Effects Of Interferon-induced APOBEC3G On Lung Cancer Cells

The innate immune system represents the first line of defense against pathogenic challenges,foreign entities,or tissue damage,orchestrated by a variety of specific cells and molecules.Among the responses,the inflammatory reaction constitutes a prominent manifestation of the innate immune system,playing a pivotal role in safeguarding the host from infections and injuries.Nevertheless,aging is associated with a progressive loss of control over the inflammatory response,culminating in the emergence of the so-called "inflammaging" phenomenon.Inflammaging is not only closely associated with the onset of numerous age-related diseases,but it is also intimately linked to the process of cellular senescence.Cellular senescence represents a crucial factor in the context of lung cancer treatment.Recent studies have indicated that tumor cells may undergo a process known as therapyinduced cellular senescence,which impedes their proliferative and metastatic potential.Nevertheless,under specific circumstances,therapy-induced cellular senescence may also foster therapeutic resistance and tumor recurrence.The APOBEC3 family of proteins represents a crucial group of innate immune system effectors that are integral to the host defense against invading pathogens.This family comprises seven members,with APOBEC3 G being the most extensively characterized.APOBEC3 G exhibits potent antiviral activity against several viral pathogens,including HIV,via its cytidine deaminase activity that promotes G-to-A hypermutation of the viral genome.Additionally,APOBEC3G’s function extends beyond viral inhibition,as it has been found to play a role in inducing genomic DNA mutations,which could be key driving factors in the onset of lung cancer.Therefore,APOBEC3 G is considered a protein closely related to the initiation and progression of lung cancer.Its dysregulation has been implicated in various human malignancies,including lung cancer,where aberrant expression and activity of APOBEC3 G can drive the accumulation of mutations and genomic instability.Therefore,unraveling the mechanistic underpinnings of APOBEC3 Gmediated mutagenesis in lung cancer holds great therapeutic potential and may inform the development of new strategies for the prevention and treatment of this devastating disease.Thus,this study aims to gain insights into the effects of interferon-induced APOBEC3 G on lung cancer cells.First,we utilized q RT-PCR to detect changes in APOBEC3 m RNA expression in various cancer cells following treatment with either interferon-α or interferon-λ.Our experimental results indicated that the m RNA levels of some members of the APOBEC3 family were significantly upregulated by both types of interferons in colorectal cancer cells(HCT116,SW1116,SW620,SW480),cervical cancer cells(He La,Si Ha),non-small cell lung cancer cells(A549,HCC827),normal colon epithelial cell lines(NCM460)and normal lung airway epithelial cell lines(BEAS-2B),especially for APOBEC3 A,APOBEC3B,and APOBEC3 G.Notably,the upregulation effect of APOBEC3 G was most significant in HCC827 cells.Meanwhile,we have validated the ability of APOBEC3 G to induce cytosine deamination leading to uracil accumulation and consequent generation of apurinic/apyrimidinic(AP)sites in HCC827 cells.To control for the potential confounding effects of interferon-induced other proteins,we established two doxycycline-inducible cell lines overexpressing APOBEC3 G,named as A3G-HCC827 and A3G-A549,as positive controls.Furthermore,we generated two APOBEC3 G knockdown cell lines,HCC827-sh A3G#1 and HCC827-sh A3G#2,using sitespecific sh RNA targeting,as negative controls.These cell lines were validated by means of Western blot analysis.Using these cell lines,we aimed to investigate the impact of upregulated APOBEC3 G expression on lung cancer cells.The results demonstrated that upregulation of APOBEC3 G led to a significant increase in comet tail length and DNA double-strand breaks,as well as activation of the ATM-Chk2 and ATR-Chk1 pathways.Additionally,upregulation of APOBEC3 G resulted in increased expression of senescence-related proteins,including p53,p21,and p16,which caused a reduction in proliferation and ultimately cell senescence in lung cancer cell lines.Consistently,b-galactosidase staining showed that the number of senescent cells was significantly increased by APOBEC3 G.Moreover,we found that senolytic drugs can eliminate senescent cells induced by APOBEC3 G upregulation.These findings reveal the potential role of APOBEC3 G as a mutagenic source in the development of lung cancer and cellular senescence,and provide a theoretical basis for the combined application of interferon and senolytic drugs in the treatment of lung cancer.