Download R032 Publication Only Basic Science: Biofilm Key proteins of

R032
Publication Only
Basic Science: Biofilm
Key proteins of Histoplasma capsulatum biofilm revealed by comparative proteomic analysis
N.S. Pitangui1, J.C.O. Sardi1, L. Scorzoni1, J.F. Da Silva1, F.P. Gullo1, G. Rodríguez-Arellanes2, M.L.
Taylor2, M.J.S. Mendes Giannini1, A.M. Fusco-Almeida1
1clinical
analysis, UNESP, Araraquara, Brazil ; 2Imunology, UNAM, City of Mexico, Mexico
Objectives: Based on the importance of biofilms and its persistence on host tissues and cell surfaces,
the present study was designed to investigate the expressed proteins by Histoplasma capsulatum
biofilms and planktonic cells, and correlate the differential proteins with interaction in host cells.
Methods: H. capsulatum biofilm assays were performed in vitro using the strain EH-315, highly
virulent. The proteomic analysis was performed by two-dimensional polyacrylamide gel electrophoresis
including sample preparation, isoelectric focusing, electrophoretic separation and image scanning and
analysis. The gels were analyzed by software ImageMaster 2D Plattinum, allowing to estimate the
isoelectric points and molecular weights of expressed proteins. Samples were sequenced using the
equipment AB SCIEX MALDI TOF/TOF 5800 System (Applied Biosystems). The spectra obtained were
processed using the Protein Pilot and the sequences were found in the Swiss databank. Results: The
proteomic analysis revealed that the samples exhibited different protein profiles, with an increased
protein expression of the strain in biofilm compared to planktonic fungal growth, being approximately
250 proteins expressed exclusively by the fungus in biofilms and others with different levels of
expression. Thus, there is a pattern of protein synthesis that correspond to a change in lifestyle of the
mobile to the sessile fungus, and the three main functional categories include proteins involved in the
metabolism of amino acids, nuclear proteins and protein translation and processing of the RNA. A
protein that highlights in proteome analysis is DASH complex subunit that is characterized as a
structural constituent of cytoskeleton and was about ten times more expressed in H. capsulatum
biofilm. Furthermore, the proteome revealed the secretion of virulence factors, which include
Metallothionein, an antioxidant protein that acts to protection the fungus against oxidative stress and,
alkaline proteinase, protein involved in the establishment of adhesion, invasion and tissue destruction.
Conclusion: The comparative proteomics reveals key proteins involved in biofilm formation,
highlighting the expression of proteins that may be involved in potential regulatory mechanism for
orchestrating the presence of a given set of yeasts inside the host cells.