Dataset Information

HyCCAPP as a tool to characterize promoter DNA-protein interactions in Saccharomyces cerevisiae.

ABSTRACT: Currently available methods for interrogating DNA-protein interactions at individual genomic loci have significant limitations, and make it difficult to work with unmodified cells or examine single-copy regions without specific antibodies. In this study, we describe a physiological application of the Hybridization Capture of Chromatin-Associated Proteins for Proteomics (HyCCAPP) methodology we have developed. Both novel and known locus-specific DNA-protein interactions were identified at the ENO2 and GAL1 promoter regions of Saccharomyces cerevisiae, and revealed subgroups of proteins present in significantly different levels at the loci in cells grown on glucose versus galactose as the carbon source. Results were validated using chromatin immunoprecipitation. Overall, our analysis demonstrates that HyCCAPP is an effective and flexible technology that does not require specific antibodies nor prior knowledge of locally occurring DNA-protein interactions and can now be used to identify changes in protein interactions at target regions in the genome in response to physiological challenges.

SUBMITTER: Guillen-Ahlers H

PROVIDER: S-EPMC5017017 | biostudies-literature | 2016 Jun

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

HyCCAPP as a tool to characterize promoter DNA-protein interactions in Saccharomyces cerevisiae.

Guillen-Ahlers Hector H Rao Prahlad K PK Levenstein Mark E ME Kennedy-Darling Julia J Perumalla Danu S DS Jadhav Avinash Y L AY Glenn Jeremy P JP Ludwig-Kubinski Amy A Drigalenko Eugene E Montoya Maria J MJ Göring Harald H HH Anderson Corianna D CD Scalf Mark M Gildersleeve Heidi I S HI Cole Regina R Greene Alexandra M AM Oduro Akua K AK Lazarova Katarina K Cesnik Anthony J AJ Barfknecht Jared J Cirillo Lisa A LA Gasch Audrey P AP Shortreed Michael R MR Smith Lloyd M LM Olivier Michael M

Genomics 20160513 6

Currently available methods for interrogating DNA-protein interactions at individual genomic loci have significant limitations, and make it difficult to work with unmodified cells or examine single-copy regions without specific antibodies. In this study, we describe a physiological application of the Hybridization Capture of Chromatin-Associated Proteins for Proteomics (HyCCAPP) methodology we have developed. Both novel and known locus-specific DNA-protein interactions were identified at the ENO ...[more]

PMID: 27184763

Similar Datasets

Project description:BackgroundSaccharomyces cerevisiae is widely studied for production of biofuels and biochemicals. To improve production efficiency under industrially relevant conditions, coordinated expression of multiple genes by manipulating promoter strengths is an efficient approach. It is known that gene expression is highly dependent on the practically used environmental conditions and is subject to dynamic changes. Therefore, investigating promoter activities of S. cerevisiae under different culture conditions in different time points, especially under stressful conditions is of great importance.ResultsIn this study, the activities of various promoters in S. cerevisiae under stressful conditions and in the presence of xylose were characterized using yeast enhanced green fluorescent protein (yEGFP) as a reporter. The stresses include toxic levels of acetic acid and furfural, and high temperature, which are related to fermentation of lignocellulosic hydrolysates. In addition to investigating eight native promoters, the synthetic hybrid promoter P3xC-TEF1 was also evaluated. The results revealed that P TDH3 and the synthetic promoter P3xC-TEF1 showed the highest strengths under almost all the conditions. Importantly, these two promoters also exhibited high stabilities throughout the cultivation. However, the strengths of P ADH1 and P PGK1 , which are generally regarded as 'constitutive' promoters, decreased significantly under certain conditions, suggesting that cautions should be taken to use such constitutive promoters to drive gene expression under stressful conditions. Interestingly, P HSP12 and P HSP26 were able to response to both high temperature and acetic acid stress. Moreover, P HSP12 also led to moderate yEGFP expression when xylose was used as the sole carbon source, indicating that this promoter could be used for inducing proper gene expression for xylose utilization.ConclusionThe results here revealed dynamic changes of promoter activities in S. cerevisiae throughout batch fermentation in the presence of inhibitors as well as using xylose. These results provide insights in selection of promoters to construct S. cerevisiae strains for efficient bioproduction under practical conditions. Our results also encouraged applications of synthetic promoters with high stability for yeast strain development.

Dataset Information

HyCCAPP as a tool to characterize promoter DNA-protein interactions in Saccharomyces cerevisiae.

Publications

HyCCAPP as a tool to characterize promoter DNA-protein interactions in Saccharomyces cerevisiae.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets