Inhibition of the RhoA GTPase Activity Increases Sensitivity of Melanoma Cells to UV Radiation Effects

Abstract

Ultraviolet radiation is the main cause of DNA damage to melanocytes and development of melanoma, one of the most lethal human cancers, which leads to metastasis due to uncontrolled cell proliferation and migration. These phenotypes are mediated by RhoA, a GTPase overexpressed or overactivated in highly aggressive metastatic tumors that plays regulatory roles in cell cycle progression and cytoskeleton remodeling. This work explores whether the effects of UV on DNA damage, motility, proliferation, and survival of human metastatic melanoma cells are mediated by the RhoA pathway. Mutant cells expressing dominant-negative (MeWo-RhoA-N19) or constitutively active RhoA (MeWo-RhoA-V14) were generated and subjected to UV radiation. A slight reduction in migration and invasion was observed in MeWo and MeWo-RhoA-V14 cells but not in MeWo-RhoA-N19 cells, which presented inefficient motility and invasiveness associated with stress fibers fragmentation. Proliferation and survival of RhoA-deficient cells were drastically reduced by UV compared to cells displaying normal or high RhoA activity, suggesting increased sensitivity to UV. Loss of RhoA activity also caused less efficient DNA repair, with elevated levels of DNA lesions such as strand breaks and cyclobutane pyrimidine dimers (CPDs). Thus, RhoA mediates genomic stability and represents a potential target for sensitizing metastatic tumors to genotoxic agents.

Impact of silver nanoparticles on the ultraviolet radiation direct and bystander effects on TK6 cell line

“…[ 37 ] Our study contrasted with the study of Espinha et al . [ 38 ] In that study, UVC radiation caused a significant decrease in cells viability and the used UVC radiation was much higher than that in our study.…”

Actin cytoskeleton dynamics in stem cells from autistic individuals

“…In order to certify that the selected inducers would actually activate RhoGTPases, we conducted an activity assay, to measure GTP-bound Cdc42, Rac and RhoA proteins. The following protocol was adapted from Espinha et al ., 2016 and Ascer et al ., 2015 33 , 34 . E. coli (strain BL21-DE3) bacteria were transformed via thermal shock with plasmids containing the sequence encoding the fusion protein RBD-GST (Rhotekin-Binding Domain-fused to Glutathione S-Transferase) or PBD-GST (PAK1-Binding Domain fused to Glutathione-S Transferase), which was kindly donated by Gary M. Bokoch from Scripps Research Institute, La Jolla, CA, USA.…”

Subversion of Ras Small GTPases in Cutaneous Melanoma Aggressiveness

“…For that purpose, RHOA was constitutively expressed or silenced in melanoma cells and subsequently the cells were exposed to UV. RHOA silencing was found to induce an increase in melanoma sensitivity to UV, increasing the number of DNA damages caused and dramatically reducing melanoma cell motility and invasion (Espinha et al, 2016). As such, RHOA can regulate DNA repair mechanisms, playing a central role in protection against UV radiation, by mediating genomic stability of melanoma cells (Espinha et al, 2016) (Table 1).…”

“…RHOA silencing was found to induce an increase in melanoma sensitivity to UV, increasing the number of DNA damages caused and dramatically reducing melanoma cell motility and invasion (Espinha et al, 2016). As such, RHOA can regulate DNA repair mechanisms, playing a central role in protection against UV radiation, by mediating genomic stability of melanoma cells (Espinha et al, 2016) (Table 1). Hence, this protein was suggested as a promising therapeutic target to sensitize melanoma cells to genotoxic damages (Espinha et al, 2016).…”

“…As such, RHOA can regulate DNA repair mechanisms, playing a central role in protection against UV radiation, by mediating genomic stability of melanoma cells (Espinha et al, 2016) (Table 1). Hence, this protein was suggested as a promising therapeutic target to sensitize melanoma cells to genotoxic damages (Espinha et al, 2016). Nevertheless, in the clinical context, immunohistochemical analyses showed that there is an increase in RHOA expression associated with smaller tumors and an increase of tumor-infiltrating lymphocytes (Kaczorowski et al, 2019) (Table 1).…”