There are three DNA polymerases, a, b, and y, in mam malian cells which can bedifferentia ted by their size, subce l lu lar location, subst ra te specificities, and susceptibilityto <a href="http://www.targetmol.com/compound/Chloramphenicol">buy
Chloramphenicol</a> specific inh ibitors. The role of each polymerase in DNA metabolismhasbeen the sub jectof many stud ies.The costs of pub lication of th is article were defrayed in partby the payment of page charges.This article must therefore be hereby marked advertise nent in accordance with U.Earl ier indirect implica te DNA polymerasepin repa ir syn thes is; however, otherstud ies indic ate th atpolymerase a is responsible for repa ir synthesis.We have been att empting to reconcile these con flicting reports and to clarifytheroles ofthedifferent DNA polymerases in repa ir synthesis.The effect of thesalt concen tration in situ on theinvolvement of DNA polymerases a and, in repa ir syn thes is has been repor ted. This study reportstheeffect of DNA polymerase inh ibitors on DNA repa ir syn thes is in subcellular preparations of hamster and human cells, using alkylating agen ts, UV irrad iation, neocarz inos tatin, and bleomycin to dam age DNA. Theeffect ofthe amount of DNA damage ontheinvolvement ofthedifferent DNA poly merases in repa ir syn thes is is also investigated.Incorporation of CHTTP into DNA was determined as described. Bleomycin and neocarz inos tatin, which re lease bases and break DNA st rands, induced DNA syn thesis when incuba ted directly wi th permeab le cells.Max imum DNA syn thes is was ob ta ined with pg dof bleomycinor pgml of neocarzinostatin.Max imum DNA syn thes is was ob ta ined bytreating G I cells with L M MNNG or mM NMU in grow th med ium for or h, respectively, at C.ForUV irrad iation, H F cells were rinsed wi th phospha tebuffered saline and exposedto UV lightatmicrowatts cm for min.Followingtrea tm entwi th thealkylating agents or UV irrad iation, cells were im med ia te ly collected, permeabilized, and assayed for DNA syn thes is in situ.Typ ical va lues for the amountof DNA synthesized in permeab le HF cells, expressed as picomoles of TTP incorpora ted X lo cellsmin were: for exponential cells. for GI cellstrea ted wi th neocarzinostatin. for G cellstrea ted wi th NMU, and. S imilar values were ob ta ined with CHO cells. Po lymerase a is inh ibited by aphidicolin, araCTP, and NEM; polymerasepis inh ibited bydd T T P and poly meraseyis inh ibited byddTT P and NEM. The effect ofthe concen tration of DNA polymerase inh ibitors on DNA repli cation and repair syn thesisin situ, aswell as on the activity of purified hamsterliver polymerase, f, was de term ined. In order to comp are theeffect of polymerase inh ibitors on in situ replicative synthesis,in situ repa ir syn thes is induced by dif fe rent DNA damaging agen ts and polymerase, f activity, re sults were expressed as per cent inh ibit ion of DNA syn thes is by each polymerase inh ibitor. For in situ repa ir syn thes is, results were correc ted fortheeffect of DNA polymerase inh ibitors on DNA syn thes is in G I CHO and HF cells in the absence of DNA damaging agents.