Philips' group happened to stimulate GFP-labeled T cells for an unrelated experiment. “Before our eyes, K-Ras shot off the membrane and into the cell interior,” says Philips. K-Ras is attached peripherally to the PM by a farnesyl lipid group and an adjacent polybasic sequence. The team hypothesized that phosphorylation by PKC within the polybasic region acted like a switch, neutralizing the positive charge and releasing K-Ras from the PM. Indeed, blocking this serine-181 phosphorylation by PKC prevented K-Ras internalization.
A phosphate-mimicking Glu-181 mutant gave another shocking result. Cells expressing this K-Ras construct died off rapidly by apoptosis, as internalized K-Ras inhabited ER, Golgi, and—more importantly—mitochondrial membranes.
The outer mitochondrial membrane hosts apoptosis regulators such as the Bcl-2 family members. Philips' group found that the PKC-phosphorylated K-Ras interacted with Bcl-Xl at mitochondrial membranes to promote apoptosis. Human T cells lacking K-Ras were resistant to activation-induced apoptosis.
Although Bcl-Xl is normally antiapoptotic, the group proposes that K-Ras either sequesters Bcl-Xl or converts it to a proapoptotic molecule.
Mutated Ras proteins are known to drive uncontrolled cell proliferation in 30 percent of human cancers. PKC agonists may be a possible strategy for treating such cancers, as the group found that oncogenic K-Ras also moved to the mitochondria when phosphorylated by PKC. Furthermore, K-Ras–driven tumors treated with the PKC agonist bryostatin-1 underwent apoptosis. PKC agonists such as bryostatin “cause K-Ras to fall off the plasma membrane,” says Philips, “and it goes from being a molecule that drives cell growth to one that kills the cell instead.”