Cell wall composition and structure of Yarrowia lipolytica transposon mutants affected in calcofluor sensitivity.
A collection of transposon-mutagenized strains of Yarrowia lipolytica was screened for wall defects by determination of their sensitivity to calcofluor white. A number of strains were hypersensitive, whereas others were resistant. Different non-allelic mutants displayed increased sensitivity to autolysis and lytic enzymes, independently of whether they were sensitive or resistant to calcofluor white. A thorough analysis of their cell walls revealed minor quantitative alterations, and no significant changes in chitin content. Electrophoretic analysis of wall-bound and excreted proteins proved to be a sensitive method that revealed defects in the cell wall structure of the mutants. Important alterations in the patterns of the wall proteins extracted by SDS or by enzymatic treatments were noticed for the mutants, as compared to the parental strain. Mutants released to the growth medium a larger number of protein species than the parental strain, suggesting impairment in wall assembly of certain polypeptides. Patterns of wall-bound and excreted proteins, as well as alterations in wall chemical composition were not diagnostic of calcofluor white sensitivity or resistance, but were specific for each mutant. Our data show that an increase in either sensitivity or resistance of Y. lipolytica to certain levels of calcofluor is equally indicative of alterations in cell wall structure, independent of chitin levels.
Departamento de Ingeniería Genética, Centro de Investigación y de Estudios Avanzados del I.P.N, Unidad Irapuato, Apartado Postal 629 Irapuato, Gto, México.
José
J
Ruiz-Herrera
Patricia
P
García-Maceira
Luis C
LC
Castillo-Barahona
Eulogio
E
Valentín
Rafael
R
Sentandreu
D001557:Benzenesulfonates / Q000494:pharmacology*; D002458:Cell Fractionation; D002473:Cell Wall / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism / Q000648:ultrastructure; D002686:Chitin / Q000378:metabolism; D002688:Chitinases / Q000378:metabolism; D003470:Culture Media; D004251:DNA Transposable Elements; D025141:Drug Resistance, Fungal / Q000235:genetics; D005656:Fungal Proteins / Q000737:chemistry / Q000302:isolation & purification; D006867:Hydrolases / Q000378:metabolism; D016254:Mutagenesis, Insertional; D009154:Mutation; D025062:Yarrowia / Q000737:chemistry* / Q000187:drug effects / Q000235:genetics*
D001557:Benzenesulfonates; D003470:Culture Media; D004251:DNA Transposable Elements; D005656:Fungal Proteins; D002686:Chitin; C037833:zymolyase; C007061:C.I. Fluorescent Brightening Agent 28; D006867:Hydrolases; D002688:Chitinases