Protection of expressed immunoglobulin genes against nuclease cleavage

Abstract

Fragmentation of the actively transcribed kappa immunoglobulin gene in mouse myeloma nuclei with micrococcal nuclease and the restriction nuclease Bsp RI reveals a chromatin structure without the regularity of repeating nucleosomes found in bulk chromatin. Such regularity is restored about 2.2 kb 3' of the coding region. An only moderately increased micrococcal nuclease sensitivity and a 65% average protection of the Bsp RI sites indicates a DNA-protein interaction in the transcribed region which is not very different from that of an inactive gene. As determined by indirect endlabeling the frequency of Bsp RI cleavage both, after very mild and exhaustive digestion, varied moderately from site to site along the gene. In addition, it was not in each case the same at analogous sites on both alleles which are both transcribed. Thus, the experiments demonstrate differences between the chromatin structures of the genes which may be related to regulatory phenomena and thereby corroborate earlier findings made with DNAase I.

Nucleosome distribution on the Jkand Ckimmunoglobulin gene segments of mouse liver chromatin

“…Another explanation of the increased cleavage may arise from a sequence specific binding of proteins in part-of the nuclei near sites 4+5 and 10 which passively organize the nucleosomes of their immediate neighborhood. This idea gains support from the increased cleavage found at site 4 in active chromatin in which we did not find evidence for regular nucleosomes (10). Moreover, the latter location coincides with or is very close to a DNAase I hypersensitive site found in immunocompetent cells (19,30).…”

“…19. The DNA probes and the marker fragments used in electrophoreses are described in the preceding paper (10). For quantitation of band intensities low density exposures of the autoradiographs were scanned with a Shimadzu CS-910 densitometer and the peak areas were evaluated.…”

Micrococcal nuclease digests intranucleosomal DNA of inactive genes more rapidly than active genes

“…Relative resistance of active genes. perhaps to single-stranded nicking in this case, has been reported before (Weischet et a/., 1983). and may reflect an altered nucleosomal conformation, analogous to 'smooth'…”

Chromatin structure of the developmentally regulated early histone genes of the sea urchinStrongylocentrotus purpuratus

“…Transcribed (3-and a-globin genes in induced murine erythroleukemia cells have been reported to exhibit regular nucleosomal organization (9), but showed no evidence of nucleosomal organization in another study (37). Transcribed Ig x genes exhibit no evidence of nucleosomal packaging (38), although weak core-and dinucleosome-sized bands have been observed (39). Upon transcriptional activation, D. melanogaster hsp70 genes (11), the Chironomus tentans secretory protein gene (40), and the 5' flanking region of the hen ovalbumin gene (10) lose nucleosomal packaging.…”