B-lymphocyte development in the bone marrow is controlled by the coordinated action of transcription factors creating regulatory networks ensuring activation of the B-lymphoid program and silencing of alternative cell fates. This process is tightly connected to malignant transformation because B-lineage acute lymphoblastic leukemia cells display a pronounced block in differentiation resulting in the expansion of immature progenitor cells. Over the last few years, high-resolution analysis of genetic changes in leukemia has revealed that several key regulators of normal B-cell development, including IKZF1, TCF3, EBF1, and PAX5, are genetically altered in a large portion of the human B-lineage acute leukemias. This opens the possibility of directly linking the disrupted development as well as aberrant gene expression patterns in leukemic cells to molecular functions of defined transcription factors in normal cell differentiation. This review article focuses on the roles of transcription factors in early B-cell development and their involvement in the formation of human leukemia.

cOne of the first lines of defense against infection is the activation of the innate immune system. It is becoming clear that autoimmune diseases, such as rheumatoid arthritis and Crohn's disease, may be caused by disturbed innate immunity, and relating granulocyte and monocyte functions to the patient genotype has become an important part of contemporary research. Although it is essential to move this field forward, a systematic study comparing the efficacy and suitability for functional studies of the various available protocols for the isolation of these immune cells has not been performed. Here, we compare human granulocyte functionality under three enrichment protocols: (i) Ficoll density gradient centrifugation, (ii) anti-CD15 antibody-conjugated microbeads (positive selection), and (iii) Polymorphoprep. Primary monocytes were isolated in parallel using (i) anti-CD14 magnetic microbeads, (ii) non-monocyte depletion by antibody-conjugated magnetic microbeads (negative selection), (iii) RosetteSep antibody cocktail, and (iv) the classical adherence protocol. The best results in terms of purity and cell functionality were obtained with positive selection by magnetic microbeads for both human granulocytes and monocytes. Whereas phagocytosis of Escherichia coli bacteria was identical in all isolation procedures tested, the granulocyte respiratory burst was higher in positively selected cells. In addition, different granulocyte enrichment procedures affect cell surface receptor expression to different extents. In toto, we propose that positive selection of granulocytes and monocytes be adopted as the procedure of choice for studies of human granulocyte and monocyte functions but caution investigators to be aware of possible alterations in cell phenotypes with different isolation procedures.

IKAROS is required for the differentiation of highly proliferative pre-B-cell precursors, and loss of IKAROS function indicates poor prognosis in precursor B-cell acute lymphoblastic leukemia (B-ALL). Here we show that IKAROS regulates this developmental stage by positive and negative regulation of superenhancers with distinct lineage affiliations. IKAROS defines superenhancers at pre-B-cell differentiation genes together with B-cell master regulators such as PAX5, EBF1, and IRF4 but is required for a highly permissive chromatin environment, a function that cannot be compensated for by the other transcription factors. IKAROS is also highly enriched at inactive enhancers of genes normally expressed in stem-epithelial cells. Upon IKAROS loss, expression of pre-B-cell differentiation genes is attenuated, while a group of extralineage transcription factors that are directly repressed by IKAROS and depend on EBF1 relocalization at their enhancers for expression is induced. LHX2, LMO2, and TEAD-YAP1, normally kept separate from native B-cell transcription regulators by IKAROS, now cooperate directly with them in a de novo superenhancer network with its own feed-forward transcriptional reinforcement. Induction of de novo superenhancers antagonizes Polycomb repression and superimposes aberrant stem-epithelial cell properties in a B-cell precursor. This dual mechanism of IKAROS regulation promotes differentiation while safeguarding against a hybrid stem-epithelial-B-cell phenotype that underlies high-risk B-ALL.

Key Points Ebf1 regulates DNA repair in a dose-dependent manner. Combined heterozygote loss of Ebf1 and Pax5 predisposes for leukemia development.

No abstract