Possible interaction between B1 retrotransposon-containing sequences and βmajorglobin gene transcriptional activation during MEL cell erythroid differentiation (original) (raw)
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DNA sequences required for regulated expression of β-globin genes in murine erythroleukemia cells
Cell, 1984
We introduced into MEL cells rabbit /3-globin gene deletion mutants and two sets of hybrid genes constructed from the inducible human @-globin gene and noninducible human y-globin gene or the murine H-2K"'"' class I MHC gene. Sl nuclease analysis of gene transcripts before and after MEL differentiation showed that induction of the rabbit @-globin gene did not require more than 58 bp of DNA 5' to the transcription initiation site. Hybrid genes were constructed with human B-globin DNA sequences from either 5' or 3' of the translation initiation site linked to the complementary parts of they or H2Kbm' genes. Both types of constructs were inducible during MEL differentiation. The relative rates of transcription of the 5'7-3'8 and 5'H2-3'8 hybrid genes show that induction of the hybrid gene transcripts results at least in part from transcriptional activation of the genes. We suggest that DNA sequences that regulate 8-globin gene transcription during MEL differentiation are located both 5' and 3' to the translation initiation site.
Activation of the beta-globin locus control region precedes commitment to the erythroid lineage
Proceedings of the National Academy of Sciences, 1992
The (3-globin locus control region (LCR) is characterized by erythroid-speciflc DNase I hypersensitive sites and is involved in the chromatin organization, trancriptional potentiation, developmental regulation, and replication tming of the entire (-globin gene duster. When and how the LCR is first activated during erythropoiesis is not known. Here we analyze the chromatin structure of the LCR during early hematopoietic differentiation using nontransformed, multipotential, growth factor-dependent, murine hematopoietic progenitor cells. We show that LCR hypersensitive sites characteristic of erythroid cells are present in three independent multilineage progenitors (FDCP (factor-dependent cell, Paterson)-mix A4, B6SUtA, and LyD9] under conditions of selfrenewal. Induction of differentiation down a nonerythroid pathway causes a progressive loss of hypersensitivity in the LCR. These results show that the (3-globin LCR is in an active chromatin configuration prior to erythroid commitment and indicate a significant role for selective gene repression in lineage specification.
The EMBO journal, 1988
We have shown that the promoter of the human beta-globin gene contains three regions in addition to the known CAC, CAAT and TATA box regions that are important for the induction of transcription in erythroid cells. By using DNaseI footprinting and gel mobility shift assays we were able to show that two of these regions bind the erythroid specific nuclear factor NF-E1 (and ubiquitous factors). The third region binds a ubiquitous CAAT-box factor (CP1). Deletion experiments suggest that only the combination of NF-E1 and CP1 binding sites, but not each of the sites alone, are capable of mediating the induction of transcription of a minimal (CAC, CAAT, TATA box) beta-globin promoter in mouse erythroleukaemia (MEL) cells.
Globin gene expression in correlation with G protein-related genes during erythroid differentiation
BMC Genomics, 2013
Background: The guanine nucleotide binding protein (G protein)-coupled receptors (GPCRs) regulate cell growth, proliferation and differentiation. G proteins are also implicated in erythroid differentiation, and some of them are expressed principally in hematopoietic cells. GPCRs-linked NO/cGMP and p38 MAPK signaling pathways already demonstrated potency for globin gene stimulation. By analyzing erythroid progenitors, derived from hematopoietic cells through in vitro ontogeny, our study intends to determine early markers and signaling pathways of globin gene regulation and their relation to GPCR expression. Results: Human hematopoietic CD34 + progenitors are isolated from fetal liver (FL), cord blood (CB), adult bone marrow (BM), peripheral blood (PB) and G-CSF stimulated mobilized PB (mPB), and then differentiated in vitro into erythroid progenitors. We find that growth capacity is most abundant in FL-and CB-derived erythroid cells. The erythroid progenitor cells are sorted as 100% CD71 + , but we did not find statistical significance in the variations of CD34, CD36 and GlyA antigens and that confirms similarity in maturation of studied ontogenic periods. During ontogeny, beta-globin gene expression reaches maximum levels in cells of adult blood origin (176 fmol/μg), while gamma-globin gene expression is consistently up-regulated in CB-derived cells (60 fmol/μg). During gamma-globin induction by hydroxycarbamide, we identify stimulated GPCRs (PTGDR, PTGER1) and GPCRs-coupled genes known to be activated via the cAMP/PKA (ADIPOQ), MAPK pathway (JUN) and NO/cGMP (PRPF18) signaling pathways. During ontogeny, GPR45 and ARRDC1 genes have the most prominent expression in FL-derived erythroid progenitor cells, GNL3 and GRP65 genes in CB-derived cells (high gamma-globin gene expression), GPR110 and GNG10 in BM-derived cells, GPR89C and GPR172A in PB-derived cells, and GPR44 and GNAQ genes in mPB-derived cells (high beta-globin gene expression).
Experimental Hematology, 2009
Objective. To investigate the mechanism(s) responsible for increased g-globin expression in vivo in decitabine-treated baboons and in vitro in cultured erythroid progenitor cells (EPC) from adult baboon bone marrow (BM). Materials and Methods. Fetal liver, adult BM erythroid cells pre-and post-decitabine, and cultured EPCs were analyzed for distribution of RNA polymerase II, histone acetylation, and histone H3 (lys4) trimethyl throughout the g-globin gene complex by chromatin immunoprecipitation. DNA methylation of the g-globin promoter was determined by bisulfite sequencing. Expression of the baboon Ig-and Vg-globin chains was determined by high performance liquid chromatography (HPLC). Expression of BCL11A, a recently identified repressor of g-globin expression, was analyzed by Western blot. Results. Increased g-globin expression in decitabine-treated baboons and cultured EPC correlated with increased levels of RNA polymerase II, histone acetylation, and histone H3 (lys4) trimethyl associated with the g-globin gene consistent with a transcriptional activation mechanism. Cultured EPC expressed the Ig-and Vg-globin chains in a pattern characteristic of fetal development. The level of DNA methylation of the g-globin gene promoter in EPC cultures was similar to BM erythroid cells from normal adult baboons. Different BCL11A isoforms were observed in BM erythroid cells and cultured EPC. Conclusion. The mechanism responsible for increased g-globin expression in cultured EPC was unexpectedly not associated with increased DNA hypomethylation of the g-globin gene promoter compared to normal BM erythroid cells, in contrast to BM erythroid cells of decitabine-treated baboons. Rather, increased fetal hemoglobin in EPC cultures was associated with a fetal Ig/Vg chain ratio and a difference in the size of the BCL11A protein compared to normal BM erythroid cells.
Developmental programs of human erythroleukemia cells: globin gene expression and methylation
Molecular and cellular biology, 1988
We investigated the programs of globin gene expression in three known (K562, HEL, and KMOE) and three novel (OCI-M1, OCI-M2, and HEL-R) human erythroleukemic cell lines of adult origin. RNAs from induced and uninduced cells were analyzed for epsilon-, gamma-, delta-, and beta-, zeta-globin-specific transcripts. While high-level gamma-globin expression was common, the lines differed in their expression of embryonic (epsilon, zeta) and adult (delta, beta) globin mRNAs. The patterns of globin gene methylation were generally consistent with their observed expression profiles, with many of the same correlations being seen in normal cells. Although the programs of globin gene expression and methylation displayed by the lines appeared to be diverse, they were not random; rather, they made developmental sense, mimicking defined globin gene programs observed during normal human development. The characteristics exhibited by several of these lines suggest that they may have been derived from t...
Cellular and Molecular Life Sciences, 1999
Communicated by F.Grosveld We have shown that the promoter of the human ,B-globin gene contains three regions in addition to the known CAC, CAAT and TATA box regions that are important for the induction of transcription in erythroid cells. By using DNaseI footprinting and gel mobility shift assays we were able to show that two of these regions bind the erythroid specific nuclear factor NF-E1 (and ubiquitous factors). The third region binds a ubiquitous CAAT-box factor (CP1). Deletion experiments suggest that only the combination of NF-E1 and CP1 binding sites, but not each of the sites alone, are capable of mediating the induction of transcription of a minimal (CAC, CAAT, TATA box) (3-globin promoter in mouse erythroleukaemia (MEL) cells.
Blood, 1985
In order to test if trans-acting regulatory factors specific for globin genes of the adult and embryonic stages of development exist in erythroid cells, transcriptionally active embryonic and adult globin genes on the same chromosome were transferred by cell fusion from the human leukemia cell K562 into phenotypically adult mouse erythroleukemia cells. Restriction-fragment-length polymorphisms of the K562 zeta (embryonic) globin genes were used to establish that all three copies of human chromosome 16 present in the K562 cell showed the same pattern of human globin gene expression after transfer to the mouse erythroleukemia cell. Adult (alpha) but not embryonic (zeta) human globin mRNA was detected in all nine of the independently derived mouse erythroleukemia hybrid cells, each of which contained human chromosome 16. Restriction endonuclease studies of the K562 alpha- and zeta-globin genes after transfer into the mouse erythroleukemia cell showed no evidence of rearrangements or de...
1998
Studies in many systems have led to the model that the human -globin locus control region (LCR) regulates the transcription, chromatin structure, and replication properties of the -globin locus. However the precise mechanisms of this regulation are unknown. We have developed strategies to use homologous recombination in a tissue culture system to examine how the LCR regulates the locus in its natural chromosomal environment. Our results show that when the functional components of the LCR, as defined by transfection and transgenic studies, are deleted from the endogenous -globin locus in an erythroid background, transcription of all -globin genes is abolished in every cell. However, formation of the remaining hypersensitive site(s) of the LCR and the presence of a DNase I-sensitive structure of the -globin locus are not affected by the deletion. In contrast, deletion of 5HS5 of the LCR, which has been suggested to serve as an insulator, has only a minor effect on -globin transcription and does not influence the chromatin structure of the locus. These results show that the LCR as currently defined is not necessary to keep the locus in an "open" conformation in erythroid cells and that even in an erythroid environment an open locus is not sufficient to permit transcription of the -like globin genes.
Blood, 1985
Analysis of methylation at the beta-globin gene cluster was carried out on DNA derived from nucleated RBCs (orthochromatic normoblasts) isolated from peripheral blood of patients with beta-thalassemia major or other congenital hemolytic anemia after splenectomy. A procedure to separate these normoblasts from the other nucleated cells of the peripheral blood was developed, providing us with a convenient source of DNA for investigating parameters related to human erythroid differentiation. Blood samples were obtained from six adult patients who express their gamma-globin genes at different levels. Inverse correlation between methylation and gene activity was consistently observed for five of the eight sites analyzed. A site 3' to the beta gene was always unmethylated, two sites flanking the epsilon gene were always found to be methylated, and two sites 5' to the two gamma genes, G gamma and A gamma, were hypomethylated in correlation with gamma gene activity of the individual ...