# A BRCT domain-containing protein induced in early phagocytosis plays a crucial role in the pathogenesis of the mucoralean Rhizopus microsporus

## TL;DR

A protein called BRCA1 helps the fungus Rhizopus microsporus survive inside immune cells, making it a key target for treating mucormycosis.

## Contribution

Identification of BRCA1 as a critical virulence factor in Rhizopus microsporus during phagocytosis.

## Key findings

- BRCA1, HDA1, and HIST1 mutants in Rhizopus microsporus show reduced virulence in mice.
- BRCA1 knockout reduces fungal burden in lungs and brain and impairs survival against immune cells.
- Gene deletions in Mucor lusitanicus do not affect virulence, showing species-specific differences.

## Abstract

Mucormycosis, caused by mucoralean fungi, is among the most lethal fungal diseases, and a deeper understanding of its pathogenesis is urgently needed. Transcriptomic profiling of virulent strain (WT) and an RNAi-deficient strain (r3b2Δ) of M. lusitanicus during phagocytosis uncovered thousands of differentially expressed genes (DEGs), highlighting early metabolic activation as a key survival strategy inside the phagosome. Enriched pathways included amino acid transport, nucleotide metabolism, and translation, reflecting an adaptive fungal response to nutrient deprivation and host immune stress. Integrative analyses of mRNA and sRNA profiles also revealed a critical role of the RNAi pathways in modulating gene expression during infection. Building on these observations, we identified four chromatin- and transcription-related candidate virulence genes, brca1, box, hist1, and hda10, which were strongly upregulated during phagocytosis and regulated by RNAi. Functional disruption of their orthologs in the clinically relevant pathogen R. microsporus significantly reduced virulence in healthy mice, particularly in the brca1, hda1, and hist1 mutants. The brca1 knockout mutant also exhibited lower fungal burden in the brain and lungs and reduced survival rates after exposure to peritoneal immune cells. In contrast, their deletion in M. lusitanicus had no detectable impact on virulence in healthy animal models, highlighting species-dependent differences in pathogenic potential. These results demonstrate that M. lusitanicus is a valuable genetic model. However, combining studies across multiple Mucorales species is essential to uncover both conserved and species-specific mechanisms of host adaptation and virulence. These insights contribute to a broader understanding of fungal adaptation, immune evasion, and the identification of novel targets for antifungal intervention.

Mucormycosis is a severe fungal infection caused by Mucorales species, often affecting immunocompromised individuals. These pathogens can survive inside macrophages, immune cells that normally destroy invading microbes. Understanding how Mucorales escape these defenses is essential for developing new treatments. In this study, we investigated early interactions between macrophages and the model organism Mucor lusitanicus. Analysis of gene expression during phagocytosis revealed activation of metabolic and stress-response pathways that promote intracellular survival, with RNA interference contributing to their regulation. Importantly, we identified several chromatin- and transcription-related genes; brca1, hist1, hda10, and box, that are induced during host–pathogen interactions. Functional experiments showed that loss of these genes in Rhizopus microsporus, one of the main causal agents of mucormycosis, reduced its ability to cause disease in mice. In particular, loss of the brca1 gene strongly impaired virulence, resulting in reduced fungal burden in the lungs and brain. It also decreases survival following exposure to peritoneal immune cells. In contrast, deletion of these genes in M. lusitanicus did not affect virulence. These findings highlight species-specific differences in virulence mechanisms. Overall, our work provides new insights into fungal adaptation to host immunity, identifying potential antifungal targets.