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Research Article

Expression of the (Recombinant) Endogenous Immunoglobulin Heavy-Chain Locus Requires the Intronic Matrix Attachment Regions

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Pages 2658-2668 | Received 16 Dec 1996, Accepted 17 Feb 1997, Published online: 29 Mar 2023
 

Abstract

The elements which regulate gene expression have traditionally been identified by their effects on reporter genes which have been transfected into cell lines or animals. It is generally assumed that these elements have a comparable role in expression of the corresponding endogenous locus. Nevertheless, several studies of immunoglobulin heavy-chain (IgH) gene expression have reported that the requirements for expressing IgH-derived transgenes differ from the requirements for expression of the endogenous IgH locus. Thus, although expression of transgenes requires multiple elements from the JH-Cμ intron—the Eμ core enhancer, the matrix attachment regions (MARs) which flank Eμ, and several switch-associated elements—B-cell lines in which expression of the endogenous heavy-chain gene is maintained at the normal level in the absence of these intronic elements have occasionally been reported. Gene targeting offers an alternative method for assessing regulatory elements, one in which the role of defined segments of endogenous genes can be evaluated in situ. We have applied this approach to the IgH locus of a hybridoma cell line, generating recombinants which bear predetermined modifications in the functional, endogenous μ heavy-chain gene. Our analysis indicates the following. (i) Ninety-eight percent of the expression of the recombinant endogenous μ gene depends on elements in the MAR-Eμ-MAR segment. (ii) Expression of the recombinant μ gene depends strongly on the MARs of the JH-Cμ intron but not on the adjoining Eμ core enhancer and switch regions; because our recombinant cell lines bear only a single copy of the μ gene, our results indicate that μ expression is activated by MAR elements lying within that same μ transcription unit. (iii) The MAR segment includes at least one activating element in addition to those defined previously by the binding of presumptive activating proteins in the nuclear matrix. (iv) Close association of the MARs with the Eμ enhancer is not required for MAR-stimulated expression. (v) The other MARs in the IgH locus do not in their normal context provide the requisite MAR function.

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