Germ-line transcripts of the immunoglobulin lambda J-C clusters in the mouse: characterization of the initiation sites and regulatory elements

Transcription of unrearranged immunoglobulin gene segments strongly correla tes with their accessibility to the V(D)J recombination machinery. The regu latory mechanisms governing this germ-line transcription are still poorly d efined. In order to identify new regulatory elements, we first carried out a detailed characterization of the transcription initiation sites for the J -C germ-line transcripts, using rapid amplification of 5' cDNA ends, assist ed by a template switching mechanism at the 5'-end of the RNA. Transcripts were observed that initiated heterogeneously, starting up to 293 (lambda1), 116 by (lambda2) and 79 by (lambda3) upstream from the respective R, gene segment. Additional RT-PCR analysis revealed the existence of germ-line tra nscripts of lambda and also of K that initiate even more upstream of these transcription initiation sites, although their frequencies were low. Promot er activity was detected in vitro 5' of J lambda2, with the minimal promote r activity mapping to the region between positions -35 and -120. In additio n, computer analysis allowed the prediction of a nuclear scaffold/matrix at tachment (S/MAR) region between the two J-C gene clusters at each hemi-locu s. This region between the lambda1/lambda3 clusters binds to the nuclear ma trix in vitro, and J-C lambda1 germ-line transcription initiates a short di stance downstream from this S/MAR element. (C) 2001 Elsevier Science Ltd. A ll rights reserved.