transcription factorstranscription factors

By Najmaldin SakiSprin 2008

HOXB < HOXA

Stem cells

Bmi1Cdkn1a

PtenEtv6Mcl1

HoxB4

HSCs

Wnt pathway

Ang-1

Notch ligand

Gp130 Ligands

TNFAlpha

HSC expansion for clinical use

via gene therapy

HSC progenyGeneral hematopoiesis

Bre:brain and reproductive organ-expressed protein

HOXB4

TNFR1

Bre Inhibit (TNF Alpha ,Fas Ligand ,various other stress-related stimuli)

Fig. 3. Induction of HOXB4 activity protects cultivated HSCs from the negative effects TNF-{alpha} on stem cell self-renewal

4-hydroxytamoxifen(TMX)-inducibleForm of HoxB4(HoxB4)

Two member of the negative feedback loop of FGF signaling

Phosphorylated in response to FGF signaling

Chemical blockage of FGF signaling augmented the long-term repopulation activity of HOXB4-expressing HSCs/HPCs in vitro

HOXB4 Modulate Wnt and Notch signaling

Hbp1:transcriptional repressor of Wnt target genes

The core of the pathway is conserved from worms and flies to mammals.(Left) At the heart of the pathway, the destruction complex consisting of GSK-3,APC, and axin captures and phosphorylates -catenin.The latter is then recognized by TrCP and ubiquitinated by an associated E3 ligase complex. Subsequently,-catenin is degraded by the proteasome. Meanwhile, TCF/LEF is bound by corepressors such as Groucho (Grg) and represses target gene transcription.(Right) When Wnt factors ligate to their cell surface receptors, the kinase activity of the destruction complex is inhibited by dishevelled (Dsh).As a consequence,-catenin accumulates and travels to the nucleus where it binds to TCF/LEF transcription factors. The resulting complex potently activates transcription of target genes.

HOXB4

HOXB4:Wnt(Narf,Nlk) & Notch(Nrarp)

Mutation or substitution of a HOX gene can lead to the replacement of one body part by an other, reflecting the key significance of these genes in determining cellular identity.

Retrovirally driven HOXB4 expression in transduced adult HSC poulations leads to a 1000-fold net increase in growth,without any apparent reduction in the ability of these cells to subsequently differentiate into lymphoid and myloid lineages,and without the cells becoming transformed.

How dose HOXB4 promote HSC proliferation?

Positive regulation by HOX proteins requires the binding of a number of co-factors , most notably members of the PBX and MEIS transcription factor families.

Convert chromatin to an inative state

The activation of transcription may require additional factors to bind to

other genes enhancer,and these factors may only be available in

specific cells and tissues

The activation of transcription may require additional factors to bind to

other genes enhancer,and these factors may only be available in

specific cells and tissues

C-myc

NotchIn addition to c-myc two components of the AP-1 Transcriptional activation complex , Jun-B & Fra-1 are also upregulated by HOXB4

HOXB4 is not the first example of a transcription factor that can both activate and repress apoptosis. One of the best characterised is the Rel/NF-ĸB transcription factor, which has both anti-and proapoptopic roles in T-cells.

Rel/NF-ĸB HOXB4

Activating proapoptopic genesActivating proapoptopic genes

Activation antiapoptopic genesActivation antiapoptopic genes

Actively promotes proliferation

SMAD 2/3

PSMAD 2/3

SMAD 4

SMAD 4

PSMAD 2/3

TGF-b

FOXOSMAD 4

PSMAD 2/3

FOXO

FOXO

PAKT

IKKb

SMAD 4

PSMAD 2/3

SMAD 4

PSMAD 2/3

P21/cipBIMFasL

PAI-1TGIFTIEG

Growth arrest Apoptosis

-

TIEG

Smad2 Smad7

Growth factor cytokines

Nucleus

STAT-5

PIM2

BAD PBAD

SurvivalApoptosis

-

Survival and apoptotic signaling pathways

DUSP6

(survival)

PIP3

PDK1

PI3K PTENP38 MAPK

CM

EM

EM

CM

EM

CM

SMAD 2/3

PSMAD 2/3

SMAD 4

SMAD 4

PSMAD 2/3

TGF-b

SMAD 4

PSMAD 2/3

SMAD 4

PSMAD 2/3

PAI-1TGIFTIEG

Growth arrest Apoptosis

-

TIEG

Smad2 Smad7

Nucleus

-

EM

TGF-beta pathway

EMGene Array

The transcription factor PU.1 is an essential regulator of haemopoiesis and a suppressor of myeloid leukaemia.Enforced expression of PU.1 in haemopoietic progenitors has suggested an instructive role of PU.1 in promoting macrophage an ddendriticcell (DC) development. PU.1 performs these important roles by regulating numerous genes within the myeloid and lymphoid lineages, including those encoding many ofthe developmentally important cytokine receptors, for example macrophage colony-stimulating factor receptor (M-CSFR) ,granulocyte–macrophage colony-stimulating factor receptor (GM-CSFR)Alpha and interleukin (IL)-7RAlpha, and by interacting with several other key transcription factors, including interferon regulatory factor (IRF)4, IRF8, acute myeloid leukaemia (AML)-1,CCAAT/enhancer-binding protein (C/EBP)a,GATA-binding protein-1 (GATA-1) and c-Jun. However, the lethality andlack oflong-term repopulatingstem cell activity associated with germline PU.1-deficiency has, until recently,precluded afullanalysis of PU.1function in adult haemopoiesis.Subsequently, overexpression studies demonstrated that continuous expression of high PU.1 levels, while permissive for macrophage development, could partially block erythroid differentiation and completely block early phases of B cell and Tcell development. recent study has even demonstrated an ability of PU.1 to reprogramme committed T cell progenitors into dendritic-like cells.

Central role of PU.1 in leukaemogenesis. Schematic of the human and mouse leukaemias in which the modulation of PU.1 function is thought to be important. Red arrows indicate the relative PU.1 activity compared with non-malignant haemopoietic progenitors. Abbreviations: DBD, PU.1 DNA-binding domain.

Pax5 and commitment to the

B cell lineage

• E2A and EBF are needed to turn on B cell specific genes including Pax5, which turns on additional B cell-specific genes

Putting it all together: growth factors + transcription factors in B cell

development

Figure 2. B-cell development and the roles of EBF. (a) Progressive stages of B-cell lymphopoiesis are shown with (i) cell designations indicated below, (ii) the status of V(D)J

recombination indicated within each cell type and (iii) characteristic cell surface markers indicated above each cell. B-biased progenitors represent the CLP-2 stage in other

models of development [1]. The approximate points at which B-cell lymphopoiesis is arrested in PU.1K/K, E2AK/K, EBFK/K and Pax5K/K mice are indicated above the cells.

Cells lacking EBF or E2A closely resemble B-biased progenitor cells but lack all Ig gene rearrangements. (b) Central role of EBF in B-cell development. Regulatory circuits in

developing B cells are shown to highlight the functions of EBF. Transcription of the ebf1 gene is a function of the transcription factors PU.1 [19] and E2A [32] and is reinforced

by IL-7R signaling [35,36]. EBF might autoregulate its own expression [18]. EBF regulates genes, including pax5 [17], RAG-1 [15], l5 and VpreB1 [15], B29 [51], B-lymphoid

tyrosine kinase (blk) [52] and CD19 [53]. Many EBF targets have been confirmed as targets of E2A, which functions in synergy with EBF [33]. A role for EBF in the regulation of

E2A expression in the B-cell compartment (broken line) is suggested by the reduction of E2A in the absence of EBF [15]. Pax5, in turn, regulates mb-1 and CD19 synergistically

with EBF [40]. Pax5 regulates many other genes required for the B-lineage program and commitment (reviewed in Ref. [1]). Abbreviations: LinK, lineage-specific markers.

A Schematic Diagram of Regulatory Network Controlling Plasma Cell Differentiation Our results suggest the diminishing of Bcl-6 as a primary event in the development of plasmacytic phenotype in Pax5-deficient DT40 cells, since Pax5 is not essential to the inhibition of Blimp-1. Thus, Pax5 is more likely needed to maintain Bcl-6 level (dashed arrow) at later stages of B cell development. The solid lines indicate the previously known inhibitory signals of this regulatory network.

Functional roles and target genes of E2A, EBF and Pax5. E2A and EBF cooperate in the specification of the B cell fate. These transcription factors regulate the expression of immunoglobulin surrogate light chain genes λ5, VpreB, the recombinase activating genes Rag1, and the mb1, B29 genes, which encode components of the pre-BCR and BCR. Pax5, which is under the control of E2A and EBF, regulates the commitment step of B cell differentiation, in which the expression of signaling receptors that determine alternative lineage choices (Notch, M-CSF-R) are repressed. Pax5 activates the expression of the B cell determinants mb-1, CD19 and BLNK/SLP65/BASH (B cell linker protein) as well as the recombination of distal variable gene segments.

Lineage commitment in the bone marrow. The position of the transcription factors on the scheme represents the stage at which their absence (based usually on knockout studies) leads to a developmental block. Proteins normally acting as inhibitors of the differentiation pathways are depicted in red. Physical interactions between transcription factors are indicated by double-pointed blue arrows. SC, stem cell; HSC, hematopoietic stem cell; CLP, common lymphoid progenitor; NK, natural killer cell.

Transcription factors in plasma cell differentiation. At the GC B cell stage, BCL-6 represses Blimp-1, a key regulator of plasma cell differentiation. Another transcription factor crucial in plasma cells, XBP-1 is repressed by Pax-5. Upon transition to the plasma cell stage, Blimp-1 expression leadstothe repression of BCL-6 transcription, and thereby to the inhibition of earlier GC B cell activities, including the transcription of a number of B cell specific transcription factors, such as Pax-5. As a consequence of Pax-5 downregulation, XBP-1, specifically required for plasma cell development, is expressed.

Structure of the vertebrate family of GATA proteins. All 6 vertebrate GATA factors share a conserved DNA-binding domain consisting of 2 zinc fingers (ZnF), a feature that defines this family of transcription factors. The different GATA factors can be divided into 2 subgroups based on amino acid sequence homology and tissue distribution: the hematopoietic subgroup (GATA 1/2/3) and the cardiac subgroup (GATA 4/5/6). Transac-tivation domains are found in either the N-terminal (N-term) and/or C-terminal (C-term) portions of the different GATA proteins. NLS, nuclear localization signal.

Proposed model for GATA factors as effectors of hormonal signaling in testicular cells. In the testis, Sertoli and Leydig cell gene expression and function are tightly regulated by the pituitary trophic hormones follicle-stimulating hormone (FSH) and leutinizing hormone (LH), respectively. Hormone binding to G-protein coupled cell surface receptors activates adenylate cyclase (AC), leading to increased intracellular cAMP levels. cAMP then binds the regulatory (R) subunit of protein kinase A (PKA), allowing dissociation of the PKA catalytic (C) subunit and its translocation to the nucleus, where it phosphorylates target proteins. In both Sertoli and Leydig cells, GATA factors are novel targets for PKA-mediated phosphorylation. As shown, phosphorylation of GATA4 at serine 261 allows for an enhanced cooperation with multiple transcriptional partners and the recruitment of the CBP coactivator. The end result is increased expression of hormonally regulated GATA-dependent target genes such as steroidogenic acute regulatory protein (Star), inhibin (Inha), and P450 aromatase (Cyp19� ).

Model for the dynamic regulation of GATA1 during haematopoiesis. The relative expression levels of the mouse GATA1 gene are indicated by various colors as shown in the box. Red arrows indicate the stages of differentiation which are inhibited when the expression levels of GATA1 are genetically altered in mice. Note that the expression levels in the testis, eosinophil and mast cell are imprecise. CLP, common lymphoid progenitor; CMP, common myeloid progenitor; ES, embryonic stem; HSC, haematopoietic stem cell; MEP, megakaryocyte/erythrocyte lineage-restricted progenitor; ProE, proerythroblast.

Model of Th17 lineage development. The differentiation of Th17 cells is initiated by the activation of naı¨ve T cells in the presence of IL-6 plus

TGF-b. This leads to the expression of ROR-gt and production of IL-17. IL-6, produced by the innate immune system, is crucial during this phase but when this cytokine is not present and T-reg cells are eliminated, IL-21 produced by NK cells and NK T cells together with TGF-b can initiate an alternate pathway of Th17 differentiation. Upon differentiation, IL-21 is also massively induced by the developing Th17 cells and acts in

autocrine fashion on Th17 cells to amplify this population. Then, IL-23 stabilizes previously differentiated Th17 cells and enables further

expansion of the Th17 lineage with sustained production of its hallmark cytokines.