Differential expression patterns of the short and long isoform of cFLIP on FasL‑mediated apoptosis

Um,Hee Jung; Chauhan,Anil Kumar; Min,Kyoung‑Jin; Kwon,Taeg Kyu

doi:10.3892/or.2018.6317

Differential expression patterns of the short and long isoform of cFLIP on FasL‑mediated apoptosis

Authors:
- Hee Jung Um
- Anil Kumar Chauhan
- Kyoung‑Jin Min
- Taeg Kyu Kwon
View Affiliations

Affiliations: Department of Immunology, School of Medicine, Keimyung University, Dalseo‑gu, Daegu 704‑701, Republic of Korea
Published online on: March 16, 2018 https://doi.org/10.3892/or.2018.6317
Pages: 2443-2449

Metrics: Total Views: 0 (Spandidos Publications: | PMC Statistics: )

Metrics: Total PDF Downloads: 0 (Spandidos Publications: | PMC Statistics: )

Cited By (CrossRef): 0 citations Loading Articles...

Abstract

cFLIP is a key regulator of the anti‑apoptotic mechanism and its association with FAS‑mediated apoptosis has been widely studied and well documented. However, the equipoise between its two isoforms i.e. the long isoform cFLIP(L) and the short isoform cFLIP(S) during FAS‑mediated apoptosis remains to be revealed. Therefore, the present study aimed to investigate the regulatory effect of these isoforms on FasL‑mediated apoptosis in renal carcinoma. Our results revealed that FasL treatment to Caki cells induced the expression of cFLIP(S) and downregulated the expression of cFLIP(L) in a concentration‑ and time‑dependent manner. Furthermore, our results indicated that cell death receptor‑mediated apoptosis inducers such as TNF‑α and TRAIL, induced apoptosis in Caki cells along with downregulation of cFLIP(L), however, they had no effect on the expression of cFLIP(S). In addition, FasL‑mediated cFLIP(L) downregulation was prevented by pan‑caspase inhibitor (z‑VAD‑fmk), however pan‑caspase inhibitor did not have an effect on FasL‑induced cFLIP(S) upregulation. Furthermore, FasL induced upregulation of the expression of cFLIP(S) at the post‑translational level. Furthermore, pretreatment of Caki cells with ROS scavengers (N‑acetylcysteine and glutathione) prevented the downregulation of cFLIP(L), the upregulation of cFLIP(S) and apoptosis induced by FasL. Collectively, these data indicated that a novel pathway of cFLIP(L)/(S) differential expression pattern was associated with FasL‑induced apoptosis and modulated by ROS generation.

View Figures

View References

1	Kikuchi H, Kuribayashi F and Imajoh-Ohmi S: Down-regulation of Fas-mediated apoptosis by plasma transglutaminase factor XIII that catalyzes fetal-specific cross-link of the Fas molecule. Biochem Biophys Res Commun. 443:13–17. 2014. View Article : Google Scholar : PubMed/NCBI
2	Wiegers GJ, Kaufmann M, Tischner D and Villunger A: Shaping the T-cell repertoire: A matter of life and death. Immunol Cell Biol. 89:33–39. 2011. View Article : Google Scholar : PubMed/NCBI
3	Kikuchi H and Nakayama T: GCN5 and BCR signalling collaborate to induce pre-mature B cell apoptosis through depletion of ICAD and IAP2 and activation of caspase activities. Gene. 419:48–55. 2008. View Article : Google Scholar : PubMed/NCBI
4	Geng L, Zhu B, Dai BH, Sui CJ, Xu F, Kan T, Shen WF and Yang JM: A let-7/Fas double-negative feedback loop regulates human colon carcinoma cells sensitivity to Fas-related apoptosis. Biochem Biophys Res Commun. 408:494–499. 2011. View Article : Google Scholar : PubMed/NCBI
5	Barnhart BC, Legembre P, Pietras E, Bubici C, Franzoso G and Peter ME: CD95 ligand induces motility and invasiveness of apoptosis-resistant tumor cells. EMBO J. 23:3175–3185. 2004. View Article : Google Scholar : PubMed/NCBI
6	Wu XX, Mizutani Y, Kakehi Y, Yoshida O and Ogawa O: Enhancement of Fas-mediated apoptosis in renal cell carcinoma cells by adriamycin. Cancer Res. 60:2912–2918. 2000.PubMed/NCBI
7	Arase H, Arase N and Saito T: Fas-mediated cytotoxicity by freshly isolated natural killer cells. J Exp Med. 181:1235–1238. 1995. View Article : Google Scholar : PubMed/NCBI
8	Itoh N, Yonehara S, Ishii A, Yonehara M, Mizushima S, Sameshima M, Hase A, Seto Y and Nagata S: The polypeptide encoded by the cDNA for human cell surface antigen Fas can mediate apoptosis. Cell. 66:233–243. 1991. View Article : Google Scholar : PubMed/NCBI
9	Kischkel FC, Hellbardt S, Behrmann I, Germer M, Pawlita M, Krammer PH and Peter ME: Cytotoxicity-dependent APO-1 (Fas/CD95)-associated proteins form a death-inducing signaling complex (DISC) with the receptor. EMBO J. 14:5579–5588. 1995.PubMed/NCBI
10	Wang X, Wang Y, Lee SJ, Kim HP, Choi AM and Ryter SW: Carbon monoxide inhibits Fas activating antibody-induced apoptosis in endothelial cells. Med Gas Res. 1:82011. View Article : Google Scholar : PubMed/NCBI
11	Gloire G, Charlier E and Piette J: Regulation of CD95/APO-1/Fas-induced apoptosis by protein phosphatases. Biochem Pharmacol. 76:1451–1458. 2008. View Article : Google Scholar : PubMed/NCBI
12	Safa AR, Day TW and Wu CH: Cellular FLICE-like inhibitory protein (C-FLIP): A novel target for cancer therapy. Curr Cancer Drug Targets. 8:37–46. 2008. View Article : Google Scholar : PubMed/NCBI
13	Malhi H and Gores GJ: TRAIL resistance results in cancer progression: A TRAIL to perdition? Oncogene. 25:7333–7335. 2006. View Article : Google Scholar : PubMed/NCBI
14	Shin EC, Seong YR, Kim CH, Kim H, Ahn YS, Kim K, Kim SJ, Hong SS and Park JH: Human hepatocellular carcinoma cells resist to TRAIL-induced apoptosis, and the resistance is abolished by cisplatin. Exp Mol Med. 34:114–122. 2002. View Article : Google Scholar : PubMed/NCBI
15	Golks A, Brenner D, Fritsch C, Krammer PH and Lavrik IN: c-FLIPR, a new regulator of death receptor-induced apoptosis. J Biol Chem. 280:14507–14513. 2005. View Article : Google Scholar : PubMed/NCBI
16	Sharp DA, Lawrence DA and Ashkenazi A: Selective knockdown of the long variant of cellular FLICE inhibitory protein augments death receptor-mediated caspase-8 activation and apoptosis. J Biol Chem. 280:19401–19409. 2005. View Article : Google Scholar : PubMed/NCBI
17	Chang DW, Xing Z, Pan Y, Algeciras-Schimnich A, Barnhart BC, Yaish-Ohad S, Peter ME and Yang X: c-FLIP(L) is a dual function regulator for caspase-8 activation and CD95-mediated apoptosis. EMBO J. 21:3704–3714. 2002. View Article : Google Scholar : PubMed/NCBI
18	Krueger A, Schmitz I, Baumann S, Krammer PH and Kirchhoff S: Cellular FLICE-inhibitory protein splice variants inhibit different steps of caspase-8 activation at the CD95 death-inducing signaling complex. J Biol Chem. 276:20633–20640. 2001. View Article : Google Scholar : PubMed/NCBI
19	Ram DR, Ilyukha V, Volkova T, Buzdin A, Tai A, Smirnova I and Poltorak A: Balance between short and long isoforms of cFLIP regulates Fas-mediated apoptosis in vivo. Proc Natl Acad Sci USA. 113:1606–1611. 2016. View Article : Google Scholar : PubMed/NCBI
20	Park YS, Kwon YJ and Chun YJ: CYP1B1 activates Wnt/β-catenin signaling through suppression of Herc5-mediated ISGylation for protein degradation on β-catenin in HeLa cells. Toxicol Res. 33:211–218. 2017. View Article : Google Scholar : PubMed/NCBI
21	Jo Y and Shin DY: Repression of the F-box protein Skp2 is essential for actin damage-induced tetraploid G1 arrest. BMB Rep. 50:379–383. 2017. View Article : Google Scholar : PubMed/NCBI
22	Wang L, Azad N, Kongkaneramit L, Chen F, Lu Y, Jiang BH and Rojanasakul Y: The Fas death signaling pathway connecting reactive oxygen species generation and FLICE inhibitory protein down-regulation. J Immunol. 180:3072–3080. 2008. View Article : Google Scholar : PubMed/NCBI
23	Krueger A, Baumann S, Krammer PH and Kirchhoff S: FLICE-inhibitory proteins: Regulators of death receptor-mediated apoptosis. Mol Cell Biol. 21:8247–8254. 2001. View Article : Google Scholar : PubMed/NCBI
24	Thome M and Tschopp J: Regulation of lymphocyte proliferation and death by FLIP. Nat Rev Immunol. 1:50–58. 2001. View Article : Google Scholar : PubMed/NCBI
25	Medan D, Wang L, Toledo D, Lu B, Stehlik C, Jiang BH, Shi X and Rojanasakul Y: Regulation of Fas (CD95)-induced apoptotic and necrotic cell death by reactive oxygen species in macrophages. J Cell Physiol. 203:78–84. 2005. View Article : Google Scholar : PubMed/NCBI
26	Sato T, Machida T, Takahashi S, Iyama S, Sato Y, Kuribayashi K, Takada K, Oku T, Kawano Y, Okamoto T, et al: Fas-mediated apoptosome formation is dependent on reactive oxygen species derived from mitochondrial permeability transition in Jurkat cells. J Immunol. 173:285–296. 2004. View Article : Google Scholar : PubMed/NCBI
27	Vercammen D, Brouckaert G, Denecker G, Van de Craen M, Declercq W, Fiers W and Vandenabeele P: Dual signaling of the Fas receptor: Initiation of both apoptotic and necrotic cell death pathways. J Exp Med. 188:919–930. 1998. View Article : Google Scholar : PubMed/NCBI