Pif1 helicase directs eukaryotic Okazaki fragments toward the two-nuclease cleavage pathway for primer removal

Eukaryotic Okazaki fragment maturation requires complete removal of the initiating RNA primer before ligation occurs. Polymerase delta (Pol delta) extends the upstream Okazaki fragment and displaces the 5' end of the downstream primer into a single-nucleotide flap, which is removed by FEN1 nuclease cleavage. This process is repeated until all RNA is removed. However, a small fraction of flaps escapes cleavage and grows long enough to be coated with RPA and requires the consecutive action of the Dna2 and FEN1 nucleases for processing. Here we tested whether RPA inhibits FEN1 cleavage of long flaps as proposed. Surprisingly, we determined that RPA binding to long flaps made dynamically by Pol delta only slightly inhibited FEN1 cleavage, apparently obviating the need for Dna2. Therefore, we asked whether other relevant proteins promote long flap cleavage via the Dna2 pathway. The Pif1 helicase, implicated in Okazaki maturation from genetic studies, improved flap displacement and increased RPA inhibition of long flap cleavage by FEN1. These results suggest that Pif1 accelerates long flap growth, allowing RPA to bind before FEN1 can act, thereby inhibiting FEN1 cleavage. Therefore, Pif1 directs long flaps toward the two-nuclease pathway, requiring Dna2 cleavage for primer removal

Phototransformation of three herbicides: chlorbufam, isoproturon and chlorotoluron: influence of irradiation on toxicity

Abstract. The direct phototransformations in aqueous solution of chlorbufam (I), chlorotoluron (II), and isoproturon (III) are compared. III With I and II photohydrolysis (substitution of chlorine atom with formation of the corresponding hydroxylated derivative) is initially almost quantitative. It does not depend on irradiation wavelength in the range 250-300 nm. In contrast with III the main reaction is a free radical rearrangement, the reaction being more efficient at 254 nm than at wavelengths longer than 270 nm. Compounds II and III were also irradiated on sand in the absence of water. Several photoproducts were identified, they result from elimination or oxidation of methyl groups of urea moiety. The toxicity of photoproducts was determined using the standard Microtox test (toxicity on the bacterium Vibrio fischeri). Photohydrolysis of I and II does not change much the toxicity of solutions whereas the oxidation, or at a lower scale the elimination of methyl group on the urea moiety, dramatically increases the toxicity

Phototransformation of halogenoaromatic derivatives in aqueous solution

The photochemical behaviour of monohalogeno-phenols and -anilines is highly dependent on the position of the halogen on the ring, but most often it is not significantly influenced by the nature of the halogen (Cl, Br, F). Photohydrolysis is the main reaction observed with 3-halogenated and it is almost specific. With 2-halogenated, photohydrolysis and photocontraction of the ring compete, the latter being very efficient with 2-halogeno-phenolates