Characterization of the Paracoccidioides beta-1,3-glucanosyltransferase family

Abstract

The cell wall of pathogenic microbes acts as an initial barrier that is in contact with hostile environments. Several proteins are associated to the cell wall, including the glucanosyltransferases, which are attached through glycosylphosphatidylinositol anchors to the wall. Here, we characterized the Paracoccidioides beta‐1,3‐glucanosyltranferase ( Gel ) family of proteins that showed significant homology to proteins belonging to the GH72 family. Immunoassays demonstrated Gel1p associated with the cell wall and with the nucleus. For Gel2p, immune labeling was associated with the cell wall and cytoplasm. Genetic complementation studies in Saccharomyces cerevisiae demonstrated that Gel2p is able to participate in the maintenance of fungal cell wall integrity, as it was able to restore the lack of Gas1p activity in a gas1Δ mutant; Gel1p was not able to do the same. On the other hand, Gel1p restores telomeric silencing in a gas1Δ mutant, providing strong support that Gel1p can be involved in transcriptional silencing in Paracoccidioides. Use of the in vivo yeast two‐hybrid system revealed proteins that interact with Paracoccidioides Gel proteins, supporting new insights into the function of Gel family members and suggesting that they could play other roles than those established at the fungal cell wall.

Thioridazine inhibits gene expression control of the cell wall signaling pathway (CWI) in the human pathogenic fungus Paracoccidioides brasiliensis

“…As a direct consequence of the down-regulation in the CWI-controlling genes, several CWI-responsive genes, involved with synthesis of the main structural components of the fungal cell wall, also had their expression ratios reduced. Figure 4b confirms down regulation of genes involved with the synthesis of β-glucans, such as Fks and Kre6 (which encode β-1,3and β-1,6-glucan synthases, respectively), as well the β-1,3-glucanosyltransferase Gel2, a GPI-anchored protein, present in the outer layer of the fungal cell wall that is responsible for elongation and cross linking of β-glucan chains (Roemer et al 1993;de Sousa Lima et al 2012). Down-regulation was also detected for the Bgl2 gene, which encodes a β-glucosyl-hydrolase (β-glucosidase 4), another cell wall-associated enzyme known to be involved in β-glucan formation, which has been shown to be involved with maintenance of cell wall plasticity throughout the life cycle of different fungal species (Hung et al 2001).…”

Analysis of Paracoccidioides secreted proteins reveals fructose 1,6-bisphosphate aldolase as a plasminogen-binding protein

“…For the ultrastructural and immunocytochemistry studies, previously described protocols were employed [76,81,82]. The yeast cells were fixed in solution containing 2% (v/v) glutaraldehyde, 2% (w/v) paraformaldehyde, and 3% (w/v) sucrose in 0.1 M sodium cacodylate buffer pH 7.2.…”

Hemoglobin Uptake by Paracoccidioides spp. Is Receptor-Mediated

“…For the ultrastructural and immunocytochemistry studies, the protocols that were previously described by Lima and colleagues [51] were employed. Transmission electron microscopy was performed using thin sections from Pb 01 yeast that were fixed in 2% (v/v) glutaraldehyde, 2% (w/v) paraformaldehyde and 3% (w/v) sucrose in 0.1 M sodium cacodylate buffer pH 7.2.…”