Determinants of hematopoietic stem cell / niche interactions in lineage commitment – HSC-FATE

The identification of the mechanisms that control stem cell self renewal and cell fate decisions are questions of critical importance for basic biology and therapeutic applications. Hematopoietic stem cells (HSC) are probably the best defined class of multi-potent adult stem cells. The defining characteristics of HSC are their ability to self-renew throughout life of the organism and to give rise to all lineages of mature blood cells. Whereas self-renewal of HSC has been extensively studied, little is known about the control mechanisms of assymetric HSC divisions that assure the balanced production of different lineage specific progeny. - Recently the major importance of the bone marrow microenvironment for HSC biology has become evident, but so far it has been widely assumed that niche occupation is mainly required for self-renewal, whereas differentiation is believed to simply imply departure from this environment. In this proposal we now want to explore, whether specific microenvironments can support differentiation of HSC along different lineages and whether differential cell adhesion and sorting events between distinct micro-environments enable asymmetric HSC divisions that assure lineage commitment. To limit our investigation to a well-defined process, we will focus on the regulation of myeloid HSC commitment by the cytokine M-CSF and the monocytic transcription factor MafB that we could recently demonstrate. - We observed that MafB is expressed at significant levels in HSC and selectively restricts HSC divisions in response to M-CSF. Both in vivo and in culture MafB-/- HSC show a strongly accelerated cell division in response to M-CSF but not to other myeloid cytokines or under T-cell and erythroid differentiation conditions. Consistent with an increased myeloid potential of MafB-/- HSC, the myeloid transcription factor PU.1 expression was strongly up-regulated. Furthermore, competitive transplantations revealed normal contribution of MafB-/- HSC to the T-cell or erythroid compartment but a strong capacity to replace competitor cells in the myelo-monocytic lineage. This lineage biased repopulation advantage was progressive, maintained long term, enhanced through serial transplantation and could be initiated by the statistical equivalent of a single stem cell in limiting dilutions. Together this indicated that MafB selectively restricts the intrinsic potential of HSC to generate myeloid progeny in response to M-CSF without compromising self-renewal capacity. - Several observations indicate that interactions with the micro-environment may be important for this phenotype. MafB -/- deficient cells show altered actin modulation in response to M-CSF and increased expression of the adhesion molecule E-Cadherin as well as components of the complex that anchors it to the actin skeleton. In other experimental models MafB mutant phenotypes often involve a disturbed balance between N-Cadherin and E-Cadherin adhesion molecules. In addition, N-Cadherin is a key molecules to anchor HSC to the self renewal niche and components of the complex linking E-Cadherin to actin are important for asymmetric stem cell divisions. - In our analysis of HSC/stromal cell interactions we will therefore focus on Cadherin adhesion molecules and their connection to the actin skeleton. We will analyze the molecular parameters that are cell intrinsic such as the MafB controlled HSC sensitivity to M-CSF signaling as well as those dependent on the micro-environment, such as the balance of cadherin adhesion molecules. We will address several precise questions with exemplary importance for HSC biology in general: - 1. Does the increased sensitivity to M-CSF and myeloid commitment of MafB-/- HSC depend on preferential interaction with M-CSF expressing bone marrow stromal cells? - 2. Does myeloid lineage commitment of HSC involve a switch from N-cadherin to E-Cadherin dependent stromal cell interactions? - 3. Are changes in the actin skeleton and/or members of the prot...