Knock-down of the TIM/TIPIN complex promotes apoptosis in melanoma cells

Abstract

The Timeless (TIM) and it's interacting partner TIPIN protein complex is well known for its role in replication checkpoints and normal DNA replication processes.Recent studies revealed the involvement of TIM and TIPIN in human malignancies; however, no evidence is available regarding the expression of the TIM/TIPIN protein complex or its potential role in melanoma. Therefore, we investigated the role of this complex in melanoma. Toassess the role of the TIM/TIPIN complex in melanoma, we analyzed TIM/ TIPIN expression data from the publicly accessible TCGA online database, Western blot analysis, and RT-qPCR in a panel of melanoma cell lines. Lentivirus-mediated TIM/ TIPIN knockdown in A375 melanoma cells was used to examine proliferation, colony formation, and apoptosis. A xenograft tumor formation assay was also performed. The TIM/TIPIN complex is frequently overexpressed in melanoma cells compared to normal melanocytes. We also discovered that the overexpression of TIM and TIPIN was significantly associated with poorer prognosis of melanoma patients. Furthermore, we observed that shRNA-mediated knockdown of TIM and TIPIN reduced cell viability and proliferation due to the induction of apoptosis and increased levels of γH2AX, a marker of DNA damage. In a xenograft tumor nude mouse model, shRNA-knockdown of TIM/TIPIN significantly reduced tumor growth. Our results suggest that the TIM/TIPIN complex plays an important role in tumorigenesis of melanoma, which might reveal novel approaches for the development of new melanoma therapies. Our studies also provide a beginning structural basis for understanding the assembly of the TIM/TIPIN complex. Further mechanistic investigations are needed to determine the complex's potential as a biomarker of melanoma susceptibility. Targeting TIM/TIPIN might be a potential therapeutic strategy against melanoma.

Prognostic relationship between TIPIN and colorectal cancer and experimental validation

“…The yeast direct homolog Tof1 is a core component of the replicasome and is required for the S-phase DNA damage checkpoint, fork stalling during disturbed DNA replication, and fork pausing during replication disorders, whereas Tipin and its yeast direct homolog Csm3 have the same role as Tof1 in DNA synthesis [14,15]. Some studies have found that TIPIN is highly expressed in melanoma, knockdown of TIPIN inhibits melanoma proliferation and promotes apoptosis, and TIPIN depletion reduces tumorigenicity of melanoma xenografts in immunocompromised mice [8]. Similarly in triple-negative breast cancer samples mRNA levels of TIPIN were significantly higher than specimens from other breast cancer subgroups and healthy tissues, protein levels of TIPIN were able to make breast cancer subtypes more invasive and proliferative, and TIPIN depletion reduced the tumorigenicity of triple-negative breast cancer cells [9].…”

“…TIM-interacting protein (TIPIN) is a 301-aa protein that interacts with TIM (TIMELESS) and was originally discovered in yeast [7]. A current study found that TIPIN is aberrantly expressed in melanoma and triple-negative breast cancer, and that low levels of TIPIN promote apoptosis in melanoma and triple-negative breast cancer [8,9]. However, no study has yet found whether there is a relationship between TIPIN and the development of colorectal cancer.…”

Integrated Bioinformatic Analysis Identifies TIPIN as a Prognostic Biomarker in Hepatocellular Carcinoma

“…Silencing of the TIPIN expression induced apoptosis and inhibited proliferation in breast cancer cell lines, making TIPIN a potential target for breast cancer therapy [32]. Furthermore, knockdown of the TIM-TIPIN complex has been reported to promote apoptosis in melanoma cell lines [33]. However, because of the limited conditions, we could not conduct molecular, cellular, and animal experiments; therefore, the Disease Markers 13 Disease Markers specific molecular mechanism of TIPIN in the progression of HCC remains to be further explored.…”