Upstream activating sequence

About: Upstream activating sequence is a research topic. Over the lifetime, 1633 publications have been published within this topic receiving 100112 citations.

Hybrid nucleic acid sequences having tandemly arranged genes

Abstract: @ A hybrid nucleic acid sequence for use in producing one or more selected polypeptides in a host cell is provided which comprises: a promoter sequence recognized by the host cell at one end; a transcription terminator sequence recognized by the host cell at the other end; and between the promoter sequence and the transcription terminator sequence: a first region which includes a ribosome-binding site, a translation initiation site, a coding region for one of said one or more selected polypeptides, and a translation termination site. but which does not include the promoter sequence or the transcription terminator sequence; and one or more second regions, each of which includes a ribosome-binding site, a translation initiation site, a coding region for the same or a different one of said one or more selected polypeptides, and a translation termination site, and which does not include the promoter sequence or the transcription terminator sequence, the reading directions of the promoter sequence, the transcription terminator sequence, the first region and each of the second regions having the same orientation.

Promoter activity of the two chicken delta-crystallin genes in a Hela cell extract.

Abstract: The in vitro transcriptional activity of the two delta-crystallin genes (5'-delta 1-delta 2-3') of the chicken was studied in a whole Hela cell extract. Both the delta 1 and delta 2 promoters were recognized by RNA polymerase II in this heterologous system. The major RNA initiation site from the delta 1 promoter was the same in vitro as that which occurs in vivo, as judged by mapping with S1-nuclease, although other minor initiation sites upstream and downstream of the major initiation site were noted. A primer extension experiment showed that the longest RNA synthesized in vitro from a delta 2 template initiated near the beginning of the first exon. The delta 1 promoter was several-fold stronger than that of delta 2 under the present in vitro conditions. Transcription from the delta 1 promoter was abolished by a competitor fragment (c'-II; includes -328 to -63) purified from the delta 2 promoter, indicating that one or more common transcription factors binding upstream from the TATA box are required for in vitro function of the two delta-crystallin promoters. Thus, in the Hela cell extract both delta-crystallin genes contain a functional promoter. We consider the possibility that the single 5'CCAAT3' sequence present in the delta 1 promoter (but lacking in the delta 2 promoter) may contribute to its greater core activity under our conditions. The greater promoter activity of the delta 1-crystallin gene in the Hela cell extract was not sufficient to account for the large ratio of delta 1 to delta 2 mRNA (approximately 50 to 100) in the embryonic chicken lens.

Expression of Caenorhabditis elegans RNA-directed RNA polymerase in transgenic Drosophila melanogaster does not affect morphological development

Abstract: Drosophila melanogaster, along with all insects and the vertebrates, lacks an RdRp gene. We created transgenic strains of Drosophila melanogaster in which the rrf-1 or ego-1 RdRp genes from C. elegans were placed under the control of the yeast GAL4 upstream activation sequence. Activation of the gene was performed by crossing these lines to flies carrying the GAL4 transgene under the control of various Drosophila enhancers. RT-PCR confirmed the successful expression of each RdRp gene. The resulting phenotypes indicated that introduction of the RdRp genes had no effect on D. melanogaster morphological development.

NMR analysis of CYP1(HAP1) DNA binding domain—CYC1 upstream activation sequence interactions: recognition of a CGG trinucleotide and of an additional thymine 5 bp downstream by the zinc cluster and the N-terminal extremity of the protein

Abstract: The DNA binding domain of the yeast transcriptional activator CYP1(HAP1) contains a zinc-cluster structure. The structures of the DNA binding domain-DNA complexes of two other zinc-cluster proteins (GAL4 and PPR1) have been studied by X-ray crystallography. Their binding domains present, besides the zinc cluster, a short linker peptide and a dimerization element. They recognize, as homodimers, two rotationally symmetric CGG trinucleotides, the linker peptide and the dimerization element playing a crucial role in binding specificity. Surprisingly, CYP1 recognizes degenerate forms of a direct repeat, CGGnnnTAnCGGnnnTA, and the role of its linker is under discussion. To better understand the binding specificity of CYP1, we have studied, by NMR, the interaction between the CYP1(55-126) peptide and two DNA fragments derived from the CYC1 upstream activation sequence 1B. Our data indicate that CYP1(55-126) interacts with a CGG and with a thymine 5 bp downstream. The CGG trinucleotide is recognized by the zinc cluster in the major groove, as for GAL4 and PPR1, and the thymine is bound in the minor groove by the N-terminal region, which possesses a basic stretch of arginyl and lysyl residues. This suggests that the CYP1(55-126) N-terminal region could play a role in the affinity and/or specificity of the interaction with its DNA targets, in contrast to GAL4 and PPR1.