Project description:New antifungal drugs are urgently needed due to the currently limited selection, the emergence of drug resistance, and the toxicity of several commonly used drugs. To identify drug leads, we screened small molecules using a Saccharomyces reporter bioassay in which the yeast heterologously expresses Hik1, a group III hybrid histidine kinase (HHK) from Magnaporthe grisea. Group III HHKs are integral in fungal cell physiology, and highly conserved throughout this kingdom; they are absent in mammals, making them an attractive drug target. Our screen identified compounds 13 and 33, which showed robust activity against numerous fungal genera including Candida, Cryptococcus and molds such as Aspergillus and Rhizopus. Drug-resistant Candida from patients were also highly susceptible to compounds 13 and 33. While the compounds do not act directly on HHKs, microarray analysis showed that compound 13 induced transcripts associated with oxidative stress, and compound 33, transcripts linked with heavy metal stress. Both compounds were highly active against Candida biofilm, in vitro and in vivo, and exerted synergy with fluconazole, which was inactive alone. Thus, we identified potent, broad-spectrum antifungal drug leads from a small molecule screen using a high-throughput, yeast reporter bioassay. Two-color experimental design testing the effects of 2 antifungal compounds (13 and 33) after 0, 20, 40 60 min. In the referred publication, the t=20, 40, 60 data was normalized against the t=0 data
Project description:New antifungal drugs are urgently needed due to the currently limited selection, the emergence of drug resistance, and the toxicity of several commonly used drugs. To identify drug leads, we screened small molecules using a Saccharomyces reporter bioassay in which the yeast heterologously expresses Hik1, a group III hybrid histidine kinase (HHK) from Magnaporthe grisea. Group III HHKs are integral in fungal cell physiology, and highly conserved throughout this kingdom; they are absent in mammals, making them an attractive drug target. Our screen identified compounds 13 and 33, which showed robust activity against numerous fungal genera including Candida, Cryptococcus and molds such as Aspergillus and Rhizopus. Drug-resistant Candida from patients were also highly susceptible to compounds 13 and 33. While the compounds do not act directly on HHKs, microarray analysis showed that compound 13 induced transcripts associated with oxidative stress, and compound 33, transcripts linked with heavy metal stress. Both compounds were highly active against Candida biofilm, in vitro and in vivo, and exerted synergy with fluconazole, which was inactive alone. Thus, we identified potent, broad-spectrum antifungal drug leads from a small molecule screen using a high-throughput, yeast reporter bioassay.
Project description:Disruption of alternative splicing frequently causes or contributes to human diseases and disorders. Consequently, there is a need for efficient and sensitive reporter assays capable of screening chemical libraries for compounds with efficacy in modulating important splicing events. Here, we describe a screening workflow employing dual Nano and Firefly luciferase alternative splicing reporters that affords highly efficient, sensitive, and linear detection of small molecule responses. Applying this system to a screen of ~95,000 small molecules, we identify compounds that selectively activate or repress a neuronal microexon network that is frequently disrupted in autism and overexpressed in neuroendocrine cancers. Remarkably, among the most potent and selective activating compounds are histone deacetylase (HDAC) inhibitors. We thus describe a high-throughput screening system for candidate splicing therapeutics, a resource of small molecule modulators of microexons, and insight into the mode of action and potential utility of HDAC inhibitors in the context of neurological disorders.
Project description:Pluripotent stem cells can be induced from somatic cells, providing an unlimited cell resource for regenerative medicine. However, genetic manipulation and difficult-to-manufacture strategies used in reprogramming limit their clinical applications. Here, we show pluripotency can be induced from mouse somatic cells by specific small-molecule compounds. The completely chemically-induced pluripotent stem cells (CiPSCs) can be stably maintained in embryonic stem cell (ESC) culture medium and resemble ESCs in terms of their gene expression profiles, epigenetic status, and potential for differentiation and germline transmission. These findings suggest that exogenous master genes are dispensable for cell fate reprogramming and pave the way for the clinical application of somatic reprogramming techniques. Chemicals' acronym: V, VPA; C, CHIR; 6, 616452; T, tranylcypromine; F, FSK; Z, DZNep; P, PGE2; R, RG108; S, SRT1720; M, 2-Me-5HT; D, D4476; B, Sodium butyrate. mRNA expression analysis of mouse embryonic fibroblasts (MEFs), GFP- cells, GFP+ clusters,GFP+ colonies, embryonic stem cells (ESCs) and CiPSCs by RNA sequencing.
Project description:Pluripotent stem cells can be induced from somatic cells, providing an unlimited cell resource for regenerative medicine. However, genetic manipulation and difficult-to-manufacture strategies used in reprogramming limit their clinical applications. Here, we show pluripotency can be induced from mouse somatic cells by specific small-molecule compounds. The completely chemically-induced pluripotent stem cells (CiPSCs) can be stably maintained in embryonic stem cell (ESC) culture medium and resemble ESCs in terms of their gene expression profiles, epigenetic status, and potential for differentiation and germline transmission. These findings suggest that exogenous master genes are dispensable for cell fate reprogramming and pave the way for the clinical application of somatic reprogramming techniques. Pluripotent stem cells can be induced from somatic cells, providing an unlimited cell resource for regenerative medicine. However, genetic manipulation and difficult-to-manufacture strategies used in reprogramming limit their clinical applications. Here, we show pluripotency can be induced from mouse somatic cells by specific small-molecule compounds. The completely chemically-induced pluripotent stem cells (CiPSCs) can be stably maintained in embryonic stem cell (ESC) culture medium and resemble ESCs in terms of their gene expression profiles, epigenetic status, and potential for differentiation and germline transmission. These findings suggest that exogenous master genes are dispensable for cell fate reprogramming and pave the way for the clinical application of somatic reprogramming techniques. Chemicals' acronyms: V, VPA; C, CHIR; 6, 616452; T, tranylcypromine; F, FSK; Z, DZNep; P, PGE2; R, RG108; S, SRT1720; M, 2-Me-5HT; D, D4476; B, Sodium butyrate.
Project description:This study aimed to construct a rapid screening system to identify novel anti-TB agents from a library of small-molecule compounds.