The adenovirus major late arrest site blocks transcription by mammalian RNA polymerase II in vitro downstream of the major late promoter but not the mouse beta-globin promoter. We localized the sequences responsible for anti-arrest to the 5' end of the beta-globin transcript and demonstrated that anti-arrest required that this region of RNA form base pairs with the nascent transcript upstream of the arrest site. Small antisense RNA or DNA oligonucleotides hybridizing upstream of the arrest site also prevented arrest when added in trans. Our results suggest that arrest is accompanied by retraction of the nascent transcript into the interior of the polymerase and that hybridization of the transcript prevents this movement, thereby allowing the polymerase to continue elongation.

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We report the mapping, sequencing, and study of the physiological role of the fourth arabinose-inducible operon from Escherichia coli, araJ. It is located at 9 min on the chromosome and codes for a single 42-kDa protein that shows no significant homology to other known proteins. Destruction of the chromosomal araJ gene does not detectably affect either of the two arabinose transport systems, the ability of cells to grow The enteric bacterium Escherichia coli can utilize L-arabinose as a sole carbon source. In the presence of arabinose, expression from the three known ara operons is induced up to 300-fold. These are the araBAD operon (5, 6), which codes for proteins involved in the catabolism of arabinose, and the araE (27) and araFGH (2, 17, 31) operons, which code for the low-and high-affinity arabinose transport systems, respectively: In addition to the promoters of these three well-characterized operons, there exists a fourth promoter in E. coli that is induced by arabinose. This promoter was originally cloned by Kosiba and Schleif (18) and at that time was misidentified as the araFGH promoter. Subsequent work (11, 13) has identified the actual araFGH promoter, and the arabinose-inducible promoter cloned by Kosiba and Schleif (18) is now known as pJ. Recent work (11,18) has shown that arabinose induces substantial transcription from pJ in vivo and in vitro. In vivo, as well as in vitro (11,18), transcription from pJ is dependent upon the arabinosespecific transcriptional regulator protein AraC and the global catabolite gene activator protein CAP, as is transcription from the araBAD, araE, and araFGH operons (7,8,32).Since all known ara mutations are confined to the araBAD, araC, araE, and araFGH operons, we were interested to learn whether pJ drives an intact gene and whether its product plays any detectable role in vivo. In particular, we wished to determine whether the araJ product was a fourth component in the high-affinity arabinose transport system, since most analogous systems contain four protein components (1, 34).Here we report the mapping of the araJ gene to 9 min on the E. coli chromosome and the cloning and sequencing of the araJ gene and surrounding regions of the chromosome. We also show that insertion/deletion mutations in araJ have no detectable effect on the ability of the bacteria to grow on arabinose and to induce the araBAD operon. MATERIALS AND METHODSMedia, strains, and general methods. Media and general methods have been described (21,33 Kosiba and Schleif (18) containing the araJ promoter. Three positive clones were selected and shown by restriction mapping to contain overlapping copies of the same piece of genomic DNA; these three clones were sent to us for further analysis.One of the lambda clones was digested with PvuI, and the 1.9-and 2.5-kb fragments that contained pJ and flanking sequences were isolated. The fragments were filled out with DNA polymerase (Klenow fragment) and then ligated into 7765 JOURNAL OF BACTERIOLOGY, Dec. 1991, p. 7765-7771