Fragments of the 5′‐flanking sequence of a human 2‐5A synthetase gene were assayed for their ability to respond to interferon‐alpha (IFN). Transient transfection assays in monkey cells demonstrated that the 5′ boundary of the sequence required for IFN‐regulated transcription is, at most, 155 nucleotides upstream from the presumed translational initiation codon. The 3′ boundary of this sequence lies within a region of multiple transcription start sites preceded by no obvious TATA box. Binding assays, using a 40‐bp probe derived from this IFN‐responsive sequence, demonstrated the presence of three IFN‐modulated, DNA‐factor band shifts using nuclear extracts prepared from human and monkey cells. The induction of these complexes in human cells by IFN occurs with kinetics which closely parallel those previously observed for the transcriptional activation of the 2‐5A synthetase gene by IFN. In vivo competition assays showed that the same 40‐bp region which bound IFN‐modulated factors could decrease the IFN‐induced activity of a co‐transfected 2‐5A synthetase promoter; this fragment, regardless of its orientation, could confer IFN‐inducibility on a heterologous promoter.

E2A is a basic helix-loop-helix (bHLH) transcription factor required for B cell lymphopoiesis and implicated in myogenesis and the regulation of insulin expression. As E2A is expressed widely in tissues, tissue-specific downstream effects are thought to result primarily from dimerization with other bHLH proteins. To investigate the degree to which regulation of E2A protein abundance may serve to regulate E2A function , expression of E2A was evaluated using immunohistochemistry on histological sections of primary human tissues. Somewhat surprisingly, nuclear staining for E2A was restricted in all tissues examined , often to a small subpopulation of cells. In some tissues , such as adult liver , expression was absent or limited to rare infiltrating lymphocytes. E2A-expressing cells were most abundant in lymphoid tissues. In tonsil , lymph node , and spleen , expression appeared most abundant and prevalent among rapidly proliferating centroblasts of the germinal center dark zone. Scattered E2A-expressing thymocytes were more numerous in the thymic cortex than medulla. In developing skeletal muscle , E2A was detectable in striated myotubes but not in more primitive mononucleated progenitors or mature muscle. Differential E2A expression was also noted in proliferating periventricular neuroepithelial cells in the developing brain. These results suggest that regulation of E2A abundance complements protein-protein interactions in modulating E2A function. (Am J Pathol 1998, 153:165-173) Embryogenesis and normal tissue homeostasis have in common the requirement for exquisite control of gene expression to generate optimal numbers of specialized, functional cells from pluripotential progenitors. Transcription factors are nuclear proteins that regulate the expression of downstream target genes. The role played by transcription factors as regulators of lineage commitment and cellular differentiation has been studied extensively in hematopoiesis and myogenesis. Protein products encoded by the transcription factor gene E2A play important roles in both of these systems.

No abstract

The chimeric transcription factor E2a-Pbx1 is expressed as a result of the 1;19 chromosomal translocation in some 5% of cases of pediatric acute lymphoblastic leukemia. We investigated the biological and transcriptional consequences of forced expression of E2a-Pbx1 in the interleukin-3 (IL-3) dependent, bone marrow-derived cell line Ba/F3. We show that forced expression of E2a-Pbx1 induces apoptosis in Ba/F3 cells without apparent effects on cell cycle progression. This pro-apoptotic effect is enhanced on cytokine deprivation. Furthermore, using cDNA representational difference analysis (RDA), we show that these cellular effects are associated with marked induction of the gene NDRG1, which was previously identified as a target of transcriptional repression by N-myc and induction by the tumor suppressor protein p53. We identify a portion of the NDRG1 promoter capable of mediating transcriptional induction by E2a-Pbx1 and show that NDRG1 is also induced on simple IL-3 deprivation of BaF3 cells. Although we show that E2a-Pbx1 induction of NDRG1 is not impaired as a result of targeting p53 using HPV E6, and therefore does not appear to be p53-dependent, our results overall are consistent with the notion that induction of NDRG1 by E2a-Pbx1 may represent part of an apoptotic or cytostatic cellular response to oncogene activation. Leukemia (2001) 15, 362-370.