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Transcriptional networks controlling lymphocyte development

Objective

Acquired immunity to foreign pathogens depends on functional B and T cells. The objective of this proposal is to elucidate the transcriptional control of lymphocyte development at three stages by deciphering the transcriptional networks specifying pro-B and pro-T cells in early lymphopoiesis and plasma cells in terminal B cell differentiation.
To this end, we generated knock-in mice carrying a biotin acceptor sequence at the C-terminus of transcription factors, which can be biotinylated by transgenic co-expression of the E. coli biotin ligase BirA. In vivo biotinylation facilitates antibody-independent precipitation of these transcription factors by streptavidin pulldown (Bio-ChIP). Preliminary Bio-ChIP sequencing experiments validated this approach for genome-wide identification of transcription factor target genes.
Bio-ChIP sequencing will be used to identify the target genes of key transcription factors controlling the development of pro-B cells (Ikaros, E2A, STAT5, EBF1, Pax5, PU.1, IRF4), pro-T cells (Notch1, RBP-J, GATA3, Ikaros, E2A, STAT5) and plasma cells (Blimp1, IRF4, XBP1). RNA sequencing of wild-type and mutant lymphocytes will determine the regulated target genes of these factors, which are ultimately relevant for the elucidation of transcriptional networks. The function of selected target genes at central nodes of these networks will be analyzed by the latest miR30-shRNA knockdown technology. Finally, regulatory complexes interacting with these transcription factors will be identified by streptavidin-pulldown purification and mass spectrometry followed by their integration into the transcriptional networks by ChIP-seq mapping to the transcription factor target genes.
These experiments will provide fundamentally new molecular insight into the generation of all three lymphocyte stages and will contribute to a better understanding of how deregulation of the transcriptional control promotes the development of lymphoid malignancies.

Field of science

  • /natural sciences/chemical sciences/analytical chemistry/mass spectrometry

Call for proposal

ERC-2011-ADG_20110310
See other projects for this call

Funding Scheme

ERC-AG - ERC Advanced Grant

Host institution

FORSCHUNGSINSTITUT FUR MOLEKULARE PATHOLOGIE GESELLSCHAFT MBH
Address
Campus-vienna-biocenter 1
1030 Wien
Austria
Activity type
Private for-profit entities (excluding Higher or Secondary Education Establishments)
EU contribution
€ 2 499 305
Principal investigator
Meinrad Busslinger (Prof.)
Administrative Contact
Tanja Winkler (Ms.)

Beneficiaries (1)

FORSCHUNGSINSTITUT FUR MOLEKULARE PATHOLOGIE GESELLSCHAFT MBH
Austria
EU contribution
€ 2 499 305
Address
Campus-vienna-biocenter 1
1030 Wien
Activity type
Private for-profit entities (excluding Higher or Secondary Education Establishments)
Principal investigator
Meinrad Busslinger (Prof.)
Administrative Contact
Tanja Winkler (Ms.)