A Dictyostelium cellobiohydrolase orthologue that affects developmental timing

Area of Science:

• Cellular Biology
• Biochemistry
• Developmental Biology

Background:

• Dictyostelium discoideum is a model organism for studying multicellularity.
• Cellulose is present in the extracellular matrix and fruiting body of D. discoideum.
• Cellulase genes, including cbhA, are found in D. discoideum.

Purpose of the Study:

• To investigate the function of the cbhA gene in Dictyostelium discoideum.
• To determine if CbhA is a functional cellobiohydrolase.
• To assess the role of CbhA in the organism's development.

Main Methods:

• Gene expression analysis of cbhA during development.
• Creation and analysis of a cbhA null mutant.
• Expression and characterization of a c-Myc epitope-tagged CbhA protein.
• Enzyme activity assays using 4-methylumbelliferyl β-D-cellobioside (4-MUC).

Main Results:

• cbhA expression is developmentally regulated, with peak expression in spores.
• A cbhA null mutant exhibits a 2-4 hour delay in development.
• Recombinant CbhA protein is secreted, binds crystalline cellulose, and is glycosylated.
• Conditioned medium from CbhA-expressing strains shows cellobiohydrolase activity.

Conclusions:

• Dictyostelium CbhA is a functional orthologue of fungal cellobiohydrolase I (CBHI).
• CbhA plays a role in the normal rate of Dictyostelium discoideum development.
• The enzyme is secreted and active in the extracellular environment.