plant surface microbiology.pdf

12 Interaction Between Soil Bacteria and Ectomycorrhiza-Forming Fungi 203
8 Biochemical Evidence for Interaction
Streptomycetes are widely distributed saprobic soil bacteria which produce a
wide range of compounds affecting other organisms. Becker et al. (1999) studied
mycorrhiza-associated Streptomyces strains with regard to their effect on
the protein pattern of ECM-forming Laccaria bicolor and Cenococcum
geophilum, and on two pathogenic fungi (Armillaria ostoyae and A. gallica).
One of the strains improved the growth of ECM fungi while inhibiting that of
the pathogens. The effects could be related to differences in fungal gene
expression (mRNA) and the protein profile obtained after in vitro translation;
new proteins were induced by the strain supporting the growth of ECM-fungi,
while the Streptomyces strain leading to adverse effects caused the disappearance
of bands.
New techniques allow for the annotation of such protein spots. Only this
way is it possible to obtain information about the function of the respective
protein. In the following, we give an example for such an approach for
Amanita muscaria.
A. muscaria is a fungus which develops ECM with a wide range of forest
trees. Grown in dual culture with bacterial isolates obtained from soil samples
in close vicinity to spruce roots, this fungus exhibited distinct changes in protein
pattern. Most effective were isolates which were members of the Actinomycetes.
After 10 weeks of dual inoculation of A. muscaria with a respective soil bacterium
in Petri dishes, the hyphae of the fungal mycelium changed their phenotype
in comparison to controls. The hyphal diameter decreased, while cell
length and the extent of hyphal branching increased. To investigate the molecular
mechanisms behind these morphological changes, the proteome of A.
muscaria was screened for differentially expressed polypeptides (two-dimensional
SDS-PAGE electrophoresis). In Fig. 1, the protein patterns for mycelium
from A. muscaria in pure culture (A) and after dual culture with the bacterium
(B) are compared. The pattern reveals about 100 well-separated protein
spots of which about 20 polypeptides were recognized as differentially
expressed. Twelve spots were excised from the gels for sequence analysis by
MALDI-TOF (matrix-assisted laser desorption/ionization time of flight) mass
spectrometry.
Reliable matches to known protein sequences with the peptide mass fingerprints
were obtained for 7 of 12 selected spots.As an example, Table 1 gives
the peptide masses obtained from protein spot no. 78. They show identity with
several predicted peptide masses of actin 1 from the saprophytic fungus
Schizophyllum commune and for actin 2 from the ectomycorrhizal fungus
Suillus bovinus (Tarkka et al. 2000).
Actins are highly conserved cytoskeletal proteins that are present in all
eucaryotic cells. They are probably involved in various processes such as cytoplasmic
streaming, cell shape determination, tip growth, cell wall deposition,