Screening for molecules that inhibit widespread bacterial transcription factors

• Grant number: 24/16116-2
• Support Opportunities: Scholarships in Brazil - Scientific Initiation
• Start date: October 01, 2024
• End date: September 30, 2025
• Field of knowledge: Biological Sciences - Genetics - Molecular Genetics and Genetics of Microorganisms
• Principal Investigator: Marilis Do Valle Marques
• Grantee: Sylvia Ellen Kempenich
• Host Institution: Instituto de Ciências Biomédicas (ICB). Universidade de São Paulo (USP). São Paulo , SP, Brazil
• Associated research grant: 21/10577-0 - Biology of Bacteria and Bacteriophages Research Center, AP.CEPID

Abstract

Transcription factors (TF) are usually considered to be poorly druggable, but this perception stems from the fact that eukaryotic TFs are predominantly intrinsically disordered proteins, which are indeed harder to inhibit with small molecules (10.1038/s41573-021-00199-0). Bacterial TFs, on the other hand, are structured and naturally regulated by small molecule allosteric modulators. The existence of anti-inducers of the Lac repressor (10.1021/bi00679a024) is an indication that we may be able to find small molecule inhibitors of TF function. We will test this proposition by searching for inhibitors of two widespread TFs, FUR and XylR. The intracellular iron concentration needs to be kept under strict regulation, since it is essential as enzyme cofactor and at the same time is a source of oxidative stress (Andrews et al. 2003; Leaden et al., 2018). Many pathogenic bacteria use siderophores and other transporters to obtain iron from the hosts (Schaible and Kaufmann 2004). The main regulatory factor of bacterial iron homeostasis is FUR (Ferric Uptake Regulator), which when bound to Fe2+ binds to specific operators in the promoter regions of its target genes, inhibiting gene expression (Lee and Hellmann, 2007). The complete Caulobacter FUR regulon has been identified (da Silva Neto et al., 2009), and includes the hutA gene encoding a hemin transporter (Balhesteros et al., 2017). Caulobacter and other bacteria can utilize D-xylose as a carbon source, and the xylose utilization operon (xylA) is repressed by the XylR regulator, a member of the LacI family of transcription factors (Meisenzahl et al., 1997; Stephens et al., 2007). We will use the hutA and xylA promoters to construct gene fusions regulated by FUR and XylR, respectively. Each reporter strain will then be used in screens against commercial small molecule libraries to identify compounds that interfere with regulation. The identification of anti-inducers in small molecule libraries will support the notion that bacterial TFs are amenable drug targets.