Genetic sensors capable of converting key metabolite levels to fluorescence signals enable the monitoring of intracellular compound concentrations in living cells， and emerge as an efficient tool in high-throughput genetic screening. However， the development of genetic sensors in yeasts lags far behind their development in bacteria. Here we report the design of a malonyl-CoA sensor in Saccharomyces cerevisiae using an adapted bacterial transcription factor FapR and its corresponding operator fapO to gauge intracellular malonyl-CoA levels. By combining this sensor with a genome-wide overexpression library， we identified two novel gene targets that improved intracellular malonyl-CoA concentration. We further utilized the resulting recombinant yeast strain to produce a valuable compound， 3-hydroxypropionic acid， from malonyl-CoA and enhanced its titer by 120%. Such a genetic sensor provides a powerful approach for genome-wide screening and could further improve the synthesis of a large range of chemicals derived from malonyl-CoA in yeast.