RNA methylation has also been shown to be altered by AdO treatment and such methylation could reg ulate protein synthesis. DAL 1 4. 1B has previ ously been shown not to be a substrate for PRMT3 or PRMT5 mediated ible cascade leading to cell death in mature oligodendro cytes. The cross talk between caspase 8 and calpain has also been identified in vascular smooth muscle cells dur ing Fas associated apoptosis. DAL 1 4. 1B could also induce apoptosis through membrane protein proteases such as calpains. A related 4. 1 family tumor suppressor, NF2, has been shown to interact with calpains in neurofi bromatosis related tumors. In the case of the NF2 tumors, some patients lacking functional mutations in the NF2 gene have concomitant overactive calpain protease activity, which effectively degrades the e isting NF2 pro tein, creating a loss of tumor suppressor protein equiv alent environment.

The potential relationship between DAL 1 4. 1B and calpains and their relationship to apop tosis is important to e amine further. arginine methylation but no information is currently available as to the presence of methylated DAL 1 4. 1B mRNA species. Further investigation into the relationship between DAL 1 4. 1B, protein methylation and apoptosis is required to determine the e act mechanism by which tumor cell growth and apoptosis are regulated by these proteins. The determination that caspase 8 activation occurs in the absence of effector caspase activation suggests a poten tially novel pathway combining aspects of both tradi tional caspase dependent cell death and the emerging effector caspase independent pathways of apoptosis.

Fur thermore, the interaction between a tumor suppressor and a post translational GSK-3 methylation enzyme is likely to be an important modulator of this pathway and so be of significant biological impor tance in controlling tumorigenesis in breast cancer cells. Conclusion In this report, caspase 8 specific activation by the tumor suppressor DAL 1 4. 1B is identified in the absence of acti vation of effector caspases 3, 6, or 7, suggesting a poten tially novel apoptotic pathway combining aspects of both traditional caspase dependent cell death and the emerg ing effector caspase independent pathways of apoptosis. Second it is shown that there is cooperation between DAL 1 4. 1B and post translational protein methylation in the induction of apoptosis in MCF 7 cells.

This suggests that the previously published interaction between the tumor essential amino acids and insulin. The MCF 7 Cl27 cell line is a DAL 4. 1B inducible cell line generated from the parental MCF 7 cell line using the Ecdysone Muristerone inducible E pression Kit. DAL 1 4. 1B e pression is induced by the addition of 2 M Murister one to the culture medium for 48 hours. Hypomethyla tion of cells was carried out by addition of 30 M AdO to the culture medium for 48 hours.