Transcription profile of Trichophyton rubrum conidia grown on keratin reveals the induction of an adhesin-like protein gene with a tandem repeat pattern

Abstract

BackgroundTrichophyton rubrum is a cosmopolitan filamentous fungus that can infect human keratinized tissue (skin, nails and, rarely, hair) and is the major agent of all chronic and recurrent dermatophytoses. The dermatophyte infection process is initiated through the release of arthroconidial adhesin, which binds to the host stratum corneum. The conidia then germinate, and fungal hyphae invade keratinized skin structures through the secretion of proteases. Although arthroconidia play a central role in pathogenesis, little is known about the dormancy and germination of T. rubrum conidia and the initiation of infection. The objective of this study was to evaluate the transcriptional gene expression profile of T. rubrum conidia during growth on keratin- or elastin-containing medium, mimicking superficial and deep dermatophytosis, respectively.ResultsA transcriptional profiling analysis was conducted using a custom oligonucleotide-based microarray by comparing T. rubrum conidia grown on elastin and keratin substrates. This comparison shows differences according to protein source used, but consisted of a very small set of genes, which could be attributed to the quiescent status of conidia. The modulated genes were related to the dormancy, survival and germination of conidia, including genes involved in the respiratory chain, signal transduction and lipid metabolism. However, an induction of a great number of proteases occurred when T. rubrum was grown in the presence of keratin such as the subtilisin family of proteases (Sub 1 and Sub 3) and leucine aminopeptidase (Lap 1 and Lap 2). Interestingly, keratin also promoted the up-regulation of a gene encoding an adhesin-like protein with a tandem repeat sequence. In silico analysis showed that the protein contains a domain related to adhesin that may play a role in host-pathogen interactions. The expression of this adhesin-like gene was also induced during the co-culture of T. rubrum with a human keratinocyte cell line, confirming its role in fungal-host interactions.ConclusionThese results contribute to the discovery of new targets involved in the adhesion of conidia and the maintenance of conidial dormancy, which are essential for triggering the process of infection and the chronicity of dermatophytosis.

Expression of genes containing tandem repeat patterns involved in the fungal‐host interaction and in the response to antifungals in Trichophyton rubrum

“…Domains with tandem repeats in the protein sequence, position of repeated amino acids and number of residues. A, Domain obtained from 8 . B‐D, Domains identified in the present study with the FaaPred tool…”

“…When conidia were co‐cultured with a human keratinocyte line (HaCat) and evaluated by qPCR, an increase in the expression of this gene was observed after 6 hours, followed by a decrease after 12 hours and another increase after 24 hours. Subsequent analysis showed that this gene encodes an adhesin rich in TRs 8 . TERG_08771 also shows similarity to an adhesin (MAD1) of Metarhizium anisopliae 25 and to a cell surface protein of A fumigatus (cspA—Afu3g08990).…”

“…The evaluation of genes containing TRs was carrying out during the early stage of interaction. Thus, a conidial suspension of T rubrum (10 7 conidia/mL) was added to minimal medium according to Cove, 1966 20 supplemented with 5% bovine keratin (Sigma‐Aldrich), and the cultures were shaken (130 rpm) for 24 hours at 28°C as described by Bitencourt et al 8 …”

“…Fluconazole is a triazole antifungal agent that inhibits the fungal cytochrome P450–dependent enzyme lanosterol 14 α‐demethylase, disrupts the fungal cell membrane, and impairs cell replication 7 . This compound has been widely used in recent clinical practice due to its smaller toxicity and cheaper production when compared to amphotericin B in its conventional and lipid form, respectively 8 . The main drawback of azole antifungals is an increase of drug resistance fungi strains.…”

“…Despite the clinical importance of dermatophytoses, the interaction between the fungus and host is not well elucidated 7 . Analysis of the gene expression profile of fungi grown in culture medium containing protein substrates such as keratin, which mimics superficial infections, has been used successfully to increase our understanding of fungal‐host interactions 8,9 …”