| Autor: |
Reveal Brad S, Paietta John V |
| Jazyk: |
angličtina |
| Rok vydání: |
2012 |
| Předmět: |
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| Zdroj: |
BMC Research Notes, Vol 5, Iss 1, p 339 (2012) |
| Druh dokumentu: |
article |
| ISSN: |
1756-0500 |
| DOI: |
10.1186/1756-0500-5-339 |
| Popis: |
Abstract Background Cystathionine γ-lyase plays a key role in the transsulfuration pathway through its primary reaction of catalyzing the formation of cysteine from cystathionine. The Neurospora crassa cystathionine γ-lyase gene (cys-16+) is of particular interest in dissecting the regulation and dynamics of transsulfuration. The aim of this study was to determine the regulatory connection of cys-16+ to the Neurospora sulfur regulatory network. In addition, the cys-16+ promoter was characterized with the goal of developing a strongly expressed and regulatable gene expression tool. Findings The cystathionine γ-lyase cys-16+ gene was cloned and characterized. The gene, which contains no introns, encodes a protein of 417 amino acids with conserved pyridoxal 5’-phosphate binding site and substrate-cofactor binding pocket. Northern blot analysis using wild type cells showed that cys-16+ transcript levels increased under sulfur limiting (derepressing) conditions and were present only at a low level under sulfur sufficient (repressing) conditions. In contrast, cys-16+ transcript levels in a Δcys-3 regulatory mutant were present at a low level under either derepressing or repressing conditions. Gel mobility shift analysis demonstrated the presence of four CYS3 transcriptional activator binding sites on the cys-16+ promoter, which were close matches to the CYS3 consensus binding sequence. Conclusions In this work, we confirm the control of cystathionine γ-lyase gene expression by the CYS3 transcriptional activator through the loss of cys-16+ expression in a Δcys-3 mutant and through the in vitro binding of CYS3 to the cys-16+ promoter at four sites. The highly regulated cys-16+ promoter should be a useful tool for gene expression studies in Neurospora |
| Databáze: |
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| Externí odkaz: |
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