Faithful genome integrity maintenance plays an essential role in cell survival. Here, we identify the RNA demethylase ALKBH5 as a key regulator that protects cells from DNA damage and apoptosis during reactive oxygen species (ROS)-induced stress. We find that ROS significantly induces global mRNA N⁶-methyladenosine (m⁶A) levels by modulating ALKBH5 post-translational modifications (PTMs), leading to the rapid and efficient induction of thousands of genes involved in a variety of biological processes including DNA damage repair. Mechanistically, ROS promotes ALKBH5 SUMOylation through activating ERK/JNK signaling, leading to inhibition of ALKBH5 m⁶A demethylase activity by blocking substrate accessibility. Moreover, ERK/JNK/ALKBH5-PTMs/m⁶A axis is activated by ROS in hematopoietic stem/progenitor cells (HSPCs) in vivo in mice, suggesting a physiological role of this molecular pathway in the maintenance of genome stability in HSPCs. Together, our study uncovers a molecular mechanism involving ALKBH5 PTMs and increased mRNA m⁶A levels that protect genomic integrity of cells in response to ROS.

It is proposed that the multiple enhancer elements associated with locus control regions and super‐enhancers recruit RNA polymerase II and efficiently assemble elongation competent transcription complexes that are transferred to target genes by transcription termination and transient looping mechanisms. It is well established that transcription complexes are recruited not only to promoters but also to enhancers, where they generate enhancer RNAs. Transcription at enhancers is unstable and frequently aborted. Furthermore, the Integrator and WD‐domain containing protein 82 mediate transcription termination at enhancers. Abortion and termination of transcription at the multiple enhancers of locus control regions and super‐enhancers provide a large pool of elongation competent transcription complexes. These are efficiently captured by strong basal promoter elements at target genes during transient looping interactions.

Helix-loop-helix (HLH) proteins play a profound role in the process of development and cellular differentiation. Among the HLH proteins expressed in differentiating erythroid cells are the ubiquitous proteins Myc, USF1, USF2, and TFII-I, as well as the hematopoiesis-specific transcription factor Tal1/SCL. All of these HLH proteins exhibit distinct functions during the differentiation of erythroid cells. For example, Myc stimulates the proliferation of erythroid progenitor cells, while the USF proteins and Tal1 regulate genes that specify the differentiated phenotype. This minireview summarizes the known activities of Myc, USF, TFII-I, and Tal11/SCL and discusses how they may function sequentially, cooperatively, or antagonistically in regulating expression programs during the differentiation of erythroid cells.

The human β-globin gene is expressed at high levels in erythroid cells and regulated by proximal and distal cis-acting DNA elements, including promoter, enhancer, and a locus control region (LCR). Transcription complexes are recruited not only to the globin gene promoters but also to the LCR. Previous studies have implicated the ubiquitously expressed transcription factor USF and the tissue-restricted activator NF-E2 in the recruitment of transcription complexes to the β-globin gene locus. Here we demonstrate that although USF is required for the efficient association of RNA polymerase II (Pol II) with immobilized LCR templates, USF and NF-E2 together regulate the association of Pol II with the adult β-globin gene promoter. Recruitment of Pol II to the LCR occurs in undifferentiated murine erythroleukemia cells, but phosphorylation of LCR-associated Pol II at serine 5 of the C-terminal domain is mediated by erythroid differentiation and requires the activity of NF-E2. Furthermore, we provide evidence showing that USF interacts with NF-E2 in erythroid cells. The data provide mechanistic insight into how ubiquitous and tissue-restricted transcription factors cooperate to regulate the recruitment and activity of transcription complexes in a tissue-specific chromatin domain.

The human beta-globin genes are expressed in a developmental stage-specific manner in erythroid cells. Gene-proximal cis-regulatory DNA elements and interacting proteins restrict the expression of the genes to the embryonic, fetal, or adult stage of erythropoiesis. In addition, the relative order of the genes with respect to the locus control region contributes to the temporal regulation of the genes. We have previously shown that transcription factors TFII-I and USF interact with the beta-globin promoter in erythroid cells. Herein we demonstrate that reducing the activity of USF decreased beta-globin gene expression, while diminishing TFII-I activity increased beta-globin gene expression in erythroid cell lines. Furthermore, a reduction of USF activity resulted in a significant decrease in acetylated H3, RNA polymerase II, and cofactor recruitment to the locus control region and to the adult beta-globin gene. The data suggest that TFII-I and USF regulate chromatin structure accessibility and recruitment of transcription complexes in the beta-globin gene locus and play important roles in restricting beta-globin gene expression to the adult stage of erythropoiesis.

Many genes residing in gene clusters and expressed in a differentiation or developmental‐stage specific manner are regulated by locus control regions (LCRs). These complex genetic regulatory elements are often composed of several DNAse I hypersensitive sites (HS sites) that function together to regulate the expression of several cis‐linked genes. Particularly well characterized is the LCR associated with the β‐globin gene locus. The β‐globin LCR consists of five HS sites that are located upstream of the β‐like globin genes. Recent data demonstrate that the LCR is required for the association of the β‐globin gene locus with transcription foci or factories. The observation that RNA polymerase II associates with the LCR in erythroid progenitor or hematopoietic stem cells which do not express the globin genes suggests that the LCR is always in an accessible chromatin configuration during differentiation of erythroid cells. We propose that erythroid specific factors together with ubiquitous proteins mediate a change in chromatin configuration that juxtaposes the globin genes and the LCR. The proximity then facilitates the transfer of activities from the LCR to the globin genes. In this article we will discuss recent observations regarding β‐globin locus activation with a particular emphasis on LCR mediated activation of adult β‐globin gene expression. J. Cell. Biochem. 105: 9–16, 2008.