(A) physiological significance of the functional interaction between mus81 and fen1 in homologous recombination repair

Abstract

Mus81-Mms4 and Fen1 are DNA endonucleases involved in processing of a variety of DNA structural intermediates formed during DNA transactions such as replication and ho-mologous recombination. Recently, it was reported that Mus81 physically interacts with Rad27 (yeast Fen1) and they are mutually stimulatory in a manner dependent on specific pro-tein-protein interactions. In this paper, we investigated the in vivo significance of the interac-tions observed between the two enzymes and showed that the N-terminal 120 amino-acid re-gion of Mus81 was required for functional interactions with Rad27. Mus81 $\Delta$ 120N lacking the N-terminal 120 amino-acid region displayed catalytic activity comparable to that wild-type when it formed a complex with Mms4, but failed to be stimulated by Rad27. Interestingly, the mus81 $\Delta$ 120N allele showed synthetic lethality when combined with sgs1$\Delta$. Considering that Sgs1 constitutes an alternative pathway in parallel to Mus81-Mms4, our results suggest that the physical and functional interactions observed between Mus81 and Rad27 are physiologically important in resolving toxic recombination intermediates. In support of this, the lethality of sgs1$\Delta$ mus81 $\Delta$ 120N was efficiently rescued by further deletion of RAD52, a key recombination mediator essential to generate the toxic intermediates. Mutagenic analyses of the N-terminal region identified two distinct motifs, named N21-26 (aa from 21-26) and N108-114 (aa from 108-114) important for the in vitro and in vivo functions of Mus81. Our findings indicate that the N-terminal region of Mus81 acts as a landing pad to interact with Rad27 and that Mus81 and Rad27 work conjointly for efficient removal of various aberrant DNA structures.