COS1, a two-component histidine kinase that is involved in hyphal development in the opportunistic pathogen Candida albicans

Abstract

Two-component histidine kinases recently have been found in eukaryotic organisms including fungi, slime molds, and plants. We describe the identification of a gene, COS1, from the opportunistic pathogen Candida albicans by using a PCR-based screening strategy. The sequence of COS1 indicates that it encodes a homolog of the histidine kinase Nik-1 from the filamentous fungus Neurospora crassa. COS1 is also identical to a gene called CaNIK1 identified in C. albicans by low stringency hybridization using CaSLN1 as a probe [Nagahashi, S., Mio, T., Yamada-Okabe, T., Arisawa, M., Bussey, H. & Yamada-Okabe, H. (1998) Microbiol. 44, 425-432]. We assess the function of COS1/CaNIK1 by constructing a diploid deletion mutant. Mutants lacking both copies of COS1 appear normal when grown as yeast cells; however, they exhibit defective hyphal formation when placed on solid agar media, either in response to nutrient deprivation or serum. In constrast to the Deltanik-1 mutant, the Deltacos1/Deltacos1 mutant does not demonstrate deleterious effects when grown in media of high osmolarity; however both Deltanik-1 and Deltacos1/Deltacos1 mutants show defective hyphal formation. Thus, as predicted for Nik-1, Cos1p may be involved in some aspect of hyphal morphogenesis and may play a role in virulence properties of the organism.

Figure 1

Predicted amino acid sequence of Cos1p aligned with Nik-1(Os-1) of N. crassa and the homolog, AnNik-1, encoded by a PCR product identified from Aspergillus nidulans (L.A.A., unpublished results). Boxed residues are identical.

Figure 2

Disruption of the COS1 gene from C. albicans. (A) Schematic showing the replacement strategy. Restriction sites are H (HindIII), Bg (BglII), X (XhoI), N (NcoI), and E (EcoRI). (B) Southern blot of genomic DNA to detect replacement of COS1 with hisG-URA3-hisG. After digestion of genomic DNA with BglII, the replacement event was detected by probing with a PCR product generated from CA13 and CA15 primers. Lanes: 1, CA I4 (wild-type parent); 2, LAC13 (Δcos1∷hisG-URA3-hisG/COS1); 3, LAC15 (Δcos1∷hisG/COS1); 4, LAC18 (Δcos1∷hisG/Δcos1∷hisG-URA3hisG); 5, CA I4 (wild type); 6, LAC13 (Δcos1∷hisG-URA3-hisG/ COS1); 7, LAC14 (Δcos1∷hisG/COS1); 8, LAC17 (Δcos1∷hisG/ Δcos1∷hisG-URA3-hisG); 9, LAC16 (Δcos1∷hisG/Δcos1∷hisG-URA3hisG).

Figure 3

Morphology of C. albicans Δcos1/Δcos1 colonies grown on solid agar Spider media. Cells were grown at 37°C on solid Spider plates for 5 days (a-d) or 4 days (e-g). (a and e) SC5314 (wild- type COS1/COS1); (b and f) LAC13 (Δcos1/COS1); (c) LAC16 (Δcos1/Δcos1); (d) LAC26 (Δcos1/Δcos1 strain in which COS1∷URA3 has been recombined back at the cos1 locus, Δcos1/COS1∷URA3); (g) LAC18 (Δcos1/Δcos1); (h) LAC23 (Δcos1/Δcos1 with COS1 expressed from a 2-μ plasmid).

Figure 4

Morphology of C. albicans Δcos1/Δcos1 colonies grown at 37°C for 3 days on agar plates containing 10% (vol/vol) fetal bovine serum. (a) SC5314 (wild-type COS1/COS1). (b) LAC13 (Δcos1/COS1). (c) LAC16 (Δcos1/Δcos1). (d) LAC 22 (Δcos1/Δcos1 with COS1 on 2-μ plasmid). (e) LAC 18 (Δcos1/Δcos1). (f) LAC 23 (Δcos1/Δcos1 with COS1 on 2-μ plasmid).