The correct spatial expression of two Drosophila bithorax complex (BX-C) genes, abdominal-A (abdA) and Abdominal-B (AbdB), is dependent on the 100-kb intergenic infraabdominal (iab) region. The iab region is known to contain a number of different domains (iab2 through iab8) that harbor cis-regulatory elements responsible for directing expression of abdA and AbdB in the second through eighth abdominal segments. Here, we use in situ hybridization to perform high-resolution mapping of the transcriptional activity in the iab control regions. We show that transcription of the control regions themselves is abundant and precedes activation of the abdA and AbdB genes. As with the homeotic genes of the BX-C, the transcription patterns of the RNAs from the iab control regions demonstrate colinearity with the sequence of the iab regions along the chromosome and the domains in the embryo under the control of the specific iab regions. These observations suggest that the intergenic RNAs may play a role in initiating cis regulation at the BX-C early in development.

Insulator DNAs and promoter competition regulate enhancerpromoter interactions within complex genetic loci. Here we provide evidence for a third mechanism: promoter-proximal tethering elements. The Scr-ftz region of the Antennapedia gene complex includes two known enhancers, AE1 and T1. AE1 selectively interacts with the ftz promoter to maintain pair-rule stripes of ftz expression during gastrulation and germ-band elongation. The T1 enhancer, located 3 of the ftz gene and Ϸ25 kb 5 of the Scr promoter, selectively activates Scr expression in the prothorax and posterior head segments. A variety of P element minigenes were examined in transgenic embryos to determine the basis for specific AE1-ftz and T1-Scr interactions. A 450-bp DNA fragment located Ϸ100 bp 5 of the Scr transcription start site is essential for T1-Scr interactions and can mediate long-range activation of a ftz͞lacZ reporter gene when placed 5 of the ftz promoter. We suggest that the Scr450 fragment contains tethering elements that selectively recruit T1 to the Scr promoter. Tethering elements might regulate enhancer-promoter interactions at other complex genetic loci. P revious studies have identified two different mechanisms for regulating enhancer-promoter interactions within complex genetic loci: promoter competition and insulator DNAs (for reviews, see refs. 1 and 2). In the case of competition, a shared enhancer selectively interacts with the strongest of linked promoters (3, 4). This preferred interaction sequesters the enhancer so it is unavailable for interacting with weaker promoters. Insulator DNAs selectively block the interaction of a distal enhancer with a target promoter when the insulator is positioned between the two (reviewed in ref. 5). This block does not interfere with the activation of proximal genes by the same enhancer. Here we present evidence for a third mechanism of regulating enhancer-promoter interactions: promoter-proximal tethering elements.The Antennapedia gene complex (ANT-C) is one of the two major Hox gene clusters in the Drosophila genome (summarized in Fig. 1). It is Ϸ500 kb in length and contains nine homeobox genes, including five homeotic selector genes that pattern the head and thorax (6, 7). The Scr selector gene is expressed in the anterior compartment of the first thoracic segment, as well as portions of the labial and maxillary head segments. This complex pattern of Scr expression depends, at least in part, on a distal enhancer, T1, that is located Ϸ25 kb 5Ј of the Scr promoter (8). T1 is located downstream of the pair-rule gene ftz, which exhibits a seven-stripe pattern of expression distinct from the Scr gene. The ftz pattern is regulated by an intergenic enhancer, AE1, located between the divergently transcribed Scr and ftz genes (9, 10). Specific T1-Scr and AE1-ftz interactions are essential for the normal patterning of the early embryo. Misexpression of either gene disrupts segmentation and causes embryonic lethality (11,12).Previous studies have shown that AE1 prefers TATAcontaining promoters (13)....

Long-range enhancer-promoter interactions are commonly seen in complex genetic loci such as Hox genes and globin genes. In the case of the Drosophila Antennapedia complex, the T1 enhancer bypasses the neighboring ftz gene and interacts with the distant Scr promoter to activate expression in posterior head segments. Previous studies identified a 450-bp promoter-proximal sequence, the tethering element, which is essential for T1-Scr interactions. To obtain a more comprehensive view of how individual enhancers selectively interact with appropriate target genes, we used bioinformatic methods to identify new cis-regulatory DNAs in the Ϸ50-kb Scr-Antp interval. Three previously uncharacterized regulatory elements were identified: a distal T1 tethering sequence mapping >40 kb from the proximal tethering sequence, a repressor element that excludes activation of Scr by inappropriate enhancers, and a new ftz enhancer that directs expression within the limits of stripes 1 and 5. Many of the regulatory DNAs in the Scr-Antp interval are transcribed, including the proximal and distal tethering elements. We suggest that homotypic interactions between the tethering elements stabilize long-range T1-Scr interactions during development.E nhancers direct localized stripes, bands, and tissue-specific patterns of gene expression in the early Drosophila embryo (1). They are typically 300 bp to 1 kb in length and contain clustered binding sites for both transcriptional activators and repressors. Enhancers usually activate nearby target genes, although there are examples where they ignore the most proximal promoters and interact with distantly linked genes. Examples include the 3Ј enhancers of the dpp gene and the T1 enhancer of Scr.The dpp enhancers fail to activate the neighboring slh and oaf genes but instead activate the expression of the distal dpp gene in imaginal disks (2). The selective regulation of dpp expression appears to depend on promoter specificity (3, 4). The oaf and slh promoters are incompatible for activation by the dpp enhancers, despite the fact that they map much closer than does the preferred dpp promoter. Similarly, the distal T1 enhancer jumps over the intervening ftz gene to activate Scr in posterior head segments (5). The failure of the T1 enhancer to activate ftz might also depend on promoter specificity. The T1 enhancer only weakly activates a minimal ftz-lacZ fusion gene, despite the fact that it contains a strong TATA element. However, the possible incompatibility between T1 and the ftz promoter is not sufficient to account for selective T1-Scr interactions, because T1 also fails to activate a Scr-lacZ fusion gene containing the minimal Scr core promoter. We have previously identified a 450-bp tethering element that maps immediately 5Ј of the Scr core promoter (6). This element is essential for T1-Scr interactions and is sufficient to mediate long-range T1-ftz interactions when placed immediately 5Ј of the ftz promoter (Fig. 1).In the present study, we have conducted a systematic analysis of cis-regulator...

Spitz et al (2003[this issue of Cell]) describe the properties of a novel cis-regulatory DNA element, the global control region (GCR), which regulates gene expression over distances of several hundred kilobases at the mouse HoxD complex. The GCR provides an explanation for the colinear genetic linkage and expression of individual Hox genes within developing limbs.