Arginine deimimase expressed in vivo activates the mitochondrial apoptosis pathway through inhibiting cytosolic ferritin and inducing chromotin autophagy

Background：Based on its low toxicity, arginine starvation therapy has the potential to treat those malignant tumors that can’t be treated by surgery. Arginine deiminase (ADI) gene is indicated to be an idea cancer-suppressor gene. ADI expressed in vivo displays higher oncolytic efficiency than ADI-PEG20 (Pegylated Arginine Deiminase by PEG 20,000)[1]. However, it is still unknown whether cytosolic ADI has the same function mechanism as ADI-PEG20 or other underlying mechanisms in cells. Methods: The interaction of ADI and other protein factors was 2 screened by yeast hybrid, and verified by co-immunoprecipitation and immunofluorescent staining. The effect of ADI inhibiting ferritin light-chain domain (FTL) on mitochondria damage was evaluated by site-directed mutation and flow cytometry. The apoptosis pathway of mitochondria control was analyzed by Western Blot and real-time PCR. The effect of p53 expression on cancer cell death was assessed by siTP53 transfection. The chromatin autophagy was explored by immunofluorescent staining and Western Blot. Results: ADI expressed in vivo inhibited the activity of cytosolic ferritin through interacting with FTL. The inactive mutant of ADI still aroused the apoptosis of some cells through mitochondria damage. Arginine starvation also induced the expression increase of p53 and p53AIP1, which aggravate cellular mitochondria damage. Chromatin autophagy appeared at the later stage of arginine starvation. DNA damage came along with the whole process of arginine starvation. Histone 3 (H3) was found in autophagosomes, which implied that cancer cells try to utilize the arginine in histones to survive during arginine starvation. Conclusions: Mitochondria damage is the major mechanism for ADI expressed in vivo to induce cancer cell death. Chromatophagy accumulations not only drive cancer cell to utilize histone arginine but also speed up cancer cell death at the later time point of arginine starvation