Generating genomic platforms to study Candida albicans pathogenesis

Mélanie Legrand; Sophie Bachellier-Bassi; Keunsook K. Lee; Yogesh Chaudhari; Hélène Tournu; Laurence Arbogast; Hélène Boyer; Murielle Chauvel; Vitor Cabral; Corinne Maufrais; Audrey Nesseir; Irena Maslanka; Emmanuelle Permal; Tristan Rossignol; Louise A. Walker; Ute Zeidler; Sadri Znaidi; Floris Schoeters; Charlotte Majgier; Renaud A. Julien; Laurence Ma; Magali Tichit; Christiane Bouchier; Patrick Van Dijck; Carol A. Munro; Christophe d’Enfert

A proof of principle of the whole procedure for protein-protein detection is shown in Figure 6 . The MAP kinase kinase Hst7 was used as bait and linked to the DNA binding domain lexA, as part of the pC2HB-GC vector. Similarly, the Cek1 kinase was expressed as a prey protein, fused to the activation domain VP16, component of the pC2HP-GC vector. pC2HB-Hst7 and pC2HP-Cek1 were transformed into opaque SC2H3α and SC2H3a strains, respectively. Mating of the resulting transformants was achieved through selection for leucine and arginine prototrophy. Upon interaction of the two proteins, the reconstituted transcriptional module promoted the expression of the HIS4 marker, allowing only the mating-derived strains to grow on a histidine-free medium, whereas the original diploid strains could not (Fig. 6B) . Through a mechanism of chromosome loss, the crossed tetraploid strains return back to a diploid state. We showed that the plasmids were stably expressed in the diploid offspring strains as these strains conserved their ability to grow in absence of arginine and leucine, indicating the presence of the prey and bait vectors respectively. In addition, all these strains were able to grow in absence of histidine, indicative of the targeted protein-protein interaction. author/funder. All rights reserved. No reuse allowed without permission.