During chemically induced differentiation of Friend virus-infected mouse erythroleukemia (MEL) cell lines, there is a biphasic down-regulation of the c-myb proto-oncogene. A plasmid containing a murine c-myb cDNA controlled by a mouse metallothionein I promoter was transfected into the C19 MEL cell line. For six transfected clones, it was found that expression of the exogenous c-myb mRNA could be up-regulated by the addition of 120 FIM ZnCl2 and that the N,N'-hexamethylenebisacetamide-induced differentiation of these transfectants was inhibited in proportion to the level of exogenous c-myb mRNA expression. By adding or removing ZnC12 at different times during the induction process, it was possible to show that up-regulation of exogenous c-myb limited to the first 2 days of induction had little or no effect on differentiation. In contrast, continuous expression of exogenous c-myb beginning at any time during the period of induction blocked further differentiation. These results suggest that during HMBA induction of MEL cells, the early down-regulation of c-myb mRNA is not necessary for terminal differentiation, whereas the down-regulation of c-myb at a later time is necessary.

Recent studies in which cloned immunoglobulin genes were introduced into cultured cells have produced two significant findings. First, the genes are expressed after transfection into lymphoid cells but not non-lymphoid cells. Second, transcription of an immunoglobulin gene requires, in addition to the promoter region, an enhancer element located downstream of the transcription start site. These findings raise the question of whether it is the promoter or the enhancer region that is responsible for the observed cell-type specificity. It has, in fact, been shown that immunoglobulin enhancers function only in lymphoid cells. We show here that the promoter for an immunoglobulin kappa light-chain gene also is strongly specific for lymphoid cells. Our result reemphasizes the importance of promoters relative to enhancers in determining which cells express which genes.

It has recently been shown that the promoter of a K immunoglobulin gene is activated for transcription by a downstream sequence element. Here we mapped the activating element to a resolution of about 20 base pairs by constructing a series of deletions in the cloned K gene. After transfection of each deleted gene into myeloma cells, a transient expression assay was used to measure the level of transcription from the K promoter. We found that the activating element extends through about 200 base pairs and encompasses a region of sequence that is conserved between mouse and human genes. As successively deeper deletions were made into the conserved region from either the 5' or 3' side, the activating ability was lost gradually rather than abruptly. Although several short segments in this region are homologous to sequences in viral enhancers, they did not seem to play a dominant role in the activating effect. We also found that the activating element remained functional when reversed in orientation or when moved upstream of the K gene.As a stem cell develops into a B-lymphocyte, rearrangements of the cellular DNA take place that form complete immunoglobulin genes from the separately encoded variable (V) and constant (C) regions (for a review, see reference 31). In B-lymphocytes that synthesize K light chains, one of many VK regions is joined to a JK segment located upstream of the unique CK region (Fig. 1A). After this rearrangement occurs, RNA transcripts of the complete K gene can be detected. The transcripts initiate from just before the rearranged VK region and proceed through the VK and JK regions, the JK-CK intron, and the CK region.As a developing B-lymphocyte enters the antibody-secreting plasma cell stage, the rate of transcription of the rearranged K gene increases, so that K transcripts may represent 5 to 10% of the total mRNA population (25). However, no transcription of the remaining unrearranged VK regions can be detected (12,19). The question therefore arises of how the cell activates the rearranged VK region for high-level transcription, while leaving the unrearranged VK regions completely inactive. Any explanation must be consistent with the fact that rearrangement does not alter the DNA sequences 5' to the VK region, where the signals for transcription initiation would normally be expected to occur (4,19).A guide to answering the question was provided by the existence of enhancer elements in certain viral genomes. Enhancers are DNA sequences that strongly stimulate transcription from many eucaryotic promoters; they can act from either an upstream or a downstream position on promoters that are thousands of base pairs (bp) away (2,7,15 in about 200 bp (1, 8), the K-activating element was only localized to the last third of the JK-CK intron (22). A logical candidate for its precise position is nrovided by a region of the intron that is strikingly well conserved in sequence among mouse, rabbit, and human genes (6). By constructing a series of K gene deletions and analyzing them in a transientexpression ...

Enhancers are DNA sequences that stimulate transcription from eukaryotic promoters. This stimulatory effect can be exerted over large distances and from a position either 5' or 3' of a promoter. Enhancers have been found in the genomes of many viruses, and in some cellular genes such as those encoding immunoglobulin heavy chain and kappa light chain. An important feature of both viral and cellular enhancers is the ability of each enhancer to stimulate transcription from many promoters other than the one with which it is found associated. However, the question of whether cellular enhancers stimulate their 'own' promoter more efficiently than other promoters has apparently not been investigated. We show here that the kappa light-chain enhancer stimulates a kappa promoter about 20-fold more than it stimulates either the simian virus 40 (SV40) early promoter or a metallothionein (MT) promoter, two promoters that are very sensitive to other enhancers. Similarly, the heavy-chain enhancer stimulates a heavy-chain promoter much more than it stimulates the SV40 and MT promoters. This synergism between immunoglobulin enhancers and promoters might be due to the action of a protein that binds specifically to each of the regulatory elements.