Journal: Oncogene

Article Title: RhoA triggers a specific signaling pathway that generates transforming microvesicles in cancer cells

doi: 10.1038/onc.2011.636

Figure Lengend Snippet: MVs promote tumor formation. ( a ) Schematic of the tumor-formation assays performed to determine the role of MVs in mediating tumor growth. Mitotically-arrested MDAMB231 cells (using mitomycin C) were co-injected into nude mice with an equal number of normal (non-transformed) NIH3T3 fibroblasts. The ability of the MVs generated by the mitotically-arrested MDAMB231 cells to confer upon the recipient NIH3T3 fibroblasts the ability to form tumors was read-out. ( b ) Table showing the results of a tumor-formation assay performed as outlined in a . Half of the mice injected with mitotically-arrested MDAMB231 and NIH3T3 cells ( Mito-C-treated 231 cells + NIH3T3 cells ) formed tumors, whereas mice injected with the mitotically-arrested cancer cells ( Mito-C-treated 231 cells ) or the NIH3T3 cells (Naïve NIH3T3 cells ), alone, failed to form tumors. Naïve MDAMB231 cells ( Naïve 231 cells ) were injected into the animals to confirm the tumor-promoting ability of these cells. ( c ) Lysates generated from cultures of NIH3T3 mouse fibroblasts and of MDAMB231 human breast cancer cells, as well as from the three tumors that formed as a result of combining the mitotically-arrested MDAMB231 and NIH3T3 cells ( Mito-C-treated 231 cells + NIH3T3 cells ) were immunoblotted with an antibody that recognizes only the mouse form of Cool-1/β-Pix ( top panel ). Note that the Cool-1/β-Pix protein can be detected in each of the tumor samples, suggesting that the tumor masses are made-up of NIH3T3 mouse fibroblasts. The blot was then re-probed with actin to confirm equal loading ( bottom panel ).

Article Snippet: The tTG and actin antibodies were from Lab Vision/Thermo, the flotillin-2 and ROCK-1 and -2 antibodies were from Santa Cruz, the Cool-1/β-Pix antibody was from BD Biosciences, while the HA and Myc antibodies were from Covance.

Techniques: Injection, Transformation Assay, Generated, Tube Formation Assay