The main component of the cell body cytoskeleton of Trypanosoma brucei is the highly organised array of stable, subpellicular microtubules on the cytoplasmic face of the plasma membrane. Although several microtubule associated proteins (MAPs) have been shown to be associated with this array, the mechanisms by which individual microtubules interact with one another and with the membrane are still largely undetermined. In this study we have used the T. brucei cytoskeleton as a complex immunogen for the production of monoclonal antibodies to define novel cytoskeletal antigens. Screening by immunofluorescence enabled the selection of an antibody, WCB-1, which detects an antigen associated specifically with the subpellicular microtubules and not with the flagellum microtubules. The antigen (WCB210) was shown to have a relative molecular mass of 210,000 by western blotting. Immunogold studies showed the epitope to be located on the membrane-facing side of the subpellicular cage; it appears to be closely associated with the cross-bridges lying between the microtubules. Unlike many MAPs this protein was shown not to be heat stable and is predicted to be a roughly globular monomer. Even though WCB210 is a very minor component of the cytoskeleton it is heavily phosphorylated. It is possible that this protein is involved in regulation of the subpellicular microtubule crossbridges by interaction with other proteins.

REFERENCES

Bennett
V.
(
1989
).
The spectrin-actin junction of erythrocyte membrane skeletons.
Biochim. Biophys. Acta
988
,
107
121
Hemphill
A.
,
Affolter
M.
,
Seebeck
T.
(
1992
).
A novel microtubule-binding motif identified in a high molecular weight microtubule-associated protein from Trypanosoma brucei.
J. Cell Biol
117
,
95
103
Sattilaro
W.
(
1986
).
Interaction of the microtubule associated protein MAP2 with actin filaments.
Biochemistry
25
,
2003
2009
Vickerman
K.
(
1985
).
Developmental cycles and biology of pathogenic trypanosomes.
Brit. Med. Bull
41
,
105
114
This content is only available via PDF.