DUSP6 regulates drug sensitivity by modulating DNA damage response.

BACKGROUND: Dual specificity phosphatase 6 (DUSP6) is a member of a family of mitogen-activated protein kinase phosphatases that dephosphorylates and inhibits activated ERK1/2. Dual specificity phosphatase 6 is dynamically regulated in developmental and pathological conditions such as cancer. METHODS: Cancer cell lines were made deficient in DUSP6 by siRNA and shRNA silencing. Sensitivity to anti-EGFR and chemotherapeutic agents was determined in viability and apoptosis assays, and in xenografts established in SCID mice. Cellular effects of DUSP6 inactivation were analysed by proteomic methods, followed by analysis of markers of DNA damage response (DDR) and cell cycle. RESULTS: We determined that depletion of DUSP6 reduced the viability of cancer cell lines and increased the cytotoxicity of EGFR and other targeted inhibitors, and cytotoxic agents, in vitro and in vivo. Subsequent phosphoproteomic analysis indicated DUSP6 depletion significantly activated CHEK2 and p38, which function in the DDR pathway, and elevated levels of phosphorylated H2AX, ATM, and CHEK2, for the first time identifying a role for DUSP6 in regulating DDR. CONCLUSION: Our results provide a novel insight into the DUSP6 function in regulating genomic integrity and sensitivity to chemotherapy in cancer.

Oncoviral DNAs induce transposition of endogenous mobile elements in the genome of Drosophila melanogaster.

Previously, we have shown that particles of Rous sarcoma virus or cloned fragments of RSV cDNA as well as DNA of oncogenic simian adenovirus Sa7, injected into the polar plasm of early Drosophila melanogaster embryos, were able to induce, with high frequency, unstable visible mutations in different groups of genetic loci. The genetic instability of the recovered mutations, i.e., their ability to revert to normal state or to generate new mutant alleles at the affected locus, was manifest in mutant lines through several generations. The molecular analysis undertaken in this study of the yellow-scute loci region which is highly sensitive to the microinjected Sa7 DNA, and of the white locus, that frequently mutates under the influence of RSV cDNA, clearly shows that the induced mutations and reversions are accompanied by insertion/excision of endogenous mobile elements. This conclusion is confirmed by in situ hybridization experiments which demonstrate that the adenovirus DNA is able to change, though with different efficiency, the chromosomal localization of certain Drosophila retrotransposons. These results partially elucidate the molecular mechanism of the genetic instability in D. melanogaster induced by microinjection of oncoviruses into early embryos, implying that is results from mobilization of endogenous transposons which play the role of insertional elements directly causing unstable mutations.

A relatively small 5' regulatory region of esterase S gene of Drosophila virilis determines the specific expression as revealed in transgenic experiments.

Expression of the esterase S gene of Drosophila virilis was studied in transgenic experiments. Truncated genomic copy of this gene including 400 bp of 5' regulatory region was integrated into the genome of Drosophila melanogaster. The products of the transferred gene were detected. It was found that strict temporal and tissue specificity of the esterase S gene expression is conserved in transformed flies. The results suggest that this specificity is evidently determined by the regulatory region of the esterase S gene and controlled by cis mechanism.

[Induction of unstable mutations in Drosophila melanogaster by microinjection of oncogenic virus DNA into polar embryonic plasm. Malignant effect of oncoviral DNA]. / Induktsiia nestabil'nykh mutatsii u Drosophila melanogaster mikroin''ektsei DNK onkogennykh virusov v poliarnuiu plazmu émbrionov. Maligniziruiushchii éffekt onkovirusnykh DNK.

We have demonstrated that the ability to induce benign neoplasms We have dominant mode of inheritance in Drosophila melanogaster is the specific feature of oncoviral DNAs. It is supposed that development of this type of neoplasms in Drosophila is connected with the changes in expression of protooncogenes in mutant genome: firstly, the genetic factors directing the development of neoplasms and Drosophila protooncogenes which shared the homology with v-src are localised in the same regions; secondly, there are structural rearrangements in c-src/fps (29A) protooncogene in mutant stocks which display the ability for neoplastic growth.

[Induction of unstable mutations in Drosophila melanogaster by microinjections of oncogenic virus DNA into the embryo polar plasma. Prolonged genetic instability of mutations at the lobe locus]. / Induktsiia nestabil'nykh mutatsii u Drosophila melanogaster mikroin"ektsiei DNK onkogennykh virusov v poliarnuiu plazmu émbrionov. Prodlennaia genet'icheskaia nestabil'nost' mutatsitii v lokuse lobe.

Mutations Lobe induced by the microinjections of RSV cDNA into Drosophila melanogaster early embryos are characterized by permanent genetic instability; the level of this instability is being changed in time. Based on the results of genetic analyses of Lobe mutations and molecular analysis of white and ADH mutations induced at high frequency in this system of gene instability, we supposed that unstable mutations which arose under the influence of retroviral cDNA are of the insertional nature.

INDUCTION OF UNSTABLE MUTATIONS IN DROSOPHILA MELANOGASTER BY THE MICROINJECTION OF ONCOGENIC VIRAL DNA INTO THE POLAR PLASM OF EMBRYOS: PROLONGED GENETIC INSTABILITY OF MUTATIONS AT THE LOBE LOCUS.

MUTATIONS IN THE LOBE LOCUS WERE PRODUCED BY MICROINJECTION OF RSV CDNA INTO EARLY EMBRYOS OF D. MELANOGASTER, AND SHOWED PROLONGED GENETIC INSTABILITY, THE EXTENT OF WHICH CHANGED SIGNIFICANTLY WITH TIME. GENETIC ANALYSIS OF LOBE-RSV MUTATIONS AND A MOLECULAR-BIOLOGICAL ANALYSIS OF STRUCTURAL REARRANGEMENTS IN THE W AND ADH LOCI, WHICH ARE FREQUENTLY MUTATED BY RETROVIRAL CDNA, SHOWED THAT THE MUTATIONS RESULTED FROM INSERTIONS ARISING IN THIS GENETIC INSTABILITY SYSTEM.