Cryptococcus neoformans can form titan-like cells in vitro in response to multiple signals
Fig 5
Influence of cell density on titan-like cell formation in C. neoformans.
(A). Cells from H99 strain inoculated in Sabouraud (white bars) or TCM (black bars) at different concentrations (103, 104, 105 and 106 cells/mL) in 96-well plates. Cell body size was measured after incubation overnight at 37°C with CO2. The asterisks indicate significant differences. B) Effect of conditioned media on cell body size. Cells from H99 strain were incubated in different conditioned TCM medium denominated (see material and methods) as TCS (supernatant of titan-like cells TCM cultures), RCS (supernatant of regular size TCM cultures), TCM + TCS (a 1:1 mixture of fresh TCM with TCS), TCM + RCS (a 1:1 mixture of fresh TCM with RCS). As controls we used a diluted TCM in H2O (1:1) and fresh TCM inoculated with 104 cells/mL (TCM) and with 106 cells/mL. (C) Cells from H99 strain were grown in TCM supplemented with 30 uM and 15 uM of the quorum sensing peptide Qsp1 (NFGAPGGAYPW). As controls, TCM supplemented with 30 uM of an inactive Qsp1 (NFGAPGAAYPW), with 30 uM of a scrambled Qsp1 peptide (AWAGYFPGPNG). TCM without any supplementation was used as control. Cell body size was measured after overnight incubation at 37°C with 5% CO2. D) Titan-like cell formation of qsp1 mutant. Cells from the KN99 (wild type) and qsp1 mutant were inoculated in TCM at 104 and 106 cells/mL, and cell body size was determined after 18 h of incubation at 37°C in the presence of 5% CO2.