An ATG5 knockout promotes paclitaxel resistance in v-Ha-ras-transformed NIH 3T3 cells

Autophagy plays a contradictory role in cell survival and death. Here, we investigated changes in paclitaxel sensitivity of cells with an ATG5 gene-knockout (KO), incapable of synthesizing an E3 ubiquitin ligase necessary for autophagy. The ATG5 KO in v-Ha-ras-transformed NIH 3T3 cells (Ras-NIH 3T3) was established using the CRISPR/Cas9 system. An LC3 immunoblot and a qRT-PCR assay were used to confirm the KO of functional ATG5. We found that the ATG5 KO led to paclitaxel resistance in Ras-NIH 3T3 cells through an ATP-binding cassette (ABC) transporter-independent mechanism. Flow cytometric analyses revealed that paclitaxel induced a remarkable significant G2/M arrest in parental cells, whereas it was relatively less effective in ATG5 KO cells. Additionally, the proportion of early apoptotic cells significantly decreased in ATG5 KO cells treated with paclitaxel than in parental cells. Interestingly, overexpression of ATG5 N-terminal cleavage product in ATG5 KO cells restored their sensitivity to paclitaxel. Taken together, our results suggest that ATG5 KO cells are resistant to paclitaxel due to the inability to produce tATG5. •An ATG5 knockout led to paclitaxel resistance in Ras-NIH 3T3 cells.•Expression of ABC transporter genes is independent of paclitaxel resistance.•Truncated ATG5 overexpression partially restores paclitaxel sensitivity.•ATG5 contributes to the cytotoxic effect of paclitaxel through tATG5 production.