Abstract
Arsenic trioxide (As2O3) has recently become one of the most effective drugs for treatment of patient with acute promyelocytic leukemia (APL), and its molecular mechanism has also been largely investigated. However, it has been reported that As2O3 resistant patients are frequently found in relapsed APL after consolidation therapy, which is due to the point mutations in B-box type 2 motifs of promyelocytic leukemia (PML) gene. In the present study, we for the first time establish whether organic arsenic species phenylarsine oxide (PAO) could induce the mutant PML-IV (A216V) protein solubility changes and degradation. Here, three different PML protein variants (i.e., PML-IV, PML-V and mutant PML-A216V) were overexpressed in HEK293T cells and then exposed to PAO in time- and dose-dependent manners. Interestingly, PAO is found to have potential effect on induction of mutant PML-IV (A216V) protein solubility changes and degradation, but no appreciable effects were found following exposure to high concentrations of iAsIII, dimethylarsinous acid (DMAIII) and adriamycin (doxorubicin), even though they cause cell death. Our current data strongly indicate that PAO has good effects on the mutant PML protein solubility changes, and it may be helpful for improving the therapeutic strategies for arsenic-resistant APL treatments in the near future.
Highlights
Arsenic trioxide (As2O3) has been widely used in traditional medicine for treatment of various diseases for more than two thousand years [1]
acute promyelocytic leukemia (APL) is a subtype of acute myeloid leukemia (AML) with distinctive biological and clinical features that has recently been found to be highly curable by all-trans retinoic acid (ATRA) and As2O3 treatment [4,5]
APL is characterized by the reciprocal chromosomal translocation t(15;17)(q22;q21), resulting in the fusion of promyelocytic leukemia (PML) gene to the retinoic acid receptor α (RARα) gene, and, the expression of PML-RARα fusion protein to block cell differentiation
Summary
Arsenic trioxide (As2O3) has been widely used in traditional medicine for treatment of various diseases for more than two thousand years [1]. LLiikkeewwiissee,, ffoorrmmaattiioonn ooff PPMMLL--nnuucclleeaarrbbooddieiess(N(NBBss))wwaassalaslosodedteetremrminiendedbybycocnofnofcoaclaml imcriocrsocoscpoypayftaefrtetrratnrsafnescfteioctnioonf PoMf PLM-ILV,PIVM,LP-MVLa-nVdaAn2d1A6V21m6Vutmanuttiannt29in3T29c3eTllsce(Fllisg(uFriegu2r).eI2n)t.eIrnetsetrinesgtliyn,giltyw, iatswfoaus nfodutnhdatthPaMt LP-MNLB-sNcBasncbaen cbleeacrlleyafrolyrmfoerdmbeyd mbyumtaunttsanPtMs PLM-IVL-(IAV2(1A62V1)6aVn)danisdsiismsiilmariltaorntoornmoarml PaMl PLM-ILV-IoVr -oVr.-AVf.tAerfteexrpeoxspuorseutroe itAosIiIAI, sthIIIe, rtehwereerewneoresignnoifisciagnntifdiicfafenrtendcieffsebreentwceesenbethtwe ePeMnLt-hNeBsPMmoLr-pNhBoslomgyorcphhanogloegsyascchoamngpeasreads compared to corresponding without arsenic treatment group, indicating the PML protein (e.g., mutant) resistance to iAsIII is probably dependence on its protein solubility changes, but not on PML-. Ww(vassv(mfswwArAuuusaaoaeeaagpgr2r2smssniia1gg1aehaaana6t6NtNreennotsltlnVeVssesseuttweerttaossomm))iipsxx-tn-nn,tsstaototepprg,g,rhhanrffaitatwwsnoontnttPnPtthhwawsooese(sMMFthaaSffnanedidddteto)LLgnccetteeePwPutttootttttAeopeApeee(erndbbdSerrrrrrOOpeoemmo)mmi2n3etttuu29heeiiohi.linnn93lnFnnaiiaennp3TeeeieavIavtgnTdedbbetect((uflhhlliielrfsete.t.resceealeaehhielittemt.c.csmeeos,,solft3t,olori,inne.seiiwiolallnln,afPfooIuuancffendrdArrretebbswrimmuuceceiiofOellt(atcfecnfiiasaPefsectterfelslxyyiee)tootcst,pc(aahhftficcPPtonnnceehheoPP)AasoxddaaPnAAPebpipnOnnnMdMoaaaOonOggnbbtbbstoiLeheLoosenonloorssedrmtmoonmcnPdmhooraroAouafmfuPmPptdfaslOtotPMPMaloaeaiaaPnbMMn-sPlnliLMPLelndtMtaePPLL-Au(pn(LptMMAaoALiOc((rdrmnfAAdio2oL2Lnddi1teett122))ngeieei6,-g,611ntmidgiViarVn66PnPPmuraneVV)MM)atMddcd-deipd)p)dmiae-LeLnLradrppcegtgo-geo-toiperrIrIroitp-tVonooVpaeaePenrttcidndn,ei,aoMeennenaadanPPitinannsdetdtLdedMMsitiienoeolrenaapigalnLgLnntkffictrrktttsr-,,-eoimoeeaVaVooemtntrrdtidhhlneaAcuiaeeaaaaee-iAnentdbxxnsntnrrnniisIpepeieondodItslIeIbbpIooinreno.Irt.ayysmersmyslH[ts[Huuun2i2ttuu(orcbd(roh,h,oF3Fee3httniwerrwia]ali]eaen-gi.ttg.nndetooeeneTutuyTttvevgmhrddddrhepPcePeeeeiiiirehraesMffMffr,3r,3ffffnnaeieeeeeAfAiAnLfdLnfrrrArrooog,eeeee-e-,sBrBIIsrrnnnnnmeIeVVeI)Is)sItttttI,,I,,, ccoonncceennttrraattiioonnss ooff PPAAOO,, aanndd iinntteerreessttiinnggllyy mmoorree tthhaann 22 μμMM ooff PPAAOO wweerree ffoouunndd ttoo bbee ssuufffificciieenntt ffoorr iinndduucciinngg mmuuttaannttPPMMLL(A(A21261V6V) p) rporteoitneisnolsuobluilbitiylictyhacnhgaensgaesswaesllwaeslPl MasLPpMroLtepinromteoidnifimcoadtioifniciantipoenllient pellet fractions after 6 h exposure (Figure 3C), indicating that PAO have strong dual effects on both normal or abnormal PML protein solubility changes along with PML proteins degradation. FFlag-PML-IV overexpressed 293T cells were exposed to: (A) low (i.e., 5, 10 and 15 μμMM); aannd ((BB)) hhiigghh cconcentrations (i.e., 20, 50 and 100 μμM) of ADR for 112 oorr 2244 hh In both cases, 4 μμMM ooff PPAAOO wweerree aallssoo uusseedd ttoo ccoommppaarree tthhee eeffffeeccttss ooff PPAAOO wwiitthh aaddrriiaammyycciinn oonn PPMMLL--IIVV pprrootteeiinn ssoolluubbility cchanges. PML-IV protein solubility changes were determined by Weesstern bblloot wwiitthh FFllaagg aannttiibbooddyy..TThheessttaarr((★)) iinnddiiccaatteess nnoonn--ssppeecciifificc pprrootteeiinnbbaannddss,,tthheewwhhiitteettrriiaannggllee((△)) iinnddiiccaatteess tthhee pprrootteeiinnddeeggrraaddaattiioonnaannddbbllaacckkttrriiaannggllee((▲) )ininddiciacatetesspprorotetieninmmooddifiifciactaitoionn
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