With yeast Saccharomyces cerevisiae, results from a variety of genetic and biochemical investigations have demonstrated that the REV genes play a major role in induction of mutations through replication processes that directly copy the damaged DNA template during DNA replication. However, in higher eucaryotes functions of homologues are poorly understood and appear somewhat different from the yeast case. It has been suggested that human REV1 interacts with human REV7, this being specific to higher eucaryotes. Here we show that purified human REV1 and REV7 proteins form a heterodimer in solution, which is stable through intensive purification steps. Results from biochemical analysis of the transferase reactions of the REV1-REV7 complex demonstrated, in contrast to the case of yeast Rev3 whose polymerase activity is stimulated by assembly with yeast Rev7, that human REV7 did not influence the stability, substrate specificity, or kinetic parameters of the transferase reactions of REV1 protein. The possible role of human REV7 is discussed.In yeast Saccharomyces cerevisiae, rev mutants, rev1, rev3, and rev7 were initially isolated on the basis of reduced mutations after UV treatment (1, 2) and classified into a subbranch of the RAD6 epistasis group, with alteration in the error-prone postreplication repair pathway (2-4). Yeast strains carrying rev mutations exhibit a reduced frequency of mutations following treatments with a variety of DNA damaging agents (5). The majority of mutations are induced through replication processes, which directly copy the damaged DNA template during DNA replication, a function of REV genes. Lesion bypass DNA replication is one cellular system activated in response to DNA damage that functions to prevent cell death caused by replication arrest (5).Rev1 protein, belonging to a family of translesion DNA polymerases (6, 7), contains a BRCA1 C terminus (BRCT) 1 domain (8) and possesses deoxycytidyl transferase activity in templatedirected reactions (9). It is capable of extending a primer terminus by insertion of dCMP opposite a variety of damaged bases and apurinic/apyrimidinic (AP) sites (9 -11). The in vivo role of the REV1 gene in translesion replication of AP sites has been well studied using a yeast system (12, 13), and dCMP residues are known to be incorporated opposite AP sites in the majority of bypass events in the wild type but not the rev1⌬ strain (13). Deoxycytidyl transferase activity of the Rev1 protein, first observed for the yeast enzyme (9), has been conserved throughout eucaryotic evolution (14 -18), implying a contribution to survival (5). However, the other role of the REV1 gene in the mutagenesis pathway is not due to its action as a deoxycytidyl transferase. Although the Rev1 protein does not allow bypass of T-T (6 -4) photoproducts in vitro, the gene is required for translesion DNA synthesis in vivo (13,19). A mutagenesisdeficient BRCT domain mutant encodes a protein with normal deoxycytidyl transferase activity (13), while methyl methane sulfonate-induced mutagenesis ...

CP/MAS 13C NMR analyses have been made for different frozen solutions of poly(vinyl alcohol) (PVA) samples with different tacticities to obtain further information about the intramolecular hydrogen bonding of PVA. Aqueous or deuterated dimethyl sulfoxide (DMSO-d 6 ) solutions (3 wt %) have been frozen in a rotating MAS rotor by cooling to −50 °C. The CH resonance lines of the frozen PVA solutions split in different ways, significantly depending on the tacticities and the solvents. These CH resonance lines can be well resolved into six to eight constituent lines by the least-squares method, whose lines have different chemical shifts evaluated by assuming the downfield shifts due to the intramolecular hydrogen bonding and the upfield shifts induced by the γ-gauche effect. Furthermore, the relative intensities of the lines thus resolved are successfully interpreted for the frozen atactic PVA solutions by the statistical treatment assuming the statistical formation of the intramolecular hydrogen bonds and the random distribution of the trans and gauche conformations along the PVA chains. The probabilities of the trans conformation and the intramolecular hydrogen bond are also determined through this analysis for atactic PVA samples. In contrast, such calculations based on Bernoullian statistics cannot be applied to the cases of the frozen isotactic PVA solutions, suggesting that certain specific conformations are preferably formed for the isotactic PVA samples.

No abstract