Immunoglobulin heavy-chain enhancer is required to maintain transfected gamma 2A gene expression in a pre-B-cell line.
The immunoglobulin heavy-chain (IgH) enhancer serves to activate efficient and accurate transcription of cloned IgH genes when introduced into B lymphomas or myelomas. The role of this enhancer after gene activation, however, is unclear. The endogenous IgH genes in several cell lines, for example, have lost the IgH enhancer by deletion and yet continue to be expressed. This might be explained if the role of the enhancer were to establish high-level gene transcription but not to maintain it. Alternatively, other enhancers might lie adjacent to endogenous IgH genes, substituting their activity for that of the lost IgH enhancer. To address both of these alternatives, we searched for enhancer activity within the flanking regions of one of these IgH enhancer-independent genes and designed an experiment that allowed us to consider separately the establishment and maintenance of expression of a transfected gene. For the latter experiment we generated numerous pre-B cell lines stably transformed with a gamma 2a gene. In this gene, the IgH enhancer lay at a site outside the heavy-chain transcription unit, between DH and JH gene segments. After expression of the transfected gene was established, selective conditions were chosen for the outgrowth of subclones that had undergone D-J joining and thus IgH enhancer deletion. Measurements of gamma 2a expression before and after enhancer deletion revealed that the enhancer was required for maintenance of expression of the transfected gene. The implication of this finding for models of enhancer function in endogenous genes is discussed.
Porton
B
B
Department of Biological Sciences, Columbia University, New York, New York 10021.
Zaller
D M
DM
Lieberson
R
R
Eckhardt
L A
LA
D000818:Animals; D001402:B-Lymphocytes / Q000502:physiology*; D015152:Blotting, Northern; D015139:Blotting, Southern; D002460:Cell Line; D004742:Enhancer Elements, Genetic; D005786:Gene Expression Regulation; D005803:Genes, Immunoglobulin; D007074:Immunoglobulin G / Q000378:metabolism; D007143:Immunoglobulin Heavy Chains / Q000235:genetics*; D007142:Immunoglobulin gamma-Chains / Q000235:genetics*; D066298:In Vitro Techniques; D051379:Mice; D012333:RNA, Messenger / Q000235:genetics; D015533:Transcriptional Activation; D014162:Transfection
D007074:Immunoglobulin G; D007143:Immunoglobulin Heavy Chains; D007142:Immunoglobulin gamma-Chains; D012333:RNA, Messenger