Regulation of oncogenic gene expression by transcription factors that function as tumor suppressors is one of the major mechanisms that regulate leukemogenesis. Understanding this complex process is essential for explaining the pathogenesis of leukemia as well as developing targeted therapies. Here, we provide an overview of the role of Ikaros tumor suppressor and its role in regulation of gene transcription in acute leukemia. Ikaros (IKZF1) is a DNA-binding protein that functions as a master regulator of hematopoiesis and the immune system, as well as a tumor suppressor in acute lymphoblastic leukemia (ALL). Genetic alteration or functional inactivation of Ikaros results in the development of high-risk leukemia. Ikaros binds to the specific consensus binding motif at upstream regulatory elements of its target genes, recruits chromatin-remodeling complexes and activates or represses transcription via chromatin remodeling. Over the last twenty years, a large number of Ikaros target genes have been identified, and the role of Ikaros in the regulation of their expression provided insight into the mechanisms of Ikaros tumor suppressor function in leukemia. Here we summarize the role of Ikaros in the regulation of the expression of the genes whose function is critical for cellular proliferation, development, and progression of acute lymphoblastic leukemia.

IKAROS, encoded by the IKZF1 gene, is a DNA-binding protein that functions as a tumor suppressor in T cell acute lymphoblastic leukemia (T-ALL). Recent studies have identified IKAROS’s novel function in the epigenetic regulation of gene expression in T-ALL and uncovered many genes that are likely to be directly regulated by IKAROS. Here, we report the transcriptional regulation of two genes, phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit delta (PIK3CD) and phosphoinositide kinase, FYVE-type zinc finger containing (PIKFYVE), by IKAROS in T-ALL. PIK3CD encodes the protein p110δ subunit of phosphoinositide 3-kinase (PI3K). The PI3K/AKT pathway is frequently dysregulated in cancers, including T-ALL. IKAROS binds to the promoter regions of PIK3CD and PIKFYVE and reduces their transcription in primary T-ALL. Functional analysis demonstrates that IKAROS functions as a transcriptional repressor of both PIK3CD and PIKFYVE. Protein kinase CK2 (CK2) is a pro-oncogenic kinase that is overexpressed in T-ALL. CK2 phosphorylates IKAROS, impairs IKAROS’s DNA-binding ability, and functions as a repressor of PIK3CD and PIKFYVE. CK2 inhibition results in increased IKAROS binding to the promoters of PIK3CD and PIKFYVE and the transcriptional repression of both these genes. Overall, the presented data demonstrate for the first time that in T-ALL, CK2 hyperactivity contributes to PI3K signaling pathway upregulation, at least in part, through impaired IKAROS transcriptional regulation of PIK3CD and PIKFYVE. Targeting CK2 restores IKAROS’s regulatory effects on the PI3K oncogenic signaling pathway.

Ikzf1 encodes a zinc finger, DNA-binding protein that functions as a tumor suppressor in acute lymphoblastic leukemia (ALL). Deletion and/or loss of Ikaros function results in the development of high-risk leukemia. In the nucleus, Ikaros forms complexes with histone deacetylase complex, NuRD, and it participates in the formation of heterochromatin. The role of Ikaros-mediated formation of heterochromatin in tumor suppression in leukemia is unknown. We determined global genomic occupancy of Ikaros, global heterochromatin distribution, chromatin accessibility, DNA methylation landscape, and gene expression in primary human T-cell ALL (T-ALL), as well as in mouse T-ALL to analyze how Ikaros regulates heterochromatin landscape and gene expression in T-ALL. Results showed that Ikaros DNA occupancy is essential for the recruitment of histone deacetylase 1 (HDAC1), Polycomb repressive complex 2 (PRC2) and formation of facultative heterochromatin, as well as the formation of constitutive heterochromatin (characterized by H3K9me3 occupancy). T-ALL cells with deletion of both Ikzf1 alleles have severely impaired HDAC1 DNA occupancy and reduced H3K27me3. Re-introduction of Ikzf1 via retroviral transduction resulted in the restoration of H3K27me3 facultative heterochromatin, along with HDAC1 DNA occupancy. The H3K27me3 genomic distribution following Ikzf1 re-introduction showed high homology to the H3K27me3 genomic distribution in normal thymocytes. Analysis of H3K9me3 genomic distribution showed that Ikzf1 deletion results in dramatic redistribution of H3K9me3 global occupancy, with reduced H3K9me3 occupancy at pericentromeric loci. Reintroduction of Ikzf1 enhances H3K9me3 enrichment in pericentromeric loci, as well as at the promoters of genes that are involves in cellular proliferation. Analysis of DNA methylation distribution showed that Ikzf1 expression regulates global DNA methylation landscape. The presence of facultative heterochromatin, with enrichment of H3K27me3, inversely correlated with DNA methylation. Global analysis of chromatin accessibility revealed that Ikaros binding resulted in the loss of chromatin accessibility at over 3400 previously-accessible chromatin sites. Dynamic analyses demonstrate the long-lasting effects of Ikaros's DNA binding on heterochromatin distribution and chromatin accessibility. Analysis of gene expression in T-ALL with both Ikzf1 alleles and in Ikzf1-defficient cells (from Ikzf1-defficient T-ALL, and from Ikzf1-wild-type T-ALL following Ikzf1 deletion by CRISPR) showed that Ikaros-induced redistribution of facultative and constitutive heterochromatin results in the repression of several genes that are critical for cell cycle progression, PI3K-AKT-mTOR, and WNT signaling pathway. In conclusion, results suggest that Ikaros' tumor suppressor function in T-ALL occurs via global regulation of the heterochromatin, DNA methylation landscape, and chromatin accessibility, as well as via epigenetic regulation of transcription of the genes that play essential roles in signaling pathways that promote cellular proliferation.