Analysis of gene expression in filamentous cells of Candida albicans grown on agar plates
Keywords:Candida albicans, filamentation, RNA extraction, qRT-PCR, RNAseq
AbstractCandida albicans (C. albicans) is a commensal organism of the human gastrointestinal and genitourinary tracts. C. albicans is also a major human pathogen, causing disease ranging from cutaneous infections to lethal systemic disease. The ability of this fungus to switch between yeast and filamentous forms of growth has long been linked to its pathogenesis. Filamentation can be induced by a variety of distinct environmental cues and can occur in either liquid or solid media. While some evidence suggests that there are differences between filamentation in solid and liquid media, gene expression analysis of filamentation in C. albicans has focused strictly on cells grown in liquid media. We have developed a method for analyzing gene expression of filamentous cells grown on solid induction media at early stages of filamentation, establishing cell plating densities, ideal collection times, and collection techniques. We have also demonstrated the utility of the approach not only in qRT-PCR assays, but high-throughput RNAseq assays as well. These assays will allow for comparison studies of C. albicans filamentation initiation in solid and liquid media.
Pfaller MA, Diekema DJ. Epidemiology of invasive candidiasis: a persistent public health problem. Clin Microbiol Rev. 2007;20(1):133-63. doi: 10.1128/CMR.00029-06. PubMed PMID: 17223626; PubMed Central PMCID: PMCPMC1797637.
Lo HJ, Kohler JR, DiDomenico B, Loebenberg D, Cacciapuoti A, Fink GR. Nonfilamentous C. albicans mutants are avirulent. Cell. 1997;90(5):939-49. PubMed PMID: 9298905.
Zheng X, Wang Y, Wang Y. Hgc1, a novel hypha-specific G1 cyclin-related protein regulates Candida albicans hyphal morphogenesis. EMBO J. 2004;23(8):1845-56. doi: 10.1038/sj.emboj.7600195. PubMed PMID: 15071502; PubMed Central PMCID: PMCPMC394249.
Saville SP, Lazzell AL, Monteagudo C, Lopez-Ribot JL. Engineered control of cell morphology in vivo reveals distinct roles for yeast and filamentous frms of Candida albicans during infection. Eukaryot Cell. 2003;2(5):1053-60. PubMed PMID: 14555488.
Hirakawa MP, Martinez DA, Sakthikumar S, Anderson MZ, Berlin A, Gujja S, et al. Genetic and phenotypic intra-species variation in Candida albicans. Genome Res. 2015;25(3):413-25. doi: 10.1101/gr.174623.114. PubMed PMID: 25504520; PubMed Central PMCID: PMCPMC4352881.
Lee KL, Buckley HR, Campbell CC. An amino acid liquid synthetic medium for the development of mycelial and yeast forms of Candida albicans. Sabouraudia. 1975;13(2):148-53. PubMed PMID: 808868.
Chen H, Fujita M, Feng Q, Clardy J, Fink GR. Tyrosol is a quorum-sensing molecule in Candida albicans. Proc Natl Acad Sci U S A. 2004;101(14):5048-52. doi: 10.1073/pnas.0401416101. PubMed PMID: 15051880; PubMed Central PMCID: PMCPMC387371.
Hornby JM, Jensen EC, Lisec AD, Tasto JJ, Jahnke B, Shoemaker R, et al. Quorum sensing in the dimorphic fungus Candida albicans is mediated by farnesol. Appl Environ Microbiol. 2001;67(7):2982-92. doi: 10.1128/AEM.67.7.2982-2992.2001. PubMed PMID: 11425711; PubMed Central PMCID: PMCPMC92970.
Staab JF, Bradway SD, Fidel PL, Sundstrom P. Adhesive and mammalian transglutaminase substrate properties of Candida albicans Hwp1. Science. 1999;283(5407):1535-8. PubMed PMID: 10066176.
Braun BR, Head WS, Wang MX, Johnson AD. Identification and characterization of TUP1-regulated genes in Candida albicans. Genetics. 2000;156(1):31-44. PubMed PMID: 10978273; PubMed Central PMCID: PMCPMC1461230.
Birse CE, Irwin MY, Fonzi WA, Sypherd PS. Cloning and characterization of ECE1, a gene expressed in association with cell elongation of the dimorphic pathogen Candida albicans. Infect Immun. 1993;61(9):3648-55. PubMed PMID: 8359888; PubMed Central PMCID: PMCPMC281060.
Schroeder A, Mueller O, Stocker S, Salowsky R, Leiber M, Gassmann M, et al. The RIN: an RNA integrity number for assigning integrity values to RNA measurements. BMC Mol Biol. 2006;7:3. Epub 2006/02/02. doi: 10.1186/1471-2199-7-3. PubMed PMID: 16448564; PubMed Central PMCID: PMCPMC1413964.
How to Cite
Authors who publish with JBM agree to the following terms:
- Authors retain copyright and grant JBM right of first publication with the work simultaneously licensed under a Creative Commons Attribution License that allows others to share the work with an acknowledgement of the work's authorship and initial publication in this journal.
- Authors are able to enter into separate, additional contractual arrangements for the non-exclusive distribution of the journal's published version of the work (e.g., post it to an institutional repository or publish it in a book), with an acknowledgement of its initial publication in this journal.
- Authors are permitted and encouraged to post their work online (e.g., in institutional repositories or on their website) prior to and during the submission process, as it can lead to productive exchanges, as well as earlier and greater citation of published work (See The Effect of Open Access).