Rad53- and Chk1-Dependent DNA Damage Response Pathways Cooperatively Promote Fungal Pathogenesis and Modulate Antifungal Drug Susceptibility

Abstract

Genome instability is detrimental for living things because it induces genetic disorder diseases and transfers incorrect genome information to descendants. Therefore, living organisms have evolutionarily conserved signaling networks to sense and repair DNA damage. However, how the DNA damage response pathway is regulated for maintaining the genome integrity of fungal pathogens and how this contributes to their pathogenicity remain elusive. In this study, we investigated the DNA damage response pathway in the basidiomycete pathogen Cryptococcus neoformans, which causes life-threatening meningoencephalitis in immunocompromised individuals, with an average of 223,100 infections leading to 181,100 deaths reported annually. Here, we found that perturbation of Rad53- and Chk1-dependent DNA damage response pathways attenuated the virulence of C. neoformans and increased its susceptibility to certain antifungal drugs, such as amphotericin B and flucytosine. Therefore, our work paves the way to understanding the important role of human fungal DNA damage networks in pathogenesis and antifungal drug susceptibility.

The TOR Pathway Plays Pleiotropic Roles in Growth and Stress Responses of the Fungal Pathogen Cryptococcus neoformans

“…Therefore, we questioned whether TOR1 overexpression might influence Rad53 activity. As previously reported (Jung et al 2019), Rad53 was quickly phosphorylated in response to MMS in the WT strain ( Figure 4C; left panel). However, MMS-mediated Rad53 phosphorylation did not occur in the TOR1oe strain ( Figure 4C; middle panel).…”

“…It has been recently demonstrated that Rad53 is the central protein kinase in the regulation of genes involved in DNA damage repair in C. neoformans (Jung et al 2019). Therefore, we questioned whether TOR1 overexpression might influence Rad53 activity.…”

“…Tor1 negatively regulates the DNA damage response in C. neoformans Another notable phenotype observed in the TOR1oe strain was an increased susceptibility to DNA damaging agents, such as hydroxyurea (HU), methyl methanesulfonate (MMS), and bleomycin (Bleo) ( Figure 4A). Suppressing Tor1 activity by rapamycin treatment completely abolished increased genotoxic stress susceptibility in the TOR1oe strain ( Figure 4A), indicating that the TOR pathway is indeed involved in the DNA damage response and adaptation, which is another virulence attribute of C. neoformans (Jung et al 2019).…”

“…To monitor Rad53 phosphorylation, the Rad53-4xFLAG (Jung et al 2019) and Rad53-4xFLAG TOR1oe strains were grown in YPD medium overnight at 30°. The overnight culture was inoculated into fresh YPD medium and further incubated for 5 hr at 30°until the OD 600 was 0.6.…”

Repression of BET activity sensitizes oral squamous cell carcinoma to PARP inhibition via inhibiting DNA homologous recombination and immune inhibition

“…Activation of the ATR-CHK1 pathway can cause cell cycle arrest to allow cellular repair or induce cell death when the damage is irreparable. [28]In our study, the PARP inhibitor olaparib induced levels of several important HR-related factors in Cal27 and Scc25 cells, indicating that HR is induced in response to olaparib. We further found that olaparib upregulates the expression of the ATR/CHK1 pathway, which may contribute to HR repair in OSCC cells.…”