BIASED READING FRAMES OF PREEXISTING DH-JH CODING JOINTS AND PREFERENTIAL NUCLEOTIDE INSERTIONS AT VH-DJH SIGNAL JOINTS OF EXCISION PRODUCTS OF IMMUNOGLOBULIN HEAVY-CHAIN GENE REARRANGEMENTS

During B cell differentiation immunoglobulin (Ig) D(H) segments join to J(H) segments, followed by joining of V(H) to DJ(H). Although circular excision products of D(H)-J(H) rearrangements have been characterized, excision products of V(H) to DJ(H) joining have never been isolated. We selectively denatured chromosomal DNA of mouse splenocytes and enriched circular DNA spanning the long distance between V(H) and D(H). Subsequent PCR amplifications allowed the identification of signal joints of V(H) to DJ(H). Sequence analysis indicated that pre-existing D(H)-J(H) coding joints of excision products showed a strong bias for reading frame 1, and the absence of reading frame 2, which would allow the expression of a truncated mu chain called D(mu) protein. When comparing the joints of the V(H)-DJ(H) and D(H)-J(H) rearrangements we observed N-nucleotide insertions to be abundant at the V(H)-D(H) signal joint, but very sparse at the D(H)-J(H) signal joint, while the coding joints of both contained abundant N-insertions. These differences in N region insertions at the signal joints suggest a differential control of the D-J and V-DJ rearrangements.