br Materials and methods br Results br Discussion

Materials and methods
Results
Discussion Human MutL-family complexes (MutLα: MLH1-PMS2, MutLβ:MLH1-PMS1, MutLγ:MLH1-MLH3) exhibit multiple molecular activities including DNA binding, DNA cleavage, ATP binding and hydrolysis [43]. In our recent study, we have uncovered an important function of MLH1 in safeguarding the integrity of genomic DNA. MLH1 represses the aberrant formation of ITSs in telomerase-expressing cells. Our data also suggest that the DNA damage response function of hMLH1 may be involved in repressing ITS formation [22]. Given the importance of the ATPase activity of MutL in DNA damage response, here, we select five separation-of-function mutants in hMLH1 to determine the role of the ATPase in ITS suppression. Results in the present study reveal that the ATPase-deficient mutants (E34A and G67R) fail to suppress ITS formation, while mutants retaining the ATPase activity (H112D, G244D, H329P) preserve MLH1’s ability to suppress ITS formation (Fig. 4). Our results therefore suggest that the ATPase activity of MLH1 is required for ITS suppression. Furthermore, we also find that the ATPase activity of MLH1 is involved in suppressing the formation of micronuclei, which is one of the characteristics indicative of GSK2292767 instability. Telomeres cap chromosome ends and prevent them from degradation and recombination [44]. However, if canonical telomeric repeats sequences are formed at intra-chromosomal regions, these sequences may contribute to chromosome breakage, recombination and rearrangement [22,[26], [27], [28], [29], [30], [31]]. In Saccharomyces cerevisiae, the Pif1 helicase inhibits de novo telomerase addition of telomeric sequences at DSBs [45]. It has been reported that yeast Pif1 reduces the level of telomerase binding to telomeres by its ATPase activity, suggesting that the ATPase activity of Pif1 is essential for inhibiting the addition of telomeric sequences at DSBs [45,46]. Our result that the ATPase activity of hMLH1 is required for preventing aberrant ITS formation is reminiscent to the function of Pif in yeast. It is noticeable that etoposide-induced MLH1 colocalization with 53BP1 foci is partial (Fig. 3B) [22], in agreement with the previous study showing that only a subset of MLH1 molecules are recruited to DSB stripes induced by laser microbeam irradiation [21]. The same is also true for MSH2 and MSH3 [21]. In addition, the number of MLH1 foci markedly outnumbers 53BP1 foci (Fig. 3B), suggesting that MLH1 molecules are abundant and not all MLH1 molecules are needed at DSBs. The interactome of MLH1 has been shown to contain proteins involved not only in MMR and DSB repair, but also in other functions including protein import/export, ubiquitin pathway, cell cycle, cellular signaling, and others [41]. Thus, it is highly possible that the DSB-independent MLH1 molecules function in other biological pathways.
Author contributions
Conflict of interests
Acknowledgement
Introducion Mycotoxins are chemical contaminants produced mainly by fungi belonging to Aspergillus, Fusarium and Penicillum genera on a variety of foods and in specific conditions of humidity and temperature, both in the field and storage [1]. Mycotoxins are found all over the world as natural contaminants of numerous food of plant origin, principally in cereal grains, and can be found in food made from animals fed on contaminated feed, such as eggs, meat, milk and dairy products [2]. Mycotoxin exposure via food and feed may result in diverse adverse health effects such as carcinogenicity, immunotoxicity, reproductive toxicity and hepatotoxicity [3], as well as alterations on energy homeostasis linked with adenosine triphosphate (ATP) depletion [4]. However, the pathways associated with impairment on energy homeostasis in animals contaminated with mycotoxins remains poorly understood. One example is the involvement of creatine kinase/phosphocreatine system (CK/PCr), an essential pathway for the maintenance of bioenergetic homeostasis.