Function And Mechanism Of ZNF268A Regulating DNA Damage Response

There are large variety of KRAB-zinc finger proteins,which play important roles in cellular life.KRAB-zinc finger proteins ZNF268 is one of them.So far,ZNF268 has been found for nearly two decades,but we still know little about it,which can partly due to ZNF268 not expressed in rodent.In previous studys,our laboratory has investigated that ZNF268 can encode and produce two different proteins named ZNF268a and ZNF268b2.ZNF268b2 can promote TNF?-induced phosphorylation of IKK?/? and activation of NF-?B pathway and result in cervical cancer development.Interestingly,althought ZNF268a can also be regarded as a positive regulator of NF-?B,it plays its role by binding to IKK? and facilitating the assembly of IKK complex.In addition,knockdown of ZNF268 total proteins(ZNF268a and ZNF268b2)can inhibit proliferation and promote apoptosis.Therefore,ZNF268a is a functional protein under cellular process and virus infection.The innate immune system is the first line of host defense against pathogens infection.Upon infection,pattern recognition receptors recognize structurally conserved components,such as DNA,and result in production of type I interferons and activation of NF-?B pathway to accomplish inhibition of viral replication and clearance of damaged cells.In addition,more and more reports indicate that self-DNA from double-strand breaks also induces innate immune response and NF-?B signaling in lower levels mediated by c GAS-STING pathway.Moreover,NF-?B can induce variety of cytokines and chemotactic factors to play significant functions in inflammatory response,immune response,proliferation,cell survival.In spite of increasing reports about how virus and self-DNA induce NF-?B pathway,people still have knowledge of blind spots about moleculars.In our study,we target to figure out the functiona of ZNF268a regulating cellular response induced by DNA damage.To investigate more function of ZNF268a,we utilize chemotherapy drugs to induce DNA damage in ZNF268a knockout cells.After DNA damage,the location of ZNF268a has no obvious transference in cells by confocal microscopy.And,we detect p-ATM and?-H2 AX through western blot and there is no change under ZNF268a deficiency.So,it's clear that ZNF268a has no influence on DNA damage repair.After DNA damage,cells suffer cell cycle arrest,autophagy and apoptosis.During these responses,ZNF268a deficiency cannot control cell cycle arrest and autophagy.But in MTT and flow cytometry,the loss of ZNF268a results in proliferation arrest and increasing apoptosis induced by CPT,ETO,TNF? and CHX.Thus,ZNF268a plays crucial role in process of cellular life.Furthermore,in order to understand how ZNF268a regulates inflammation and cell death induced by intrinsic DNA fragment,we first establish ZNF268a knockout U2 OS cells by two sg RNA.And then,we detect m RNA level of pro-inflammatory cytokines by RT-PCR,such as IL-6,IL-8,TNF?,and the data indicates the stimulation of chemotherapy drugs increases expression of pro-inflammatory cytokines,but the loss of ZNF268a impairs it.Finally,Dual-luciferase reporter assay verifies our expectation that ZNF268a deficiency suppressed NF-?B activity gently induced by ETO.In sum,our results suggest ZNF268a may promote NF-?B activity stimulated by chemotherapy drugs.In conclusion,ZNF268a deficiency suppresses cell proliferation and promotes apoptosis induced by chemotherapy drugs.Furthermore,loss of ZNF268a can inhibit NF-?B activity in some degree.These results can not only improve our knowledge about ZNF268 protein,but also contribute to understanding the relationship between intrinsic and extrinsic inflammation responses which may have crosstalk with each other.