Screening of Candida albicans GRACE library revealed a unique pattern of biofilm formation under repression of the essential gene ILS1

Abstract

Candida albicans biofilm formation is governed by a regulatory circuit comprising nine transcription factors which control a network of target genes. However, there are still unknown genes contributing to biofilm features. Thus, the GRACE library was screened to identify genes involved in mature biofilm development. Twenty-nine conditional mutants were selected for a second screening revealing three groups of genes: twenty- two conditional mutants were defective for normal growth and unable to form biofilms; six strains, conditionally defective in genes ARC40, ARC35, ORF19.2438, SKP1, ERG6 , and ADE5,7 that are likely essential or involved in general cell processes, grew normally as free-floating cells but produced less biofilm; finally, the conditional strain for a putative essential isoleucyl- tRNA synthetase gene, ILS1 , was unable to grow as yeast-phase cells but was capable of producing a tridimensional biofilm structure in spite of reduced metabolic activity. This unique biofilm still relied on the classical biofilm genes, while it differentially induced groups of genes involved in adhesion, protein synthesis, cell wall organization, and protein folding. Although the conditional mutant repressed genes annotated for morphology and homeostasis processes affecting morphology and metabolism, the dynamic cell growth enabled the formation of a complex biofilm community independent of ILS1 .

Signal-mediated localization of Candida albicans pheromone response pathway components

“…Samples were tested for quality control using Bioanalyzer and submitted for sequencing to the Genome Quebec Innovation Centre using Illumina miSeq sequencing platform. RNA-seq data was processed and analyzed as described by Costa et al (2019) . Raw and processed data have been deposited in NCBI’s Gene Expression Ominibus and are accessible through GEO Series accession number GSE158062 ( https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?…”

Application of the Mutant Libraries for Candida albicans Functional Genomics

“…The GRACE library [ 22 ] was further used to identify genes involved in biofilm formation under a strong biofilm-induced condition (RPMI-MOPS medium). A strain expressing very low level of the essential gene ILS1 , encoding isoleucyl-tRNA synthetase, was incapable of yeast-phase growth but capable of producing the wild-type three-dimensional biofilm with decreased metabolic level [ 50 ]. Besides the classical regulatory clusters in biofilm, this study reveals a different regulation profile that depends on ILS1 .…”

Functional analysis of the Candida albicans kinome reveals Hrr25 as a regulator of antifungal susceptibility

“…In addition, a collection of protein kinases and their regulators under the control of a tetracycline-inducible promoter was used to define the impact of kinase overexpression on filamentous growth of C. albicans ( Bar-Yosef et al., 2018 ; Ramírez-Zavala et al., 2013 ). Finally, the gene replacement and conditional expression (GRACE) library has been employed to study genes important for fitness, drug susceptibility, and virulence in C. albicans ( Caplan et al., 2018 ; Chen et al., 2018 ; Costa et al., 2019 ; Fu et al., 2021 ; Lee et al., 2016a ; O’Meara et al., 2015 ; Roemer et al., 2003 ). Yet, given that this functional genomics resource remains limited in genome coverage, a comprehensive phenotypic analysis of the C. albicans kinome in antifungal susceptibility has yet to be performed.…”