Mus musculus Transcriptome or Gene expression

The general transcription factor IIH (TFIIH) is a multi-subunit complex involved in basal transcription as well as nucleotide excision repair. Consequently, mutations in several TFIIH subunits, including the helicase XPD, can lead to defective nucleotide excision repair (NER) and decreased basal transcription. In humans, germline mutations in XPD can lead to xeroderma pigmentosum (XP), trichothiodystrophy (TTD) and Cockayne syndrome and go along with reduced NER capacity. However, while XP patients suffer from a highly increased risk of skin cancer including melanoma, TTD patients have developmental defects, but no predisposition to cancer. To better understand the reason for the lack of tumorigenic potential, we developed cell culture models of normal and TTD-mutant melanoma cells containing the XPD-R722W mutation. Our results show that the generation of TTD-mutant melanoma cell clones was possible, thus revealing that TTD mutations are generally compatible with melanoma cell growth. XPD-R722W melanoma cells were more sensitive to UV irradiation than control cells. Furthermore, proliferation and migration of XPD-R722W cells were substantially reduced compared to control cells, demonstrating that the TTD-causing mutation affects tumorigenic features beyond UV sensitivity. To identify factors mediating this tumor-suppressive effect, we performed comparative RNA sequencing of control and XPD-R722W melanoma cells. We identified DNA damage-inducible transcript-4 like protein (DDIT4L, also called REDD2) as the topmost upregulated gene in XPD-mutant mouse melanoma cells. REDD2 has been previously implicated in the inhibition of mTOR - a pathway, which is activated in most melanomas. Interestingly, we found that high REDD2 expression is predictive for enhanced overall survival of skin cutaneous melanoma patients, and melanoma cell lines exhibit very low or even undetectable amounts of REDD2, implying that it might fulfill a tumor-suppressive role in melanoma. Functional studies revealed that REDD2 expression leads to delayed tumorigenic abilities such as proliferation and migration in melanoma cells and suppresses the phosphorylation of the mTORC1 targets and translation regulators p70-S6K and 4EBP1. In summary, our data reveal that the mTOR inhibitor REDD2 is involved in the suppression of melanoma cells expressing TTD-causing XPD mutations.