Evidence for physical interaction between the immunoglobulin heavy chain variable region and the 3′ regulatory region

B cell-specific expression of immunoglobulin heavy chain (IgH) genes utilizes two cis regulatory regions, the intronic enhancer (E mu), located in the J(H)-C mu intron, and a complex regulatory region that lies 3 ' to the IgH gene cluster, 3 ' RR. We hypothesized that the 3 ' RR is involved in IgH gene transcription in plasma cells via physical interaction between distal 3 ' RR enhancers and target V-H sequences, with loop formation by intervening DNA. In support of this hypothesis we report sequence data at DNA recombination breakpoints as evidence for loop formation preceding DNA inversion in a plasma cell line. In addition, using the chromosome conformation capture technique, physical interactions between VH and 3 ' RR were analyzed directly and detected in MPC11 plasma cells and variants and normal splenic B cells but not detected in splenic T cells or in non-B cells. V-H-3 ' RR interactions were present in the absence of E mu, but when the hs1,2 enhancer was replaced by a Neo(R) gene in a variant cell line lacking E mu, H chain expression was lost, and interactions between VH and 3 ' RR and among the 3 ' RR regulators themselves were severely disrupted. In addition, the chromosome conformation capture technique detected interactions between the myc promoter and 3 ' RR elements in MPC11, which like other plasmacytomas contains a reciprocal translocation between the c-myc and the IgH locus. In sum, our data support a hypothesis that cis V-H-3 ' RR and myc-3 ' RR interactions involve physical interactions between these DNA elements.