Candida albicans RLM1 Diversity Regulates Cell Wall Composition and Virulence

Candida albicans RLM1, a transcription factor essential for cell wall integrity, contains a polymorphic CAA/G repeat (poly-Q) associated with recurrent vulvovaginal candidiasis. We aim to investigate how poly-Q repeat variations alter C. albicans phenotypic changes to elucidate the correlation between CAI microsatellites and virulence. Using CRISPR/Cas9, we generated RLM1-edited C. albicans strains with varying poly-Q repeats, including an 11_Q variant from a clinical isolate and the 39_Q allele from the parental SC5314 strain. The RLM1-edited strains with higher poly-Q exhibited greater cell wall stress tolerance and reduced echinocandin sensitivity. Transmission electron microscopy revealed significant alterations in cell wall thickness and cell volume in the low poly-Q strain treated with sub-MIC caspofungin, while no such changes were observed in high poly-Q strains. Corresponding changes in cell wall composition, as evidenced by the ratios of glucose to mannose, were observed in the caspofungin-treated yeast cells. The Galleria mellonella infection model demonstrated that the virulence of the high poly-Q strains was significantly enhanced, accompanied by higher adhesion and survival in activated THP-1 macrophages. Compared with the RLM1 39_Q/41_Q strain, the 11_Q/Δ strain displayed differential expression of envelope-associated genes and reduced induction of downstream cell wall synthesis genes under caspofungin stress. These findings indicate that RLM1 polymorphisms modulate C. albicans virulence through changes in cell wall composition and transcriptional regulation and support a role for RLM1 genetic variation in host immune evasion.