Unknown,Transcriptomics,Genomics,Proteomics

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Gfi1 regulates survival and lineage commitment of hematopoietic precursors and prevents myeloproliferative diseases

ABSTRACT: Hematopoietic stem cells (HSCs) and lymphoid-primed multi-potential progenitors (LMPPs) are able to initiate both lymphoid and myeloid differentiation. We show here that the transcriptional repressor Gfi1 (growth factor independence 1) implements a specific gene expression program in HSCs and LMPPs that is critical for their survival and lymphoid differentiation potential. We present evidence that Gfi1 is required to maintain expression of genes involved in lymphoid development such as Flt-3, IL7R, Ebf1, Rag1, CCR9 and Notch1 and controls myeloid lineage commitment by regulating expression of genes such as Hoxa9 or M-CSFR. Gfi1 also inhibits apoptosis in HSCs by repressing pro-apoptotic genes such as Bax or Bak. As a consequence, Gfi1-/- mice show defects in self renewal, survival and both myeloid and lymphoid development of HSCs and LMPPs. Co-expression of a Bcl-2 transgene can partially restore the function of HSCs in Gfi1-/- mice, but not the defects in early lymphoid development. Of interest, Gfi1-/- x Bcl-2 transgenic mice show an accelerated expansion of myeloid cells and succumb to a fatal myeloproliferative disease resembling chronic myelomonocytic leukemia (CMML). Our data show that Gfi1 protects HSCs against apoptosis, ensures the proper development of LMPPs and plays a role in the development of myeloid leukemia. We used microarrays to detail the global gene expression changes following knockout of Gfi1 in mouse LSK cells We compared LSK cells isolated from Gfi1 knockout mice with wildtype cells to determine global gene expression changes by microarray analysis

ORGANISM(S): Mus musculus

SUBMITTER: Ludger Klein-Hitpass

PROVIDER: E-GEOD-20282 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Growth factor independence 1 protects hematopoietic stem cells against apoptosis but also prevents the development of a myeloproliferative-like disease.

Khandanpour Cyrus C Kosan Christian C Gaudreau Marie-Claude MC Dührsen Ulrich U Hébert Josée J Zeng Hui H Möröy Tarik T

Stem cells (Dayton, Ohio) 20110201 2

The regulation of gene transcription is elementary for the function of hematopoietic stem cells (HSCs). The transcriptional repressor growth factor independence 1 (Gfi1) restricts HSC proliferation and is essential to maintain their self-renewal capacity and multipotency after transplantation. In addition, Gfi1(-/-) HSCs are severely compromised in their ability to compete with wild-type (wt) HSCs after transplantation. We now report that Gfi1 protects HSCs against stress-induced apoptosis, prob ...[more]

PMID: 21732494

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Project description:Growth factor independent 1 (Gfi1) is a transcriptional repressor originally identified as a common integration site in Moloney-murine-leukemia-virus-induced T-cell leukemia. Gfi1-/- mice display increased apoptosis of developing thymocytes and T lymphopenia; however, there are contradictory reports of the absolute number of Gfi1-/- early T lineage progenitors. We used floxed alleles of Gfi1 crossed to various T-cell-specific Cre transgenes to map the requirements for Gfi1 during lymphoid priming and development. We show that Gfi1 is necessary for the proper formation and function of both lymphoid-primed multipotent progenitors and early T lineage progenitors. These defects correlate with a global inability of Gfi1-/- progenitors to enforce the activation of lymphoid genes including IL7R, Rag1, Flt3 and Notch1. Forced expression of intracellular Notch1 fails to rescue the Gfi1-/- defective lymphoid gene signature or Gfi1-/- T cell development. Instead, activation of Notch1 in Gfi1-/- cells results in a potent synthetic lethal phenotype that is most dramatic in immature thymocytes, but absent in mature peripheral T cells where developmental transcriptional programs are silent. Moreover, we find that the requirement for Gfi1-transcriptional integration of Notch-driven lymphoid transcriptional programs is cell autonomous. Our data indicate that Gfi1 is required at multiple independent stages of lymphoid development. In hematopoietic progenitors Gfi1 is necessary to integrate Notch1 signaling, mediate lymphoid priming, the formation of early T lineage progenitors and subsequent T lineage commitment. Lineage negative cells were purified by magnetic beads from RosaCreERT2 Gfi1 ex4-5 floxed mice and an activated Notch1 signal was introduced using a GFP+ retroviral vector. GFP+ progenitors were FACS-sorted and cultured in semi-solid media for one week to allow sufficient time to to instruct lymphoid differentiation, then replated in 1uM 4-OHT or EtOH control. After an additional 7 days, CFU were disrupted and RNA was isolated for global gene expression using microarrays.

Project description:C/EBPalpha is a transcription factor critically involved in myeloid development and indispensable for formation of granulocytes. To track the cellular fate of stem and progenitor (LSK) cells, which express C/EBPalpha, we developed a mouse model expressing Cre recombinase from the Cebpa promoter and an inducible EYFP allele. We show that Cebpa/EYFP+ cells represent a significant subset of LSK cells, which predominantly give rise to myeloid cells in steady state hematopoiesis. C/EBPalpha induced a robust myeloid gene expression signature and downregulated E2A-induced regulators of early lymphoid development. In addition, Cebpa/EYFP+ cells comprise a fraction of early thymic progenitors (ETP) with robust myeloid potential. However, Cebpa/EYFP+ LSK and ETP cells retained the ability to develop into erythroid and T-lymphoid lineages, respectively. These findings support an instructive, but argue against a lineage restrictive role of C/EBPalpha in multipotent hematopoietic and thymic progenitors. We performed global gene expression profiling of double-sorted Cebpa/EYFP+ and Cebpa/EYFP- LSK cells of pooled Cebpa Cre/wt R26 EYFP reporter mice to identify differentially regulated genes in Cebpa+ versus Cebpa- LSK cells. RNA was isolated from three biological replicates of Cebpa/EYFP+ LSK cells and two biological replicates of Cebpa/EYFP- LSK cells. To determine if the identified genes were truly dependent on Cebpa expression, we also performed global gene expression profilling of Cebpa/EYFP+ and Cebpa/EYFP- fetal liver LSK cells of Cebpa Cre/fl R26 EYFP mice. Induction of Cebpa/Cre expression in these mice leads to Cre-mediated recombination of the floxed wt Cebpa allele resulting in a complete Cebpa knock-out. In this case, RNA was isolated from two biological replicates of either Cebpa/EYFP+ and Cebpa/EYFP- LSK cells. In addition, we included one biological replicate of Cebpa/EYFP+ and Cebpa/EYFP- fetal liver LSK cells of Cebpa Cre/wt R26 EYFP mice to determine the correlation of differentially regulated genes in bone marrow and fetal liver LSK cells.

Dataset Information

Gfi1 regulates survival and lineage commitment of hematopoietic precursors and prevents myeloproliferative diseases

Publications

Growth factor independence 1 protects hematopoietic stem cells against apoptosis but also prevents the development of a myeloproliferative-like disease.

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