scholarly journals The binding of the ubiquitous transcription factor Sp1 at the locus control region represses the expression of  -like globin genes

2005 ◽  
Vol 102 (28) ◽  
pp. 9896-9900 ◽  
Author(s):  
D. Feng ◽  
Y. W. Kan
Keyword(s):  
Transcription Factor ◽  
Control Region ◽  
Locus Control Region ◽  
Globin Genes ◽  
Transcription Factor Sp1 ◽  
Ubiquitous Transcription Factor

Position independence and proper developmental control of gamma-globin gene expression require both a 5' locus control region and a downstream sequence element

1994 ◽  
Vol 14 (9) ◽  
pp. 6087-6096
Author(s):  
Q Li ◽  
J A Stamatoyannopoulos
Keyword(s):  
Transgenic Mice ◽  
Control Region ◽  
Locus Control Region ◽  
Regulatory Sequences ◽  
Globin Genes ◽  
Sequence Element ◽  
Position Effects ◽  
Developmental Control ◽  
Developmental Patterns ◽  
Gamma Globin

We have analyzed the expression of human gamma-globin genes during development in F2 progeny of transgenic mice carrying two types of constructs. In the first type, gamma-globin genes were linked individually to large (approximately 4-kb) sequence fragments spanning locus control region (LCR) hypersensitive site 2 (HS2) or HS3. These LCR fragments contained not only the core HS elements but also extensive evolutionarily conserved flanking sequences. The second type of construct contained tandem gamma- and beta-globin genes linked to identical HS2 or HS3 fragments. We show that gamma-globin expression in transgenic mice carrying HS2 gamma or HS3 gamma constructs is highly sensitive to position effects and that such effects override the cis regulatory elements present in these constructs to produce markedly different developmental patterns of gamma-globin expression in lines carrying the same transgene. In contrast, gamma-globin expression in both HS2 gamma beta and HS3 gamma beta mice is sheltered from position effects and the developmental patterns of gamma-globin expression in lines carrying the same transgene are identical and display stage-specific regulation. The results suggest that cis regulatory sequences required for proper developmental control of fetal globin expression in the presence of an LCR element reside downstream from the gamma genes.

scholarly journals Promoters of the murine embryonic  -like globin genes Ey and  h1 do not compete for interaction with the  -globin locus control region

2003 ◽  
Vol 100 (3) ◽  
pp. 1111-1115 ◽  
Author(s):  
X. Hu ◽  
M. Bulger ◽  
J. N. Roach ◽  
S. K. Eszterhas ◽  
E. Olivier ◽  
...  
Keyword(s):  
Control Region ◽  
Locus Control Region ◽  
Globin Genes

Correct Function of the Locus Control Region May Require Passage Through a Nonerythroid Cellular Environment

Blood ◽  
1999 ◽  
Vol 93 (2) ◽  
pp. 703-712 ◽  
Author(s):  
George Vassilopoulos ◽  
Patrick A. Navas ◽  
Evangelia Skarpidi ◽  
Kenneth R. Peterson ◽  
Chris H. Lowrey ◽  
...  
Keyword(s):  
Gene Expression ◽  
Control Region ◽  
Globin Gene ◽  
Locus Control Region ◽  
Erythroid Cell ◽  
Globin Genes ◽  
Globin Mrna ◽  
L Cells ◽  
Correct Function ◽  
Globin Gene Expression

Abstract The function of the β-globin locus control region (LCR) has been studied both in cell lines and in transgenic mice. We have previously shown that when a 248-kb β-locus YAC was first microinjected into L-cells and then transferred into MEL cells by fusion, the YAC loci of the LxMEL hybrids displayed normal expression and developmental regulation.To test whether direct transfer of a β-globin locus (β-YAC) into MEL cells could be used for studies of the function of the LCR, a 155-kb β-YAC that encompasses the entire β-globin locus was used. This YAC was retrofitted with a PGK-neo selectable marker and with two I-PpoI sites at the vector arm-cloned insert junctions, allowing detection of the intact globin loci on a single I-PpoI fragment by pulsed field gel electrophoresis (PFGE). ThePpo-155 β-YAC was used to directly lipofect MEL 585 cells. In 7 β-YAC MEL clones with at least one intact copy of the YAC, the levels of total human globin mRNA (ie, ɛ + γ + β) per copy of integrated β-YAC varied more than 97-fold between clones. These results indicated that globin gene expression was strongly influenced by the position of integration of the β-YAC into the MEL cell genome and suggested that the LCR cannot function properly when the locus is directly transferred into an erythroid cell environment as naked β-YAC DNA. To test whether passage of the β-YAC through L-cells before transfer into MEL cells was the reason for the previously observed correct developmental regulation of human globin genes in the LxMEL hybrid cells, we transfected the YAC into L-cells by lipofection. Three clones carried the intact 144-kb I-PpoI fragment and transcribed the human globin genes with a fetal-like pattern. Subsequent transfer of the YAC of these L(β-YAC) clones into MEL cells by fusion resulted in LxMEL hybrids that synthesized human globin mRNA. The variation in human β-globin mRNA (ie, ɛ + γ + β) levels between hybrids was 2.5-fold, indicating that globin gene expression was independent of position of integration of the transgene, as expected for normal LCR function. The correct function of the LCR when the YAC is first transferred into the L-cell environment raises the possibility that normal activation of the LCR requires interaction with the transcriptional environment of an uncommitted, nonerythroid cell. We propose that the activation of the LCR may represent a multistep process initiated by the binding of ubiquitous transcription factors early during the differentiation of hematopoietic stem cells and completed with the binding of erythroid type of factors in the committed erythroid progenitors.

scholarly journals Human β-Globin Locus Control Region HS5Contains CTCF- and Developmental Stage-Dependent Enhancer-BlockingActivity in ErythroidCells

2003 ◽  
Vol 23 (24) ◽  
pp. 8946-8952 ◽  
Author(s):  
Keiji Tanimoto ◽  
Akiko Sugiura ◽  
Akane Omori ◽  
Gary Felsenfeld ◽  
James Douglas Engel ◽  
...  
Keyword(s):  
Developmental Stage ◽  
Control Region ◽  
Yeast Artificial Chromosome ◽  
Globin Gene ◽  
Artificial Chromosome ◽  
Locus Control Region ◽  
Ctcf Binding ◽  
Ctcf Binding Site ◽  
Globin Genes ◽  
Mechanistic Basis

ABSTRACT The human β-globin locus contains five developmentally regulatedβ -type globin genes. All five genes depend on the locus control region (LCR), located at the 5′ end of the locus, for abundant globin gene transcription. The LCR is composed of five DNase I-hypersensitive sites (HSs), at least a subset of which appear to cooperate to form a holocomplex in activating genes within the locus. We previously tested the requirement for proper LCR polarity by inverting it in human β-globin yeast artificial chromosome transgenic mice and observed reduced expression of all theβ -type globin genes regardless of developmental stage. This phenotype clearly demonstrated an orientation-dependent activity of the LCR, although the mechanistic basis for the observed activity was obscure. Here, we describe genetic evidence demonstrating that human HS5 includes enhancer-blocking (insulator) activity that is both CTCF and developmental stage dependent. Curiously, we also observed an attenuating activity in HS5 that was specific to the ε-globin gene at the primitive stage and was independent of the HS5 CTCF binding site. These observations demonstrate that the phenotype observed in the LCR-inverted locus was in part attributable to placing the HS5 insulator between the LCR HS enhancers (HS1 to HS4) and the promoter of the β-globin gene.

scholarly journals Developmental regulation of human gamma- and beta-globin genes in the absence of the locus control region

Blood ◽  
1994 ◽  
Vol 84 (5) ◽  
pp. 1656-1665 ◽  
Author(s):  
J Starck ◽  
R Sarkar ◽  
M Romana ◽  
A Bhargava ◽  
AL Scarpa ◽  
...  
Keyword(s):  
Transgenic Mice ◽  
Control Region ◽  
Locus Control Region ◽  
Fetal Life ◽  
Globin Genes ◽  
Beta Globin ◽  
Mouse Development ◽  
Rnase Protection ◽  
Adult Mice ◽  
Low Levels

Two lines of transgenic mice carrying a normal 40-kb Kpn I beta-globin cluster transgene lacking the locus control region (LCR) were analyzed for the expression of human gamma- and beta-globin genes during mouse development. After RNase protection assays, the ratios of human G gamma- , A gamma-, or beta-mRNAs relative to endogenous mouse zeta + alpha mRNAs were obtained for each stage of development. The two gamma transgenes were expressed in day-11.5 blood (embryonic stage) and day- 13.5 blood (early fetal stage), but their expression was markedly decreased by day 16.5 of fetal life. Expression of the beta transgene was essentially absent at day 13.5, appeared at a low level by day 16.5, and was maximal by day 18.5, reaching a level similar to that observed in adult mice. Therefore, developmentally regulated expression of the human gamma- and beta-globin transgenes was obtained in the absence of the LCR. The relative expression of human gamma- and beta- globin genes was also examined in mice carrying 40-kb Kpn I beta- cluster transgenes with two different base substitutions associated with nondeletion forms of hereditary persistence of fetal hemoglobin (HPFH), -202 C-->G G gamma HPFH and -117 G-->A A gamma HPFH. The ratio of G gamma- to beta-globin transcripts was markedly increased in red blood cells of adult mice from three different lines carrying the transgene with the -202 G gamma HPFH mutation. This result confirms our previous preliminary results (Tanaka et al: Ann NY Acad Sci, 612:167, 1990) indicating that the -202 G gamma HPFH phenotype was reproduced in transgenic mice. The relatively low levels of G gamma-mRNA expression in adult mice carrying the non-HPFH transgene excludes a major influence of the 3′ beta-globin enhancer, present upstream of the G gamma gene because of the tandem repeat insertion, as a factor in the persistent G gamma gene expression observed in blood of adult mice carrying the -202 G gamma HPFH transgene. This conclusion is also supported by the fact that, in mice carrying the -117 A gamma HPFH transgene, G gamma-globin mRNA was detected in blood of adult animals only at low levels similar to that observed in the non-HPFH lines. However, the A gamma-HPFH phenotype was not reproduced in the transgenic lines carrying the -117A gamma HPFH mice.

scholarly journals The hypersensitive sites of the murine β-globin locus control region act independently to affect nuclear localization and transcriptional elongation

Blood ◽  
2012 ◽  
Vol 119 (16) ◽  
pp. 3820-3827 ◽  
Author(s):  
M. A. Bender ◽  
Tobias Ragoczy ◽  
Jongjoo Lee ◽  
Rachel Byron ◽  
Agnes Telling ◽  
...  
Keyword(s):  
Control Region ◽  
Globin Gene ◽  
Nuclear Periphery ◽  
Locus Control Region ◽  
Transcriptional Elongation ◽  
Globin Genes ◽  
Hypersensitive Sites ◽  
Nuclear Location ◽  
High Level ◽  
Rna Transcript

Abstract The β-globin locus control region (LCR) is necessary for high-level β-globin gene transcription and differentiation-dependent relocation of the β-globin locus from the nuclear periphery to the central nucleoplasm and to foci of hyperphosphorylated Pol II “transcription factories” (TFys). To determine the contribution of individual LCR DNaseI hypersensitive sites (HSs) to transcription and nuclear location, in the present study, we compared β-globin gene activity and location in erythroid cells derived from mice with deletions of individual HSs, deletions of 2 HSs, and deletion of the whole LCR and found all of the HSs had a similar spectrum of activities, albeit to different degrees. Each HS acts as an independent module to activate expression in an additive manner, and this is correlated with relocation away from the nuclear periphery. In contrast, HSs have redundant activities with respect to association with TFys and the probability that an allele is actively transcribed, as measured by primary RNA transcript FISH. The limiting effect on RNA levels occurs after β-globin genes associate with TFys, at which time HSs contribute to the amount of RNA arising from each burst of transcription by stimulating transcriptional elongation.

Recruitment of Rad50 and MCM Complexes on the Human Beta Globin Locus Control Region (LCR): A Novel Role for LCR Hypersensitive Site 4 (HS4).

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 273-273
Author(s):  
Subhradip Karmakar ◽  
Milind C. Mahajan ◽  
Sherman M. Weissman
Keyword(s):  
Dna Replication ◽  
Control Region ◽  
Protein Complex ◽  
Locus Control Region ◽  
Partial Purification ◽  
Open Chromatin ◽  
Gene Promoters ◽  
Globin Genes ◽  
Beta Globin ◽  
Wide Range

Abstract The human beta globin locus control region (LCR) aids in the establishment of an open chromatin configuration thereby facilitating the expression of linked downstream globin genes in a temporal and spatial manner along the developmental ontogeny. The LCR also functions as a classical enhancer of the beta like globin genes. The four DNAase-I hypersensitive regions (HS 1–4) on the LCR are believed to act as nucleation centers for the assembly of multi-protein transcriptional complexes that interact with the distant gene promoters. Present work is focused on characterizing the protein complex assemblies on the HS4 region and delineating their function in the context of goblin gene regulation. Our finding shows an assembly of a 2–5 MDa complex on the HS4 locus that is both erythroid restricted and sequence specific. Electrophoretic mobility shift assays (EMSA) were performed using K562 and HeLa nuclear extracts and ∼35mer double stranded oligonucleotides with 7 bp overlaps tiling the entire core HS4 sequence. We detected prominent K562 specific bands formed with an AT rich 39 bp HS4 sequence. Sequence specificity of DNA-protein complex formation on this AT rich sequence was determined by competitive EMSA assays. Gel supershift assays with a wide range of antibodies and subsequent partial purification of this multiprotein complex followed by the LC-MS/MS analysis reveal the presence of a mini chromosome maintenance (MCM) complex that is associated with the DNA replication, DNA repair associated Rad50-Mre-11-NBS-1 complex, histone acetyl transferase p300 and DNA binding transcription factors ILF2 (NF45) and ILF3 (NF90). The association of these proteins was determined by co-immunoprecipitation experiments. Chromatin immunoprecipitation (ChIP) experiments demonstrate the in vivo recruitment of these proteins on the HS4 region of the LCR. We are currently working on further characterization and functional studies of this complex and its significance on the role of the LCR in DNA replication and repair.

Sign in / Sign up

Export Citation Format

Share Document