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Loss of Ikaros DNA-binding function confers integrin-dependent survival on pre-B cells and progression to acute lymphoblastic leukemia.

Joshi I, Yoshida T, Jena N, Qi X, Zhang J, Van Etten RA, Georgopoulos K - Nat. Immunol. (2014)

Bottom Line: Deletion of the DNA-binding domain of the transcription factor Ikaros generates dominant-negative isoforms that interfere with its activity and correlate with poor prognosis in human precursor B cell acute lymphoblastic leukemia (B-ALL).Here we found that conditional inactivation of the Ikaros DNA-binding domain in early pre-B cells arrested their differentiation at a stage at which integrin-dependent adhesion to niches augmented signaling via mitogen-activated protein kinases, proliferation and self-renewal and attenuated signaling via the pre-B cell signaling complex (pre-BCR) and the differentiation of pre-B cells.Our results explain how normal pre-B cells transit from a highly proliferative and stroma-dependent phase to a stroma-independent phase during which differentiation is enabled, and suggest potential therapeutic strategies for Ikaros-mutant B-ALL.

View Article: PubMed Central - PubMed

Affiliation: Cutaneous Biology Research Center, Massachusetts General Hospital, Harvard Medical School, Charlestown, Massachusetts, USA.

ABSTRACT
Deletion of the DNA-binding domain of the transcription factor Ikaros generates dominant-negative isoforms that interfere with its activity and correlate with poor prognosis in human precursor B cell acute lymphoblastic leukemia (B-ALL). Here we found that conditional inactivation of the Ikaros DNA-binding domain in early pre-B cells arrested their differentiation at a stage at which integrin-dependent adhesion to niches augmented signaling via mitogen-activated protein kinases, proliferation and self-renewal and attenuated signaling via the pre-B cell signaling complex (pre-BCR) and the differentiation of pre-B cells. Transplantation of polyclonal Ikaros-mutant pre-B cells resulted in long-latency oligoclonal pre-B-ALL, which demonstrates that loss of Ikaros contributes to multistep B cell leukemogenesis. Our results explain how normal pre-B cells transit from a highly proliferative and stroma-dependent phase to a stroma-independent phase during which differentiation is enabled, and suggest potential therapeutic strategies for Ikaros-mutant B-ALL.

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Signaling pathways in WT and Ikaros-deficient pre-B cellsa-b, Immunoblot analysis of proliferation and survival (a) and differentiation (b) signaling pathways activated by IL-7R and pre-BCR is shown. Representative expression and activity of pre-BCR-affiliated PTKs and downstream differentiation-inducing signaling effectors, as described in Supplementary Fig. 4a, are shown from two WT and three IkE5Δ/Δ independent stromal cultures of primary cells after limited in vitro propagation. β-tubulin, T-Btk or T-p38 serve as loading controls for WT and IkE5Δ/Δ pre-B cells and non-adherent WT pre-B cells. c, Intracellular Ca2+ levels (Fura Red, left panel) at steady state and Ca2+ flux (Green/Fura Red, right panel) measured after anti-IgM-stimulation of WT and IkE5Δ/Δ adherent and non-adherent pre-B cells. Fura Red staining and MFI shown on the left site inversely correlates with Ca2+ levels. Data are representative of two independent WT and mutant pre-B cell cultures.
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Figure 4: Signaling pathways in WT and Ikaros-deficient pre-B cellsa-b, Immunoblot analysis of proliferation and survival (a) and differentiation (b) signaling pathways activated by IL-7R and pre-BCR is shown. Representative expression and activity of pre-BCR-affiliated PTKs and downstream differentiation-inducing signaling effectors, as described in Supplementary Fig. 4a, are shown from two WT and three IkE5Δ/Δ independent stromal cultures of primary cells after limited in vitro propagation. β-tubulin, T-Btk or T-p38 serve as loading controls for WT and IkE5Δ/Δ pre-B cells and non-adherent WT pre-B cells. c, Intracellular Ca2+ levels (Fura Red, left panel) at steady state and Ca2+ flux (Green/Fura Red, right panel) measured after anti-IgM-stimulation of WT and IkE5Δ/Δ adherent and non-adherent pre-B cells. Fura Red staining and MFI shown on the left site inversely correlates with Ca2+ levels. Data are representative of two independent WT and mutant pre-B cell cultures.

Mentions: The survival and proliferative expansion of pre-B cells are supported by a combination of pre-BCR and IL-7R signaling that activates the PI3K-Akt and Erk1-2 MAPK pathways (Supplementary Fig 4a and Fig. 4a, b). Both PI3K-Akt and Erk1-2 were active in WT adherent but not in non-adherent pre-B cells (Fig. 4a), which are in the process of exiting the cell cycle (Supplementary Fig. 3b) and upregulating expression of small pre-B cell markers (Fig. 3c, d). Activation of Akt was similar in IkE5Δ/Δ adherent pre-B cells compared to WT, but activation of Erk1 and Erk2 was greatly increased (Fig. 4a). Consistent with a higher Erk1-2 MAPK activity, an increase in Cyclin D2 (Fig. 4a) and cell cycle (Fig. 2d, e) was observed in IkE5Δ/Δ compared to WT large pre-B cells.


Loss of Ikaros DNA-binding function confers integrin-dependent survival on pre-B cells and progression to acute lymphoblastic leukemia.

Joshi I, Yoshida T, Jena N, Qi X, Zhang J, Van Etten RA, Georgopoulos K - Nat. Immunol. (2014)

Signaling pathways in WT and Ikaros-deficient pre-B cellsa-b, Immunoblot analysis of proliferation and survival (a) and differentiation (b) signaling pathways activated by IL-7R and pre-BCR is shown. Representative expression and activity of pre-BCR-affiliated PTKs and downstream differentiation-inducing signaling effectors, as described in Supplementary Fig. 4a, are shown from two WT and three IkE5Δ/Δ independent stromal cultures of primary cells after limited in vitro propagation. β-tubulin, T-Btk or T-p38 serve as loading controls for WT and IkE5Δ/Δ pre-B cells and non-adherent WT pre-B cells. c, Intracellular Ca2+ levels (Fura Red, left panel) at steady state and Ca2+ flux (Green/Fura Red, right panel) measured after anti-IgM-stimulation of WT and IkE5Δ/Δ adherent and non-adherent pre-B cells. Fura Red staining and MFI shown on the left site inversely correlates with Ca2+ levels. Data are representative of two independent WT and mutant pre-B cell cultures.
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Related In: Results  -  Collection

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Figure 4: Signaling pathways in WT and Ikaros-deficient pre-B cellsa-b, Immunoblot analysis of proliferation and survival (a) and differentiation (b) signaling pathways activated by IL-7R and pre-BCR is shown. Representative expression and activity of pre-BCR-affiliated PTKs and downstream differentiation-inducing signaling effectors, as described in Supplementary Fig. 4a, are shown from two WT and three IkE5Δ/Δ independent stromal cultures of primary cells after limited in vitro propagation. β-tubulin, T-Btk or T-p38 serve as loading controls for WT and IkE5Δ/Δ pre-B cells and non-adherent WT pre-B cells. c, Intracellular Ca2+ levels (Fura Red, left panel) at steady state and Ca2+ flux (Green/Fura Red, right panel) measured after anti-IgM-stimulation of WT and IkE5Δ/Δ adherent and non-adherent pre-B cells. Fura Red staining and MFI shown on the left site inversely correlates with Ca2+ levels. Data are representative of two independent WT and mutant pre-B cell cultures.
Mentions: The survival and proliferative expansion of pre-B cells are supported by a combination of pre-BCR and IL-7R signaling that activates the PI3K-Akt and Erk1-2 MAPK pathways (Supplementary Fig 4a and Fig. 4a, b). Both PI3K-Akt and Erk1-2 were active in WT adherent but not in non-adherent pre-B cells (Fig. 4a), which are in the process of exiting the cell cycle (Supplementary Fig. 3b) and upregulating expression of small pre-B cell markers (Fig. 3c, d). Activation of Akt was similar in IkE5Δ/Δ adherent pre-B cells compared to WT, but activation of Erk1 and Erk2 was greatly increased (Fig. 4a). Consistent with a higher Erk1-2 MAPK activity, an increase in Cyclin D2 (Fig. 4a) and cell cycle (Fig. 2d, e) was observed in IkE5Δ/Δ compared to WT large pre-B cells.

Bottom Line: Deletion of the DNA-binding domain of the transcription factor Ikaros generates dominant-negative isoforms that interfere with its activity and correlate with poor prognosis in human precursor B cell acute lymphoblastic leukemia (B-ALL).Here we found that conditional inactivation of the Ikaros DNA-binding domain in early pre-B cells arrested their differentiation at a stage at which integrin-dependent adhesion to niches augmented signaling via mitogen-activated protein kinases, proliferation and self-renewal and attenuated signaling via the pre-B cell signaling complex (pre-BCR) and the differentiation of pre-B cells.Our results explain how normal pre-B cells transit from a highly proliferative and stroma-dependent phase to a stroma-independent phase during which differentiation is enabled, and suggest potential therapeutic strategies for Ikaros-mutant B-ALL.

View Article: PubMed Central - PubMed

Affiliation: Cutaneous Biology Research Center, Massachusetts General Hospital, Harvard Medical School, Charlestown, Massachusetts, USA.

ABSTRACT
Deletion of the DNA-binding domain of the transcription factor Ikaros generates dominant-negative isoforms that interfere with its activity and correlate with poor prognosis in human precursor B cell acute lymphoblastic leukemia (B-ALL). Here we found that conditional inactivation of the Ikaros DNA-binding domain in early pre-B cells arrested their differentiation at a stage at which integrin-dependent adhesion to niches augmented signaling via mitogen-activated protein kinases, proliferation and self-renewal and attenuated signaling via the pre-B cell signaling complex (pre-BCR) and the differentiation of pre-B cells. Transplantation of polyclonal Ikaros-mutant pre-B cells resulted in long-latency oligoclonal pre-B-ALL, which demonstrates that loss of Ikaros contributes to multistep B cell leukemogenesis. Our results explain how normal pre-B cells transit from a highly proliferative and stroma-dependent phase to a stroma-independent phase during which differentiation is enabled, and suggest potential therapeutic strategies for Ikaros-mutant B-ALL.

Show MeSH
Related in: MedlinePlus