Transcriptional control of haematopoietic specification and differentiation

Lead Research Organisation: University of Oxford

Abstract

Our interests focus on the mechanisms that underlie blood formation and differentiation into mature blood cells (such as those fighting infection, involved in coagulation or in oxygen transport). Many inherited and acquired diseases are associated with anaemia and leukaemia in which production of certain types of blood cells is disrupted. Understanding these disorders depends on knowing how blood normally forms in the bone marrow and throughout development.
Recently, there has been considerable accumulation of knowledge about the factors that direct formation of the blood. One such factor (called SCL, Stem Cell Leukaemia) plays several critical roles in this process. We plan to understand how this factor controls the formation of blood at a molecular level.
Our main experimental model makes use of mouse embryonic stem (mES) cells that can reproduce the early steps of embryonic development in vitro (in a culture dish).
We aim at understanding how a blood-specific protein complex forms, how it regulates the coordinated expression of the critical players involved in this pathway and how, when it is deregulated, it may lead to leukaemia and anaemia.

Technical Summary

Understanding how stem cells are specified during embryonic development and how lineages differentiate to produce mature and specialised cells are central questions in biology and are of crucial interest to elucidate pathways involved in diseases.
The main aim of this programme is to characterise some of the molecular mechanisms engaged by key regulators of blood development (such as SCL/Tal1), with a particular focus on the early stages of mesoderm patterning and haematopoietic specification.
Functional identification of protein complexes and their nuclear targets combined with molecular characterisation of their mechanisms of action and structural biology approaches will give insights into genetic interactions and transcriptional networks critical in decision pathways. Using in vitro differentiation of mouse ES cells, mouse haematopoiesis and Xenopus embryos as experimental models, we will start building networks of genetic interactions at the heart of haematopoietic development.
These approaches are instrumental for our understanding of how a haematopoietic stem cell-specific protein complex might form and how, when this process is deregulated, this leads to leukaemia. Moreover, this might help characterise the pathways leading to definitive haematopoiesis and HSCs from hES and iPS cells that remain poorly characterised. This is important if one wants to be able to produce HSCs in vitro for regenerative medicine purposes.

Publications

10 25 50
 
Description Project Grant
Amount £80,000 (GBP)
Organisation Leukaemia and Lymphoma Research 
Sector Charity/Non Profit
Country United Kingdom
Start 01/2012 
End 01/2013
 
Description Project Grant
Amount £300,000 (GBP)
Organisation British Heart Foundation (BHF) 
Sector Charity/Non Profit
Country United Kingdom
Start 05/2017 
End 05/2020
 
Description Responsive mode Research Grant
Amount £343,000 (GBP)
Organisation Biotechnology and Biological Sciences Research Council (BBSRC) 
Sector Public
Country United Kingdom
Start 01/2015 
End 01/2018
 
Title MKD1 cell line 
Description Generation of a new mouse megakaryocytic cell line 
Type Of Material Cell line 
Year Produced 2012 
Provided To Others? Yes  
Impact Very good alternative to using primary mouse mekagaryocyte progenitors. 
 
Title Scl:mCherry ES cell line 
Description Insertion of the mCherry reporter gene into the Scl locus in mouse ES cells 
Type Of Material Cell line 
Year Produced 2018 
Provided To Others? Yes  
Impact We can now follow the cellular fate of Scl-null cells. This has allowed to report a cell fate change from blood to cardiac/paraxial lineages in absence of this transcriptional regulator. This is a critical finding for our mechanistic understanding of lineage specification. 
URL https://www.nature.com/articles/s41467-018-07787-6
 
Title X-ray structure of LMO2-LDB1(LID) 
Description Crystal structure of a complex of two transcriptional regulators involved in many critical developmental and differentiation pathways. 
Type Of Material Biological samples 
Provided To Others? No  
Impact A better understanding of the mechanism of action of a key component of a multiprotein complex. This has increased our knowledge on how multiprotein complexes form. 
 
Title X-ray structure of a quaternary protein complex bound to DNA 
Description The atomic structure of a key quaternary protein complex bound to DNA was solved by X-ray crystallography 
Type Of Material Biological samples 
Provided To Others? No  
Impact Better understanding of some of the molecular mechanisms involved in the transcriptional regulation of haematopoiesis and leukaemogenesis. These observations will apply to other tissue-specific transcriptional complexes. 
 
Title Proteomics in Flk1+ cells 
Description Proteomics database of SCL partners in blood-fated mesodermal cells. 
Type Of Material Database/Collection of data 
Provided To Others? No  
Impact This is contributing to our understanding of the molecular mechanisms underlying lineage specification. More specifically, how a tissue-specific regulator activates a lineage-specific gene expression programme and represses alternative fates in the same cell. This is an important concept likely to apply to other cellular systems. 
 
Title Whole genome sequencing databases (Flk1+ cells) 
Description RNA-seq, ChIP-seq, ATAC-seq databases from blood-fated mesodermal cells 
Type Of Material Database/Collection of data 
Year Produced 2018 
Provided To Others? Yes  
Impact This has provided new insight into the molecular mechanisms underlying lineage specification and highlighted the functional importance of repression mechanisms. More specifically, how a tissue-specific regulator activates a lineage-specific gene expression programme and represses alternative fates in the same cell. This is an important concept likely to apply to other cellular systems. 
URL https://www.nature.com/articles/s41467-018-07787-6
 
Description Crystal structure 
Organisation University of Oxford
Department Wellcome Trust Centre for Human Genetics
Country United Kingdom 
Sector Charity/Non Profit 
PI Contribution We have provided them with a very interesting protein complex to study by X-ray crystallography.
Collaborator Contribution They have solved the crystal structure of the transcription factor SCL and its partners. We are now further studying this structure in functional assays.
Impact PMID: 21076045 PMID: 21045296 PMID: 23831025 The structure has provided us with a structural explanation for how this complex operates. This has important implications for our understanding of the transcriptional regulation of haematopoiesis and leukaemogenesis.
Start Year 2007
 
Description Function of ETO2 in haematopoietic development 
Organisation Medical Research Council (MRC)
Department MRC Molecular Haematology Unit
Country United Kingdom 
Sector Academic/University 
PI Contribution My PhD student collaborated with the partner lab to develop a new project based on findings that stemmed from previous studies in our lab.
Collaborator Contribution The collaborative lab provided us with expertise, advice and reagents to develop this new project
Impact PMID: 23318133
Start Year 2007
 
Description GATA1 transcriptional regulation 
Organisation Medical Research Council (MRC)
Department MRC Molecular Haematology Unit
Country United Kingdom 
Sector Academic/University 
PI Contribution Expertise in technologies
Collaborator Contribution Expertise in specific technologies
Impact PMID: 20154211 PMID: 18625887 PMID: 16551635
 
Description HSC production from mouse and human ES cells 
Organisation Medical Research Council (MRC)
Department MRC Molecular Haematology Unit
Country United Kingdom 
Sector Academic/University 
PI Contribution Expertise in mouse and human ES cell cultures and differentiation
Collaborator Contribution Expertise in haematopoiesis during embryonic development to help design protocols aiming at reproducing blood development in vitro.
Impact We are currently characterising ES cell-derived haematopoiesis in order to be in a better position to produce blood stem cells in vitro. If successful, we will be closer to making blood stem cells from patient's own cells. This is a long-term project.
Start Year 2014
 
Description Mouse and human ES cell differentiation cultures with NIH 
Organisation National Institutes of Health (NIH)
Country United States 
Sector Public 
PI Contribution Expertise in mouse ES cells and developmental haematopoiesis
Collaborator Contribution Expertise in human ES cell differentiation cultures that complement our mouse ES cell expertise.
Impact Mouse ES cell differentiation protocol that recapitulates some of the milestones occurring during embryonic development of blood stem cells.
Start Year 2014
 
Description Normal human hematopoiesis 
Organisation University of Oxford
Country United Kingdom 
Sector Academic/University 
PI Contribution Intellectual
Collaborator Contribution Intellectual and experimental
Impact Better understanding of the hematopoietic hierarchy in humans.
Start Year 2015
 
Description Relationship between arterial and hematopoietic differentiation 
Organisation National Institutes of Health Clinical Center
Country United States 
Sector Hospitals 
PI Contribution Initiated the project. Expertise in ES cell differentiation, hematopoietic development.
Collaborator Contribution Expertise in vasculogenesis
Impact PMID: 27250641
Start Year 2012
 
Description Role of Gata1s in fetal megakaryopoiesis 
Organisation University of Oxford
Department Radcliffe Department of Medicine
Country United Kingdom 
Sector Academic/University 
PI Contribution Co-supervision of a post-doctoral scientist. Bringing expertise in Cellular and Molecular Biology.
Collaborator Contribution Expertise in megakaryopoiesis and function of GATA1 in hematopoiesis
Impact No outputs yet.
Start Year 2016
 
Description School visit 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Schools
Results and Impact 100 pupils attended the talks on biomedical careers. This has inspired pupils to study biology at University.

5th and 6th form pupils came to the lab for work experience
Year(s) Of Engagement Activity 2012
 
Description School visit (MCS) 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Schools
Results and Impact The talk inspired discussions and reflection on biomedical/science-related careers.

The talk motivated students to embark on scientific studies. Some came to my lab for work experience the following summer.
Year(s) Of Engagement Activity 2014
 
Description School visit (MCS) 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Schools
Results and Impact 50 pupils attended the career talk who engaged in discussions about science-related careers after the presentation.

6th form students came for a one week lab experience in Summer 2013
Year(s) Of Engagement Activity 2013
 
Description School visit (MCS) 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Schools
Results and Impact The talk inspired discussions and reflection on biomedical/science-related careers.

Following the talk, I had request from pupils wanting to do work experience in my lab. We hosted 4 of those in the summer.
Year(s) Of Engagement Activity 2015,2016,2017,2018,2019
 
Description WIMM Public lectures 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? Yes
Geographic Reach Local
Primary Audience Public/other audiences
Results and Impact Mixture of school pupils and people with an interest in science. All the talks were very well perceived and prompted discussions afterwards.

Below is the letter sent by the Head of Science of one of the schools whose pupils attended the lectures:


"Thank you for an excellent evening of lectures last Wednesday. Our boys were very pleased to be there. The talks were pitched at exactly their level and addressed a range of current topics that particularly interest them. Our discussions continued over a meal afterwards in a local restaurant. They had been inspired and stimulated beyond the confines of the A-level syllabus.

Thank you to all who made the evening so successful.

Yours sincerely",
Year(s) Of Engagement Activity 2011
 
Description Women in science meeting 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Other academic audiences (collaborators, peers etc.)
Results and Impact The talk stimulated discussions and increased interactions between women in our Institute

Organisation of specific workshops
Year(s) Of Engagement Activity 2014
 
Description Workshop (can we have it all?) 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Other academic audiences (collaborators, peers etc.)
Results and Impact Discussions on common issues experienced by women in science

Decisions made on actions to implement within our Institute to improve women's assertiveness
Year(s) Of Engagement Activity 2014
 
Description Young PI programme (how to manage people) 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Postgraduate students
Results and Impact Researchers came to me after the presentation to further discuss specific aspects of my talk.

Better of understanding of what it takes to manage people as a PI.
Year(s) Of Engagement Activity 2014