Systematic gene overexpression inCandida albicansidentifies a regulator of early adaptation to the mammalian gut

Abstract

Candida albicans is part of the human gastrointestinal (GI) microbiota. To better understand how C. albicans efficiently establishes GI colonisation, we competitively challenged growth of 572 signature‐tagged strains (~10% genome coverage), each conditionally overexpressing a single gene, in the murine gut. We identified CRZ2, a transcription factor whose overexpression and deletion respectively increased and decreased early GI colonisation. Using clues from genome‐wide expression and gene‐set enrichment analyses, we found that the optimal activity of Crz2p occurs under hypoxia at 37°C, as evidenced by both phenotypic and transcriptomic analyses following CRZ2 genetic perturbation. Consistent with early colonisation of the GI tract, we show that CRZ2 overexpression confers resistance to acidic pH and bile salts, suggesting an adaptation to the upper sections of the gut. Genome‐wide location analyses revealed that Crz2p directly modulates the expression of many mannosyltransferase‐ and cell‐wall protein‐encoding genes, suggesting a link with cell‐wall function. We show that CRZ2 overexpression alters cell‐wall phosphomannan abundance and increases sensitivity to tunicamycin, suggesting a role in protein glycosylation. Our study reflects the powerful use of gene overexpression as a complementary approach to gene deletion to identify relevant biological pathways involved in C. albicans interaction with the host environment.

Recent advances in understanding Candida albicans hyphal growth

“…Other notable approaches that are changing how we work with and view this fungal pathogen include experimental or micro-evolution approaches 13– 15 , in particular with respect to host niche environments. In addition, large-scale approaches, such as population and genetic diversity analyses via genome sequences of large numbers of isolates 16 , and the establishment of genomic platforms that facilitate the study of gene function at a genome-wide level 17– 20 pave the way for future multi-omic studies.…”

“…These studies yielded important information on gene essentiality and azole resistance in this fungal pathogen. Extensive effort was also invested in generating a genomic platform centered on an ORFeome collection representing the majority of open reading frames (ORFs) in Gateway donor vectors, together with a wide range of expression vectors 18 facilitating genome-wide overexpression analyses and protein–protein interaction studies 16, 20, 34 . Together, these new technologies have facilitated recent advances in hyphal growth signaling, host interactions, and membrane traffic.…”

Identification and Characterization of a Neutral Locus for Knock-in Purposes in C. parapsilosis

“…We first analyzed CpRP10, the ortholog of RPS1 in C. albicans, as it is one of the most preferred loci for integrating exogenous DNA in that species (Chauvel et al, 2012;Gerami-Nejad et al, 2013). We adopted an integrative plasmid designed for overexpression that takes advantage of the Gateway TM cloning method (Znaidi et al, 2018). We generated a GFP-expressing derivative utilizing the LEU2 gene for selection.…”

“…The most preferred locus for knock-in approaches in C. albicans is RPS1, also known as RP10 or orf19.3002 (encodes the ortholog of the ribosomal protein 10 from Saccharomyces cerevisiae), because constructs integrated into this region provide strong, homogenous expression that have been exploited in many studies (Swoboda et al, 1995;Chauvel et al, 2012;Legrand et al, 2018;Znaidi et al, 2018). In agreement with this, we identified the ortholog of RPS1 in C. parapsilosis (CPAR2_110290) using the BLAST search function of the Candida Genome Database and named it CpRP10 (Skrzypek et al, 2017).…”

Short-Term Probiotic Administration Increases Fecal-Anti Candida Activity in Healthy Subjects

“…Candida albicans ( C. albicans ) is a natural inhabitant of the gut and at the same time may behave as an aggressive pathogen causing life-threatening disease [1]. The human immune system has developed multiple strategies to fight fungal infections, but at the same time Candida spp.…”