Set1-mediated histone H3K4 methylation is required for azole induction of the ergosterol biosynthesis genes and antifungal drug resistance inCandida glabrata

Abstract

Candida glabrata is an opportunistic pathogen that has developed the ability to adapt and thrive under azole treated conditions. The common mechanisms that can result in Candida drug resistance are due to mutations or overexpression of the drug efflux pump or the target of azole drugs, Cdr1 and Erg11, respectively. However, the role of epigenetic histone modifications in azole-induced gene expression and drug resistance are poorly understood in C. glabrata. In this study, we show for the first time that Set1 mediates histone H3K4 mono-, di-, and trimethylation in C. glabrata. In addition, loss of SET1 and histone H3K4 methylation results in increased susceptibility to azole drugs in both C. glabrata and S. cerevisiae. Intriguingly, this increase in susceptibility to azole drugs in strains lacking Set1-mediated histone H3K4 methylation is not due to altered transcript levels of CDR1, PDR1 or Cdr1’s ability to efflux drugs. Genome-wide transcript analysis revealed that Set1 is necessary for azole-induced expression of 12 genes involved in the late biosynthesis of ergosterol including ERG11 and ERG3. Importantly, chromatin immunoprecipitation analysis showed that histone H3K4 trimethylation was detected on chromatin of actively transcribed ERG genes. Furthermore, H3K4 trimethylation increased upon azole-induced gene expression which was also found to be dependent on the catalytic activity of Set1. Altogether, our findings show that Set1-mediated histone H3K4 methylation governs the intrinsic drug resistant status in C. glabrata via epigenetic control of azole-induced ERG gene expression.IMPORTANCEC. glabrata is the second most commonly isolated species from Candida infections, coming in second to C. albicans. Treatment of C. glabrata infections are difficult due to their natural resistance to antifungal azole drugs and their ability to adapt and become multidrug resistant. In this study, we investigated the contributing cellular factors for controlling drug resistance. We have determined that an epigenetic mechanism governs the expression of genes involved in the late ergosterol biosynthesis pathway, an essential pathway that antifungal drugs target. This epigenetic mechanism involves histone H3K4 methylation catalyzed by the Set1 methyltransferase complex (COMPASS). We also show that Set1-mediated histone H3K4 methylation is needed for expression of specific azole induced genes and azole drug resistance in C. glabrata. Identifying epigenetic mechanisms contributing to drug resistance and pathogenesis could provide alternative targets for treating patients with fungal infections.

The SET-domain protein CgSet4 negatively regulates antifungal drug resistance via the ergosterol biosynthesis transcriptional regulator CgUpc2a

“…Next, we generated and characterized deletion strains for CgSET1 , and CgSET3–6 genes, as we had recently shown that the loss of CgSET2 gene resulted in decreased sensitivity to the azole drug FLC ( 28 ). In consistence with the recent studies ( 28 , 38 ), phenotypic analysis showed increased sensitivity and a moderate level of resistance toward FLC in Cgset1Δ and Cgset2Δ mutants, respectively, compared with wild-type ( wt ) cells ( Fig. 1 A ).…”

“…To ascribe histone H3 lysine methyltransferase activity to SET domain–containing proteins in C. glabrata , we performed Western analysis and examined trimethylation of lysine 4 and lysine 36 residues in histone H3 in Cgset1Δ – Cgset6Δ mutants. As reported recently ( 28 , 38 ), the Cgset1Δ mutant exhibited no H3K4me3, whereas the Cgset2Δ mutant displayed very low levels of H3K36me3 ( Fig. 1 E ).…”

“…S1 , A and B , respectively. Notably, CgSet1 and CgSet2 enzymes have recently been shown to be required for monomethylation, dimethylation, and trimethlyation of lysine-4, and trimethylation of lysine-36 residues, in histone H3, respectively ( 28 , 38 ), while this work was underway.…”

“…Chromatin architecture is pivotal to gene expression regulation, with histone post-translational modifications playing a key role in maintenance of chromatin homeostasis ( 58 ). The epigenetic control of antifungal resistance mechanisms in C. glabrata is beginning to be elucidated ( 28 , 38 , 56 ). Toward this end, we report an essential role for the SET domain–containing protein CgSet4 in the transcriptional downregulation of CgPDR1 and CgUPC2A genes, which code for the master regulator of MDR and ergosterol biosynthesis genes, respectively, in C. glabrata .…”

Post-Translational Modifications of Histones Are Versatile Regulators of Fungal Development and Secondary Metabolism

“…To date, only HAT/HDAC inhibitors are histone PTM modifiers that are used as antifungal agents. Nevertheless, the potential function of the SET1-dependant H3K4me in the drug resistance of C. glabrata to azoles and of S. cerevisiae to Brefeldin A has been highlighted [ 180 , 181 ]. The wide variety of histone PTMs and their potential to be targeted in antifungal treatments represent a promising field that remains largely un-investigated.…”