Glycolipid-protein interaction in the mechanism of signal transduction: studies with a photoactivable ganglioside analogue.
An increasing body of evidence suggests that glycolipid domains are present on the plasma membrane surface of mammalian cells and play a key role in signal transduction. We have investigated the modulation of glycolipid-protein interaction consequent to a specific event occurring at the plasma membrane. For this purpose, a new photoactivable, radioactive derivative of GM1 ganglioside, carrying a phenyldiazirine fatty acid labelled with 125I, has been used with rat cerebellar granule cells in culture. Upon incubation of photoactivable GM1 with the cells followed by illumination, several proteins become radioactive and were detectable on the two dimensional-electrophoresis, which points to their interaction with the ganglioside. Upon addition of cytotoxic doses of glutamate, known to induce indirectly the activation of protein kinase C (PKC), one of the proteins crosslinked by photoactivable GM1 in control cells of molecular mass about 92 kDa and pI about 4, was not anymore detectable; this suggests its exclusion from the glycolipid domains. On the contrary, another protein, of about 15 kDa and pI 6.5, previously not crosslinked, was interacting with the ganglioside derivative after glutamate treatment. Comparable effects were exerted by phorbol-2-myristate-3-acetate, which directly induces the activation of PKC. These results show that PKC activation, a key step of inbound trans-membrane signalling, affects the interaction between glycolipids and proteins at the plasma membrane surface, possibly within a mixed domain. The dynamic modulation of ganglioside-protein interaction may affect the involvement of glycolipid domains in membrane-located events such as signal transmission and lipid/protein sorting.
Masserini
M
M
Department of Medical Chemistry and Biochemistry, University of Milano, Italy. masserin@imiucca.csi.unimi.it
Pitto
M
M
Ferraretto
A
A
Brunne
J
J
Palestini
P
P
D000818:Animals; D002531:Cerebellum / Q000378:metabolism*; D005677:G(M1) Ganglioside / Q000378:metabolism*; D006017:Glycolipids / Q000378:metabolism; D007457:Iodine Radioisotopes; D011493:Protein Kinase C / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction
D006017:Glycolipids; D007457:Iodine Radioisotopes; D005677:G(M1) Ganglioside; D011493:Protein Kinase C