Response of human DNA polymerase iota to DNA lesions

Abstract

Lesion bypass is an important mechanism to overcome replication blockage by DNA damage. Translesion synthesis requires a DNA polymerase (Pol). Human Pol iota encoded by the RAD30B gene is a recently identified DNA polymerase that shares sequence similarity to Pol eta. To investigate whether human Pol iota plays a role in lesion bypass we examined the response of this polymerase to several types of DNA damage in vitro. Surprisingly, 8-oxoguanine significantly blocked human Pol iota. Nevertheless, translesion DNA synthesis opposite 8-oxoguanine was observed with increasing concentrations of purified human Pol iota, resulting in predominant C and less frequent A incorporation opposite the lesion. Opposite a template abasic site human Pol iota efficiently incorporated a G, less frequently a T and even less frequently an A. Opposite an AAF-adducted guanine, human Pol iota was able to incorporate predominantly a C. In both cases, however, further DNA synthesis was not observed. Purified human Pol iota responded to a template TT (6-4) photoproduct by inserting predominantly an A opposite the 3' T of the lesion before aborting DNA synthesis. In contrast, human Pol iota was largely unresponsive to a template TT cis-syn cyclobutane dimer. These results suggest a role for human Pol iota in DNA lesion bypass.

Roles of specialized DNA polymerases in mutagenesis by 8-hydroxyguanine in human cells

“…2). Although the incorporation of 2'-deoxyribonucleotides opposite 8-OH-Gua in template DNA by DNA pol ι in vitro was reported [31][32][33], the result obtained in this study suggests that the contribution of the DNA pol to 8-OH-Gua bypass is minimal, at best, at least in this human cell line. REV1 is a member of the Y-family DNA pols, but it acts as deoxycytidyl transferase opposite a range of damaged bases [23,24].…”

“…It has been shown that TLS DNA pols bypassed 8-OH-Gua in vitro [25][26][27][28][29][30][31][32][33][34]. Thus, to understand the mutagenesis and carcinogenesis processes by 8-OH-Gua, it is important to examine the contributions of the polymerases to the induction and suppression of the G:CT:A mutations caused by this lesion in living mammalian cells.…”

UV-B Radiation Induces Epithelial Tumors in Mice Lacking DNA Polymerase η and Mesenchymal Tumors in Mice Deficient for DNA Polymerase ι

“…5D). Several in vitro experiments have indicated that the Pol protein participates in the bypass of two major UV-induced DNA lesions, CPDs and pyrimidine pyrimidone photoproducts [(6-4)PPs] (27,60,(63)(64)(65)76). According to these previous studies, Pol protein bypasses thymine-thymine CPDs in a highly error-prone manner under certain conditions (60,63,64).…”

Miscoding properties of 1,N6-ethanoadenine, a DNA adduct derived from reaction with the antitumor agent 1,3-bis(2-chloroethyl)-1-nitrosourea

“…Studies using in vitro replication systems have revealed that the three related Y family enzymes, pols η (Rad30A), ι (Rad30B) and κ (DinB1), are capable of bypassing various DNA lesions [e.g. [28][29][30][31][32][33][34][35][36] that are normally blockers of classical DNA polymerases such as pol α and pol β. However, such translesion syntheses can be error-free or error-prone.…”

“…29,34,[41][42][43]. A unique feature of this enzyme is that it preferentially incorporates G, rather than A, opposite the template T, leading to a T:G mismatch [29,43,44].…”