics and chromatin accessibility approaches will provide information on the evolution of the molecular landscape as the cells transit through the stages leading to production of HSCs. Ultimately, our goal is to translate our findings to the clinical setting.
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