Conserved cis-elements bind a protein complex that regulates Drosophila ras2/rop bidirectional expression

Abstract

Summary The Drosophila ras2 promoter region exhibits bidirectional activity, as has been demonstrated for the human c-Ha-rasl and the mouse c-Ki-ras. Here we address a unique case of ras regulation, as Drosophila ras2 provides the only example to date in which the flanking gene (rop) and its product have been isolated. A linking mechanism of control suggests a mutual interaction between the two gene products. Our studies indicate that the Drosophila ras2 promoter region shares with the human c-Ha-rasl promoter a CACCC box and an AP-l-like sequence. A 14 bp promoter fragment which holds a CACCC element is demonstrated to interact with a specific transcription factor (factor B). This CACCC promoter element represents a stretch of imperfect palindrome. We present evidence that this factor can form a complex with another specific DNA-binding protein (factor A). The binding sites (A + B) for these protein factors are essential for 95% expression of both genes flanking the promoter (ras2 and rop). Region A consists of four overlapping consensus sequences: a TATA-like element, a DSE-like motif (the core sequence of the serum response element), a DRE octamer, which has been shown to play a role in cell proliferation, and a 5 bp direct repeat representing the GATA consensus sequence. Factor A has a very weak affinity to the full promoter region, but when complexed with factor B binding efficiency is enhanced. We also show that alterations of DNA-protein binding specificities can be achieved by supplementing the growth media with different sera.

beta2 knockout mice develop parenchymal iron overload: A putative role for class I genes of the major histocompatibility complex in iron metabolism.

“…1B). Similar genomic structures have been seen in many other gene systems (8)(9)(10)(11)(12)(13). In addition, a plasmid containing a TL and HEPH gene and the ,BGAP motif has been shown to express both genes after being transfected into various cell lines (7).…”

Over‐expression of DREF in the Drosophila wing imaginal disc induces apoptosis and a notching wing phenotype

“…Experiments are now underway to address this point. In addition, it has been reported that the promoter of the Drosophila ras2 gene also contains the DRE-like sequence (Lightfoot et al 1994), suggesting that this is another candidate DREF-target gene. Moreover, we searched the Drosophila genome project database for DRE-like sequences and found them in the promoter region of the Dsor1 gene, a Drosophila homologue of mammalian MEK (Tsuda et al 1993).…”

“…Promoter regions of DNA replication-and proliferation-related genes such as those for the DNA polymerase a 180 and 73 kDa subunits, PCNA, dE2F, cyclin A, D-ras2 and D-raf, contain a DNA replication-related element (DRE), and the requirement of this DRE for promoter activity has been confirmed in both cultured cells (Hirose et al 1993;Lightfoot et al 1994;Ohno et al 1996;Takahashi et al 1996;Ryu et al 1997) and transgenic flies (Yamaguchi et al 1995Sawado et al 1998). Moreover, a specific DRE-binding factor (DREF) has been identified as a homodimer of 80 kDa polypeptide (Hirose et al 1993) and its cDNA has been cloned .…”

“…However, by searching the Drosophila genome database we found DRE-like sequences in the 5 H -flanking regions of many genes other than DNA replicationand proliferation-related genes, suggesting that DREF plays multiple roles in vivo. While the effect of an over-expression of CR1, which is a dominant negative form of DREF in imaginal discs (Hirose et al 1999), and the effect of the mutation in the DRE on the promoter activity in cultured cells have been studied extensively (Hirose et al 1993;Lightfoot et al 1994;Ohno et al 1996;Takahashi et al 1996;Yamaguchi et al 1996;Ryu et al 1997;Sawado et al 1998), the consequences of elevating DREF activity in developing tissues have hitherto remained unclear.…”

Differential expression during embryogenesis of three genes clustered in the Ras1 region of Drosophila melanogaster

“…The putative promoters regulating the divergent transcription of these two genes should be included in this region, which contains the octamer TATCGATA. This octamer is also found within a box required for the full activity of the bidirectional promoter regulating the divergent transcription of the Ras2 and Rop genes (Cohen et al, 1988), in both directions (Lev et al, 1989;Lightfoot et al, 1994). Recently this octamer has been (lanes 1-4, 7, and 8 ) or embryos (lanes 5, 6, 9-1 1) were hybridized with probes 1 to 8, respectively Lane 9 was hybridized with a probe prepared from Rlcl coding sequence Lanes 10 and 11 were hybridized with probes prepared from cDNAs of Rlbl and Rasl, respectively termed DRE (for DNA replication-related element), since it was found as an essential element in the promoters of two Drosophila genes encoding proteins associated with DNA replication, DNA polymerase a, and the proliferating cell nuclear antigen (Hirose et al, 1993(Hirose et al, , 1994.…”