Functional Coupling of a Nematode Chemoreceptor to the Yeast Pheromone Response Pathway

Tehseen, Muhammad; Dumancic, Mira M.; Briggs, Lyndall J.; Wang, Jian; Berna, Amalia Z.; Anderson, Alisha; Trowell, Stephen

doi:10.1371/journal.pone.0111429

Cited by 6 publications

(3 citation statements)

References 58 publications

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“…Tehseen et al demonstrated that the Caenorhabditis elegans olfactory GPCR ODR-10 was functionally expressed in yeast by using chimeric Gpa1/C. elegans Gα [97]. Mukherjee et al constructed a medium-chain fatty acid biosensor by using the olfactory receptor OR1G1 that functionally expressed in yeast [98].…”

Section: Peripheral Membrane Proteinsmentioning

confidence: 99%

Biosensing Techniques in Yeast: G-Protein Signaling and Protein-Protein Interaction Assays for Monitoring Ligand Stimulation and Oligomer Formation of Heterologous GPCRs

Nakamura¹,

Kondo²,

Ishii³

2018

Peripheral Membrane Proteins

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Guanine nucleotide-binding proteins (G-proteins) act as transducers of external stimuli for intracellular signaling, and control various cellular processes in cooperation with seven transmembrane G-protein-coupled receptors (GPCRs). Because GPCRs constitute the largest family of eukaryotic membrane proteins and enable the selective recognition of a diverse range of molecules (ligands), they are the major molecular targets in pharmaceutical and medicinal fields. In addition, GPCRs have been known to form heteromers as well as homomers, which may result in vast physiological diversity and provide opportunities for drug discovery. G-proteins and their signal transduction machinery are universally conserved in eukaryotes; thereby, the yeast Saccharomyces cerevisiae has been used to construct artificial in vivo GPCR biosensors. In this chapter, we focus on the yeast-based GPCR biosensors that can detect ligand stimulation and oligomer formation, and summarize their techniques using the G-protein signaling and protein-protein interaction assays.

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Section: Peripheral Membrane Proteinsmentioning

confidence: 99%

Biosensing Techniques in Yeast: G-Protein Signaling and Protein-Protein Interaction Assays for Monitoring Ligand Stimulation and Oligomer Formation of Heterologous GPCRs

Nakamura¹,

Kondo²,

Ishii³

2018

Peripheral Membrane Proteins

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“…A technique that is capable of onsite VOC collection and detection would allow law enforcement agencies to rapidly assess potentially hazardous conditions and aid investigators in analysing VOCs in laboratories. One potential method for onsite detection is the use of biosensors that are capable of detecting targets in the liquid phase, and thus, a system that collects VOCs from a liquid substrate could be a potential method of onsite analysis [20,21].…”

Section: Introductionmentioning

confidence: 99%

Application of a Microfluidic Gas-to-Liquid Interface for Extraction of Target Amphetamines and Precursors from Air Samples

et al. 2020

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The investigation of clandestine laboratories poses serious hazards for first responders, emergency services, investigators and the surrounding public due to the risk of exposure to volatile organic compounds (VOCs) used in the manufacture of illicit substances. A novel gas sampling interface using open microfluidic channels that enables the extraction of VOCs out of the gas phase and into a liquid, where it can be analysed by conventional detection systems, has recently been developed. This paper investigates the efficiency and effectiveness of such a gas-to-liquid (GTL) extraction system for the extraction of amphetamine-type substances (ATS) and their precursors from the vapour phase. The GTL interface was evaluated across a range of different ATS and their precursors (methamphetamine, dimethylamphetamine, N-formylmethamphetamine, benzaldehyde, phenyl-2-propanone, ephedrine and pseudoephedrine) at concentrations ranging between 10 and 32 mg m−3. These gas samples were produced by a gas generation system directly in Tedlar® bags and gas canisters for controlled volume sampling. When using gas sampled from Tedlar® bags, four of the seven compounds were able to be extracted by the GTL interface, with the majority of the VOCs having extraction yields between 0.005% and 4.5%, in line with the results from an initial study. When samples were taken from gas canisters, only benzaldehyde was able to be detected, with extraction efficiencies between 0.2% and 0.4%. A custom-built mount for the GTL interface helped to automate the extraction process, with the aim of increasing extraction efficiency or reducing variability. However, the extraction efficiency did not improve when using this accessory, but the procedure did become more efficient. The results from the study indicated that the GTL interface could be employed for the collection of gaseous ATS and incorporated into mobile detection systems for onsite collection and analysis of volatile compounds related to ATS manufacture.

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“…Yeast cultures, which can be grown rapidly and at low cost, are ideal for use in automated screens [ 8 – 11 ]. Yeast cells are suitable hosts for the expression of nematode proteins [ 12 – 18 ], including enzymes essential for different life-cycle stages of the parasites, many of which cannot be propagated in vitro [ 17 ]. We engineered Saccharomyces cerevisiae strains to express either different parasite drug targets [ 9 ], or their equivalent human proteins, such that the growth of the yeast is dependent on the functioning of these heterologous proteins.…”

Section: Introductionmentioning

confidence: 99%

Yeast-Based High-Throughput Screens to Identify Novel Compounds Active against Brugia malayi

et al. 2016

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BackgroundLymphatic filariasis is caused by the parasitic worms Wuchereria bancrofti, Brugia malayi or B. timori, which are transmitted via the bites from infected mosquitoes. Once in the human body, the parasites develop into adult worms in the lymphatic vessels, causing severe damage and swelling of the affected tissues. According to the World Health Organization, over 1.2 billion people in 58 countries are at risk of contracting lymphatic filariasis. Very few drugs are available to treat patients infected with these parasites, and these have low efficacy against the adult stages of the worms, which can live for 7–15 years in the human body. The requirement for annual treatment increases the risk of drug-resistant worms emerging, making it imperative to develop new drugs against these devastating diseases.Methodology/Principal FindingsWe have developed a yeast-based, high-throughput screening system whereby essential yeast genes are replaced with their filarial or human counterparts. These strains are labeled with different fluorescent proteins to allow the simultaneous monitoring of strains with parasite or human genes in competition, and hence the identification of compounds that inhibit the parasite target without affecting its human ortholog. We constructed yeast strains expressing eight different Brugia malayi drug targets (as well as seven of their human counterparts), and performed medium-throughput drug screens for compounds that specifically inhibit the parasite enzymes. Using the Malaria Box collection (400 compounds), we identified nine filarial specific inhibitors and confirmed the antifilarial activity of five of these using in vitro assays against Brugia pahangi.Conclusions/SignificanceWe were able to functionally complement yeast deletions with eight different Brugia malayi enzymes that represent potential drug targets. We demonstrated that our yeast-based screening platform is efficient in identifying compounds that can discriminate between human and filarial enzymes. Hence, we are confident that we can extend our efforts to the construction of strains with further filarial targets (in particular for those species that cannot be cultivated in the laboratory), and perform high-throughput drug screens to identify specific inhibitors of the parasite enzymes. By establishing synergistic collaborations with researchers working directly on different parasitic worms, we aim to aid antihelmintic drug development for both human and veterinary infections.

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Functional Coupling of a Nematode Chemoreceptor to the Yeast Pheromone Response Pathway

Cited by 6 publications

References 58 publications

Biosensing Techniques in Yeast: G-Protein Signaling and Protein-Protein Interaction Assays for Monitoring Ligand Stimulation and Oligomer Formation of Heterologous GPCRs

Biosensing Techniques in Yeast: G-Protein Signaling and Protein-Protein Interaction Assays for Monitoring Ligand Stimulation and Oligomer Formation of Heterologous GPCRs

Application of a Microfluidic Gas-to-Liquid Interface for Extraction of Target Amphetamines and Precursors from Air Samples

Yeast-Based High-Throughput Screens to Identify Novel Compounds Active against Brugia malayi

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