scholarly journals Involvement of a second lymphoid-specific enhancer element in the regulation of immunoglobulin heavy-chain gene expression.

1990 ◽  
Vol 10 (6) ◽  
pp. 3155-3162 ◽  
Author(s):  
T A Libermann ◽  
M Lenardo ◽  
D Baltimore
Keyword(s):  
Binding Site ◽  
Heavy Chain ◽  
Critical Role ◽  
Immunoglobulin Heavy Chain ◽  
Lymphoid Cells ◽  
Chain Gene ◽  
Specific Expression ◽  
Enhancer Activity ◽  
Enhancer Element ◽  
Dna Motif

To determine whether enhancer elements in addition to the highly conserved octamer (OCTA)-nucleotide motif are important for lymphoid-specific expression of the immunoglobulin heavy-chain (IgH) gene, we have investigated the effect of mutating the binding site for a putative additional lymphoid-specific transcription factor, designated NF-microB, in the murine IgH enhancer. We demonstrate that the NF-microB-binding site plays a critical role in the IgH enhancer, because mutation of the microB DNA motif decreased transcriptional activity of the IgH enhancer in cells of the B-cell lineage but not in nonlymphoid cells. This effect was comparable to or even stronger than the effect of a mutation in the OCTA site. Moreover, combined mutation of both microB and OCTA sites further reduced enhancer activity in lymphoid cells. Interestingly, alteration of either the microB or E3 site in a 70-base-pair fragment of the IgH enhancer that lacks the binding site for OCTA abolished enhancer activity in lymphoid cells completely. Nevertheless, a multimer of the microB motif alone showed no enhancer activity. DNase footprinting analysis corroborated the functional data showing that a lymphoid-specific protein binds to the microB DNA motif. Our results suggest that the microB element is a new crucial element important for lymphoid-specific expression of the IgH gene but that interaction with another enhancer element is essential for its activity.

scholarly journals Involvement of a second lymphoid-specific enhancer element in the regulation of immunoglobulin heavy-chain gene expression

1990 ◽  
Vol 10 (6) ◽  
pp. 3155-3162
Author(s):  
T A Libermann ◽  
M Lenardo ◽  
D Baltimore
Keyword(s):  
Binding Site ◽  
Heavy Chain ◽  
Critical Role ◽  
Immunoglobulin Heavy Chain ◽  
Lymphoid Cells ◽  
Chain Gene ◽  
Specific Expression ◽  
Enhancer Activity ◽  
Enhancer Element ◽  
Dna Motif

To determine whether enhancer elements in addition to the highly conserved octamer (OCTA)-nucleotide motif are important for lymphoid-specific expression of the immunoglobulin heavy-chain (IgH) gene, we have investigated the effect of mutating the binding site for a putative additional lymphoid-specific transcription factor, designated NF-microB, in the murine IgH enhancer. We demonstrate that the NF-microB-binding site plays a critical role in the IgH enhancer, because mutation of the microB DNA motif decreased transcriptional activity of the IgH enhancer in cells of the B-cell lineage but not in nonlymphoid cells. This effect was comparable to or even stronger than the effect of a mutation in the OCTA site. Moreover, combined mutation of both microB and OCTA sites further reduced enhancer activity in lymphoid cells. Interestingly, alteration of either the microB or E3 site in a 70-base-pair fragment of the IgH enhancer that lacks the binding site for OCTA abolished enhancer activity in lymphoid cells completely. Nevertheless, a multimer of the microB motif alone showed no enhancer activity. DNase footprinting analysis corroborated the functional data showing that a lymphoid-specific protein binds to the microB DNA motif. Our results suggest that the microB element is a new crucial element important for lymphoid-specific expression of the IgH gene but that interaction with another enhancer element is essential for its activity.

Pi, a pre-B-cell-specific enhancer element in the immunoglobulin heavy-chain enhancer

1993 ◽  
Vol 13 (10) ◽  
pp. 5957-5969
Author(s):  
T A Libermann ◽  
D Baltimore
Keyword(s):  
B Cell ◽  
Heavy Chain ◽  
Mutational Analysis ◽  
Protein S ◽  
Immunoglobulin Heavy Chain ◽  
Chain Gene ◽  
Cell Stage ◽  
Enhancer Activity ◽  
Enhancer Element ◽  
Light Chain Gene

We have identified a new immunoglobulin heavy-chain enhancer element, designated pi, between the microE2 and microE3 elements. The pi enhancer element is transcriptionally active primarily during early stages of B-cell development but becomes virtually inactive during B-cell maturation at about the stage of immunoglobulin kappa light-chain gene rearrangement. Mutational analysis suggests that the pi element is crucial for immunoglobulin heavy-chain enhancer activity at the pre-B-cell stage but is almost irrelevant for enhancer activity at the mature B-cell or plasma-cell stage. The activity of the pi enhancer element correlates with the presence of an apparently pre-B-cell-specific protein-DNA complex. The similarity of the pi site to recognition sequences for members of the ets gene family suggests that the protein(s) interacting with the pi site most likely are ets-related transcription factors.

scholarly journals Pi, a pre-B-cell-specific enhancer element in the immunoglobulin heavy-chain enhancer.

1993 ◽  
Vol 13 (10) ◽  
pp. 5957-5969 ◽  
Author(s):  
T A Libermann ◽  
D Baltimore
Keyword(s):  
B Cell ◽  
Heavy Chain ◽  
Mutational Analysis ◽  
Protein S ◽  
Immunoglobulin Heavy Chain ◽  
Chain Gene ◽  
Cell Stage ◽  
Enhancer Activity ◽  
Enhancer Element ◽  
Light Chain Gene

We have identified a new immunoglobulin heavy-chain enhancer element, designated pi, between the microE2 and microE3 elements. The pi enhancer element is transcriptionally active primarily during early stages of B-cell development but becomes virtually inactive during B-cell maturation at about the stage of immunoglobulin kappa light-chain gene rearrangement. Mutational analysis suggests that the pi element is crucial for immunoglobulin heavy-chain enhancer activity at the pre-B-cell stage but is almost irrelevant for enhancer activity at the mature B-cell or plasma-cell stage. The activity of the pi enhancer element correlates with the presence of an apparently pre-B-cell-specific protein-DNA complex. The similarity of the pi site to recognition sequences for members of the ets gene family suggests that the protein(s) interacting with the pi site most likely are ets-related transcription factors.

scholarly journals Pan/E2A expression precedes immunoglobulin heavy-chain expression during B lymphopoiesis in nontransformed cells, and Pan/E2A proteins are not detected in myeloid cells.

1994 ◽  
Vol 14 (6) ◽  
pp. 4087-4096 ◽  
Author(s):  
Y Jacobs ◽  
X Q Xin ◽  
K Dorshkind ◽  
C Nelson
Keyword(s):  
B Cell ◽  
Cell Line ◽  
Heavy Chain ◽  
B Lymphocyte ◽  
Myeloid Cells ◽  
Immunoglobulin Heavy Chain ◽  
Cell Development ◽  
B Cell Development ◽  
Chain Gene ◽  
Enhancer Element

A newly developed rat long-term bone marrow culture system was used to study the role of Pan/E2A basic helix-loop-helix transcription factors during B-cell development. In this system, B-lymphocyte progenitors actively differentiate into mature B cells. Monoclonal (Yae) and polyclonal (anti-Pan) antibodies were employed to characterize the expression of Pan proteins by Western blot assay during hematopoiesis and to examine the components of immunoglobulin heavy-chain gene enhancer element-binding species by electrophoretic mobility shift assay. During B-cell development, the appearance of Pan/E2A proteins preceded the expression of immunoglobulin heavy-chain protein. A Pan-containing immunoglobulin heavy-chain enhancer element (mu E5)-binding species (BCF1), composed of immunoreactive Pan-1/E47 but not Pan-2/E12, was observed concomitantly with the detection of Pan/E2A proteins. In addition to BCF1, other mu E5-binding species were detected which were not recognized by the Yae antibody. Two of these species were present in primary B-lymphocyte and myeloid cultures and were recognized by an anti-upstream stimulatory factor antiserum. Although Pan/E2A proteins have been proposed to be ubiquitous, Pan/E2A proteins were not detected in primary myeloid cultures composed mainly of granulocytes and macrophages or in the macrophage cell line J774. The absence of Pan/E2A proteins in differentiated myeloid cells correlated with low steady-state levels of Pan/E2A RNA. However, Pan/E2A proteins were present in a promyeloid cell line, 32DCL3, suggesting that extinction of Pan/E2A expression may play a role in myelopoiesis.

scholarly journals Genes activated in the presence of an immunoglobulin enhancer or promoter are negatively regulated by a T-lymphoma cell line.

1988 ◽  
Vol 8 (5) ◽  
pp. 1932-1939 ◽  
Author(s):  
D M Zaller ◽  
H Yu ◽  
L A Eckhardt
Keyword(s):  
Heavy Chain ◽  
Immunoglobulin Heavy Chain ◽  
Negative Control ◽  
Lymphoma Cell ◽  
Chain Gene ◽  
Immunoglobulin Genes ◽  
Heavy Chain Gene ◽  
Immunoglobulin Heavy Chain Gene ◽  
Specific Expression ◽  
T Lymphoma

The tissue-specific expression of immunoglobulin genes can be partially explained by a requirement for activating factors found only in B lymphocytes and their derivatives. However, loss of immunoglobulin expression upon fusion of an immunoglobulin-producing myeloma cell with a T lymphoma cell (BW5147) or fibroblast (L cell) suggests that negatively acting factors also play a role in the tissue specificity of immunoglobulin genes. Expression of a cloned immunoglobulin heavy-chain gene introduced into myeloma cells was suppressed after fusion of the myeloma transformants with BW5147. The presence of either the immunoglobulin heavy-chain enhancer or promoter conferred suppression, under similar conditions, upon a heterologous gene that is normally expressed in both B and T lymphocytes. These immunoglobulin heavy-chain gene control regions, or gene modifications induced by them, are subject to negative control by T-lymphocyte-derived factors.

scholarly journals Identification of a novel factor that interacts with an immunoglobulin heavy-chain promoter and stimulates transcription in conjunction with the lymphoid cell-specific factor OTF2.

1990 ◽  
Vol 10 (5) ◽  
pp. 2145-2153 ◽  
Author(s):  
B K Yoza ◽  
R G Roeder
Keyword(s):  
B Cell ◽  
Heavy Chain ◽  
Regulatory Element ◽  
Immunoglobulin Heavy Chain ◽  
Upstream Region ◽  
Chain Gene ◽  
Specific Factor ◽  
Mobility Shift ◽  
Specific Expression ◽  
Cell Extracts

The tissue-specific expression of the MOPC 141 immunoglobulin heavy-chain gene was studied by using in vitro transcription. B-cell-specific transcription of this gene was dependent on the octamer element 5'-ATGCAAAG-3', located in the upstream region of this promoter and in the promoters of all other immunoglobulin heavy- and light-chain genes. The interaction of purified octamer transcription factors 1 and 2 (OTF1 and OTF2) with the MOPC 141 promoter was studied by using electrophoretic mobility shift assays and DNase I footprinting. Purified OTF1 from HeLa cells and OTF1 and OTF2 from B cells bound to identical sequences within the heavy-chain promoter. The OTF interactions we observed extended over the heptamer element 5'-CTCAGGA-3', and it seems likely that the binding of the purified factors involves cooperation between octamer and heptamer sites in this promoter. In addition to these elements, we identified a second regulatory element, the N element with the sequence 5'-GGAACCTCCCCC-3'. The N element could independently mediate low levels of transcription in both B-cell and HeLa-cell extracts, and, in conjunction with the octamer element, it can promote high levels of transcription in B-cell extracts. The N element bound a transcription factor, NTF, that is ubiquitous in cell-type distribution, and NTF was distinct from any of the previously described proteins that bind to similar sequences. Based on these results, we propose that NTF and OTF2 interactions (both with their cognate DNA elements and possibly at the protein-protein level) may be critical to B-cell-specific expression and that these interactions provide additional pathways for regulating gene expression.

scholarly journals Binding in vitro of multiple cellular proteins to immunoglobulin heavy-chain enhancer DNA.

1986 ◽  
Vol 6 (12) ◽  
pp. 4168-4178 ◽  
Author(s):  
C L Peterson ◽  
K Orth ◽  
K L Calame
Keyword(s):  
Heavy Chain ◽  
Protein Interactions ◽  
Binding Sites ◽  
Immunoglobulin Heavy Chain ◽  
Lymphoid Cells ◽  
Protein Protein Interactions ◽  
Protein Activity ◽  
Enhancer Element ◽  
Enhancer Sequences

Seven protein-binding sites on the immunoglobulin heavy-chain (IgH) enhancer element have been identified by exonuclease III protection and gel retardation assays. It appears that the seven sites bind a minimum of four separate proteins. Three of these proteins also bind to other enhancers or promoters, but one protein seems to recognize exclusively IgH enhancer sequences. A complex of four binding sites, recognized by different proteins, is located within one 80-base-pair region of IgH enhancer DNA. Close juxtaposition of enhancer proteins may allow protein-protein interactions or be part of a mechanism for modulating enhancer protein activity. All IgH enhancer-binding proteins identified in this study were found in extracts from nonlymphoid as well as lymphoid cells. These data provide the first direct evidence that multiple proteins bind to enhancer elements and that while some of these proteins recognize common elements of many enhancers, others have more limited specificities.

Genes activated in the presence of an immunoglobulin enhancer or promoter are negatively regulated by a T-lymphoma cell line

1988 ◽  
Vol 8 (5) ◽  
pp. 1932-1939
Author(s):  
D M Zaller ◽  
H Yu ◽  
L A Eckhardt
Keyword(s):  
Heavy Chain ◽  
Immunoglobulin Heavy Chain ◽  
Negative Control ◽  
Lymphoma Cell ◽  
Chain Gene ◽  
Immunoglobulin Genes ◽  
Heavy Chain Gene ◽  
Immunoglobulin Heavy Chain Gene ◽  
Specific Expression ◽  
T Lymphoma

The tissue-specific expression of immunoglobulin genes can be partially explained by a requirement for activating factors found only in B lymphocytes and their derivatives. However, loss of immunoglobulin expression upon fusion of an immunoglobulin-producing myeloma cell with a T lymphoma cell (BW5147) or fibroblast (L cell) suggests that negatively acting factors also play a role in the tissue specificity of immunoglobulin genes. Expression of a cloned immunoglobulin heavy-chain gene introduced into myeloma cells was suppressed after fusion of the myeloma transformants with BW5147. The presence of either the immunoglobulin heavy-chain enhancer or promoter conferred suppression, under similar conditions, upon a heterologous gene that is normally expressed in both B and T lymphocytes. These immunoglobulin heavy-chain gene control regions, or gene modifications induced by them, are subject to negative control by T-lymphocyte-derived factors.

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