Transcription termination at the chicken beta H-globin gene.

Abstract

We characterized the transcription termination region of the chicken 3H.-globin gene. First we located the region by nuclear runon transcription in vitro. Then we sequenced and subcloned it into a chloramphenicol acetyltransferase (CAT) expression vector for assay in vivo. The region of PH termination contains two interesting elements located about 1 kilobase downstream of the pH gene poly(A) site. Either element alone can block CAT expression if inserted between the promoter and the poly(A) site of the cat gene in pRSVcat. The first element in the termination region is an unusually large inverted repeat in the DNA (AG = -71 kcal). The second element, 200 base pairs further downstream, is an RNA polymerase II promoter which directs transcription back upstream on the complementary strand. This transcription converges on and collides with that from the 13H gene at or near the inverted repeat where transcription from both directions stops. We propose that the inverted repeat is a strong pause site which positions the converging polymerases for mutual site-specific termination.Transcription termination signals have been identified and studied extensively both in vivo and in vitro for Escherichia coli RNA polymerase and eucaryotic RNA polymerases I and III (4, 26-28, 37, 44). In E. coli, two main types of termination are recognized, factor dependent and factor independent (44). Factor-dependent termination signals precede the actual sites of termination and exhibit considerable sequence flexibility, which makes them difficult to recognize. A protein factor, usually p, interacts with the RNA to assist termination by the polymerase. Factor-independent terminators, in contrast, have well-defined sequence signals and are readily recognizable by their G+C-rich palindrome, which precedes a cluster of dT residues in the nontemplate strand of the DNA at the site of termination. Additional modes of procaryotic transcription termination also occur and may be of physiological significance. These include transcriptional interference of converging (33) or tandem (1) transcription units and transcriptional arrest caused by blockage of the DNA template by a bound factor (50).In eucaryotes, most RNA polymerase III terminators, like the factor-independent terminators in E. coli, possess a dT cluster and exhibit no obligatory requirement for a factor, but they lack a palindrome (37, 44). Eucaryotic RNA polymerase I termination (27), although factor dependent, bears little resemblance to p-dependent termination in E. coli but instead resembles aspects of the protein blockage mechanism described by Sellitti et al. (50). For polymerase I terminators, it is the DNA to which the factor binds (27), and the signal follows, rather than precedes, the site of termination (27,28).RNA polymerase II appears to incorporate elements of all the above termination mechanisms in its repertoire of strategies. Transcription of the human gastrin gene appears to terminate upstream of the termination signal as for RNA polymerase I (3, 49). The terminator for...

Transcription Termination and 3′-End Processing of the Spliced Leader RNA in Kinetoplastids

“…Transcription termination by pol II has been studied primarily on protein-coding genes (74). The transcription termination signals are bipartite and include the polyadenylation signal (7) in conjunction with a variety of downstream elements, such as poly(T) tracts with intrinsic bent DNA but no adjacent upstream secondary structure (37), trans-acting factors binding to the DNA (4,13,14) or the RNA polymerase (75), or the structure of the transcript (56). Since long (Ͼ6-nt) poly(T) tracts may cause pausing but should not effect pol II termination on protein-coding genes, a different mechanism must exist for mRNA termination in trypanosomatids.…”

Polyadenylation and transcription termination in gene constructs containing multiple tandem polyadenylation signals

“…RNA polymerase II transcription termination occurs hundreds or thousands of nucleotides downstream of polyadenylation sites (7,13,14). Although there have been a number of reports of specific sequences which may act as termination sequences (15)(16)(17)(18)(19)(20)(21)(22)(23)(24)(25), in vivo, transcription termination often depends on 3'-end processing (9)(10)(11)(12)26). Mutations in the conserved polyadenylation AAUAAA signal of the mouse major 3-globin gene or the human globin gene, respectively (11,12), or mutations in both the AAUAAA hexanucleotide and the downstream GT-rich region of the SV40 early polyadenylation site (9) can inhibit polyadenylation as well as transcription termination.…”

RNA polymerase II transcription termination is mediated specifically by protein binding to a CCAAT box sequence.

“…It is possible that sites of RNA (or DNA) secondary structure may also be involved in signalling termination. For example, analysis of the chicken pH-globin gene shows that termination occurs proximal to a region that contains a very strong potential hairpin loop structure (30), suggesting that regions of strong secondary structure, in addition to factor-binding sites, are capable of inducing termination of transcription. Analysis of three other pol II termination sites, the mouse major ,-globin termination element (11), the gastrin gene terminator (2,33), and the histone H2A gene termination region (19), revealed that all are composed of long stretches of AT-rich sequences.…”