Immunoglobulin kappa gene enhancers synergistically activate gene expression but independently determine chromatin structure.
Previous studies have located transcriptional enhancer elements within both the intron and 3'-region of the mouse kappa immunoglobulin gene. Here we address the role of these two enhancers in specifying gene activity and specific chromatin structures. MOPC41 kappa gene constructs, either intact or containing deletions of one or both enhancers, were introduced into S194 mouse plasmacytoma cells for transient and stable expression studies. Transient expression assays revealed that the basal level expression exhibited by enhancerless constructs was activated 100-200-fold by the two enhancers together in a synergistic fashion. A similar trend was observed when both enhancers were present in stably integrated constructs, although the synergy was less pronounced. Analysis of DNase I hypersensitive sites in the chromatin revealed that stably integrated constructs established hypersensitive sites about the enhancer sequences. These sites demonstrated the same nuclease susceptibility as those associated with the endogenous gene(s), and their establishment was independent of the presence of the other enhancer. Thus, although both enhancers are required for maximal gene expression, the elements act independently in determining specific chromatin structures.
Blasquez
V C
VC
Department of Chemistry and Biochemistry, University of Notre Dame, Indiana 46556.
Hale
M A
MA
Trevorrow
K W
KW
Garrard
W T
WT
D000818:Animals; D015152:Blotting, Northern; D015139:Blotting, Southern; D002467:Cell Nucleus / Q000502:physiology; D002843:Chromatin / Q000502:physiology*; D002999:Clone Cells; D004273:DNA, Neoplasm / Q000235:genetics / Q000302:isolation & purification; D003850:Deoxyribonuclease I; D004742:Enhancer Elements, Genetic; D015972:Gene Expression Regulation, Neoplastic; D005803:Genes, Immunoglobulin; D007145:Immunoglobulin kappa-Chains / Q000235:genetics*; D007438:Introns; D051379:Mice; D010954:Plasmacytoma; D015183:Restriction Mapping; D017384:Sequence Deletion; D014162:Transfection; D014407:Tumor Cells, Cultured
D002843:Chromatin; D004273:DNA, Neoplasm; D007145:Immunoglobulin kappa-Chains; D003850:Deoxyribonuclease I