Ras Oncoproteins Transfer from Melanoma Cells to T Cells and Modulate Their Effector Functions

Abstract

Lymphocytes establish dynamic cell–cell interactions with the cells they scan. Previous studies show that upon cell contact, various membrane-associated proteins, such as Ras-family proteins, transfer from B to T and NK lymphocytes. Mutations in RAS genes that encode constitutively active, GTP-bound, oncoproteins are rather common in human cancers; for instance, melanoma. Cancer immunoediting has been postulated to contribute to the elimination of malignant melanoma. Thus, we asked whether Ras oncoproteins can transfer from melanoma to T cells, including tumor-infiltrating lymphocytes (TILs), and subsequently induce functional effects in the adopting T cells. To explore this issue, we genetically engineered an HLA-A2+ melanoma cell line, MEL526, to express GFP or GFP-tagged H-Ras mutants stably. In this study, we show by an in vitro coculture system that GFP-tagged H-Ras, but not GFP, transfers from MEL526 to T cells and localizes to the inner aspect of their plasma membrane. This cell-contact-dependent process was increased by TCR stimulation and did not require strict Ag specificity. Importantly, we found a positive correlation between the levels of the acquired constitutively active H-RasG12V and ERK1/2 phosphorylation within the adopting TILs. We also show a significant increase in IFN-γ production and cytotoxic activity in TILs that acquired H-RasG12V compared to TILs that acquired a different H-Ras mutant. In conclusion, our findings demonstrate a hitherto unknown phenomenon of intercellular transfer of Ras oncoproteins from melanoma to TILs that consequently augments their effector functions.

Ras Signaling Inhibitors Attenuate Disease in Adjuvant-Induced Arthritis via Targeting Pathogenic Antigen-Specific Th17-Type Cells

“…Interestingly, previous reports suggest that inner plasma membrane (PM)-anchored Ras proteins can transfer from antigen-presenting B cells to T cells both through the immunological synapse and through actin supported long-range PM extensions termed tunneling nanotubes. These findings further support the importance of Ras GTPases in immunity, by showing that active Ras-GTPase signals can spread between immune cells (12, 13) and even from cancer cells expressing oncogenic Ras to T cells (14). …”

H-Ras transfers from B to T cells via tunneling nanotubes

“…12, 23, 25 We have previously validated this observation regarding the transfer of GFP-H-Ras from B721.221 cells to NK cells 12 as well as from melanoma transfectants to T cells. 27 In these studies, by immunofluorescence staining with anti-Ras mAbs, we found a positive correlation between the immunostaining and the amount of GFP-H-RasG12V acquired by the T cells only in permeabilized cells – but not in intact cells. Moreover, our previous 12, 27 and present findings show that GFP-H-RasG12V is primarily distributed in the PM of the adopting T cells.…”

“…27 In these studies, by immunofluorescence staining with anti-Ras mAbs, we found a positive correlation between the immunostaining and the amount of GFP-H-RasG12V acquired by the T cells only in permeabilized cells – but not in intact cells. Moreover, our previous 12, 27 and present findings show that GFP-H-RasG12V is primarily distributed in the PM of the adopting T cells. Taken together, all these multimodality data suggest that the transferred GFP-H-RasG12V is in fact incorporated into the inner aspect of the PM of the adopting T cells and signals to induce the phosphorylation of its downstream effector ERK1/2.…”

“…In a very recent study, we have furthermore demonstrated that the transfer of H-RasG12V from melanoma cells to melanoma-specific cytotoxic T cells augments their effector functions, including cytokine secretion and cytotoxic degranulation. 27 However, with current technologies it is difficult to determine the relative functional contribution of membrane-associated H-RASG12V proteins transferring either via TNTs or via trogocytosis.…”

Intercellular transfer of Ras: Implications for immunity

“…Such connections allow the transfer of membrane-anchored proteins, plasma membrane (PM) fragments, and even microRNAs 1 - 5 . This “unorthodox” intercellular information transfer occurs through various incompletely defined mechanisms.…”