Dataset Information

The Sulfolobus initiator element is an important contributor to promoter strength.

ABSTRACT: Basal elements in archaeal promoters, except for putative initiator elements encompassing transcription start sites, are well characterized. Here, we employed the Sulfolobus araS promoter as a model to study the function of the initiator element (Inr) in archaea. We have provided evidence for the presence of a third core promoter element, the Sulfolobus Inr, whose action depends on a TATA box and the TFB recognition element (BRE). Substitution mutations in the araS Inr did not alter the location of the transcription start site. Using systematic mutagenesis, the most functional araS Inr was defined as +1 GAGAMK +6 (where M is A/C and K is G/T). Furthermore, WebLogo analysis of a subset of promoters with coding sequences for 5' untranslated regions (UTRs) larger than 4 nucleotides (nt) in Sulfolobus solfataricus P2 identified an Inr consensus that exactly matches the functional araS Inr sequence. Moreover, mutagenesis of 3 randomly selected promoters confirmed the Inr sequences to be important for basal promoter strength in the subgroup. Importantly, the result of the araS Inr being added to the Inr-less promoters indicates that the araS Inr, the core promoter element, is able to enhance the strength of Inr-less promoters. We infer that transcription factor B (TFB) and subunits of RNA polymerase bind the Inr to enhance promoter strength. Taken together, our data suggest that the presence or absence of an Inr on basal promoters is important for global gene regulation in Sulfolobus.

SUBMITTER: Ao X

PROVIDER: S-EPMC3811598 | biostudies-literature | 2013 Nov

REPOSITORIES: biostudies-literature

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The Sulfolobus initiator element is an important contributor to promoter strength.

Ao Xiang X Li Yingjun Y Wang Fan F Feng Mingxia M Lin Yanxu Y Zhao Shumiao S Liang Yunxiang Y Peng Nan N

Journal of bacteriology 20130913 22

Basal elements in archaeal promoters, except for putative initiator elements encompassing transcription start sites, are well characterized. Here, we employed the Sulfolobus araS promoter as a model to study the function of the initiator element (Inr) in archaea. We have provided evidence for the presence of a third core promoter element, the Sulfolobus Inr, whose action depends on a TATA box and the TFB recognition element (BRE). Substitution mutations in the araS Inr did not alter the location ...[more]

PMID: 24039266

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Project description:Trichoderma reesei can produce up to 100 g/liter of extracellular proteins. The major and industrially relevant products are cellobiohydrolase I (CBHI) and the hemicellulase XYNI. The genes encoding both enzymes are transcriptionally activated by the regulatory protein Xyr1. The first 850 nucleotides of the cbh1 promoter contain 14 Xyr1-binding sites (XBS), and 8 XBS are present in the xyn1 promoter. Some of these XBS are arranged in tandem and others as inverted repeats. One such cis element, an inverted repeat, plays a crucial role in the inducibility of the xyn1 promoter. We investigated the impact of the properties of such cis elements by shuffling them by insertion, exchange, deletion, and rearrangement of cis elements in both the cbh1 and xyn1 promoter. A promoter-reporter assay using the Aspergillus nigergoxA gene allowed us to measure changes in the promoter strength and inducibility. Most strikingly, we found that an inverted repeat of XBS causes an important increase in cbh1 promoter strength and allows induction by xylan or wheat straw. Furthermore, evidence is provided that the distances of cis elements to the transcription start site have important influence on promoter activity. Our results suggest that the arrangement and distances of cis elements have large impacts on the strength of the cbh1 promoter, whereas the sheer number of XBS has only secondary importance. Ultimately, the biotechnologically important cbh1 promoter can be improved by cis element rearrangement.IMPORTANCE In the present study, we demonstrate that the arrangement of cis elements has a major impact on promoter strength and inducibility. We discovered an influence on promoter activity by the distances of cis elements to the transcription start site. Furthermore, we found that the configuration of cis elements has a greater effect on promoter strength than does the sheer number of transactivator binding sites present in the promoter. Altogether, the arrangement of cis elements is an important factor that should not be overlooked when enhancement of gene expression is desired.

Dataset Information

The Sulfolobus initiator element is an important contributor to promoter strength.

Publications

The Sulfolobus initiator element is an important contributor to promoter strength.

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