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Identification of novel putative regulators of the major apoptotic nuclease DNA Fragmentation Factor

100%

Hanus J., Kalinowska-Herok M., Widlak P.

2010

vol. 57

issue 4

521-527

Yeast two- and three-hybrid systems were used to screen cDNA libraries from HeLa cells and human brain tissue to identify novel protein partners of DNA Fragmentation Factor, the major apoptotic nuclease. The two-hybrid system revealed the DFF45 inhibitory subunit of the nuclease as the only identified partner of the DFF40 catalytic subunit. Similar analysis revealed several protein candidates that potentially interact with the DFF45 subunit: FBXO28, FOSL1, PGK1, PCNT, FHL1 and GFAP. Recombinant GFAP protected DFF45 against cleavage with caspase-3 and prevented activation of the DFF nuclease in vitro. In addition, three-hybrid system results revealed a putative novel protein partner of the DFF40-DFF45 heterodimer. The candidate cDNA contained two open reading frames that mapped to an intron of the GBF1 gene. Products of the candidate cDNA derived from a cell-free transcription/translation system inhibited DNA cleavage by recombinant caspase-activated DFF. This putative partner of DFF may have functional importance in regulating the apoptotic response because its RNAi silencing facilitated cleavage of the DFF45 inhibitor subunit and affected chromatin fragmentation in HeLa cells undergoing apoptosis. This hypothetical protein, named DRIG based on an acronym specifying its genomic location, could be a novel factor involved in regulation of DFF40 apoptotic nuclease.

Badanie wpływu Selolu na ekspresję genów kodujących transportery błonowe i enzymy metabolizujące leki w komórkach nowotworowych wrażliwych i opornych

72%

Dudkiewicz Wilczyńska J., Grabowska A., Książek I., Nowak K., Anuszewska E.

Annales Academiae Medicae Silesiensis

2011

vol. 65

issue 5-6

7-13

The objective of this study was to demonstrate diff erences in the gene expression of human cervical cancer cells (HeLa) and vinblastine-resistant KB-V1 subline treated with doxorubicin alone and combination of Selol 5% and doxorubicin. Ongoing studies seek to clarify the mechanism of action of Selol in diff erent types of cancer cells, including those which show multidrug resistance. Cells treatment with the tested compounds in the group of genes tested in HeLa cells causes other changes than in KB-V1 cells. In the resistant cells, exposure to Selol 5% and doxorubicin, released the cytotoxic eff ects by changing the expression of ABCC2 and BCL2L1 genes. The observed dependence also allows better understanding the molecular mechanisms of resistance in the KB-V1 cell line.

Celem pracy było wykazanie różnic w ekspresji genów komórek ludzkiego nowotworu szyjki macicy (HeLa) i opornej na winblastynę podlinii KB-V1, poddanych działaniu samej doksorubicyny oraz po łącznym podaniu Selolu 5% i doksorubicy. Prowadzone badania zmierzają do wyjaśnienia mechanizmu działania Selolu w różnych typach komórek nowotworowych, w tym opornych wielolekowo. Poddanie komórek działaniu testowanych związków powoduje inne zmiany w grupie badanych genów w komórkach HeLa niż w komórkach KB-V1. Łączne podanie Selolu 5% i doksorubicyny wyzwala efekt cytotoksyczny w komórkach opornych KB-V1, co przypuszczalnie jest związane ze zmianą ekspresji genów ABCC2 i BCL2L1. Zaobserwowana zależność pozwala także lepiej zrozumieć molekularne podłoże oporności komórek linii KB-V1.

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1 Acta Biochimica Polonica

1 Annales Academiae Medicae Silesiensis

1 2011

1 2010

1 Anuszewska E.

1 Dudkiewicz Wilczyńska J.

1 Grabowska A.

1 Hanus J.

1 Kalinowska-Herok M.

1 Książek I.

1 Nowak K.

1 Widlak P.

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Identification of novel putative regulators of the major apoptotic nuclease DNA Fragmentation Factor

Badanie wpływu Selolu na ekspresję genów kodujących transportery błonowe i enzymy metabolizujące leki w komórkach nowotworowych wrażliwych i opornych