Barrier-to-Autointegration Factor (BAF) evaluation during DNA-damage response.

Barrier-to-Autointegration factor (BAF) is a DNA binding protein able to interact with both the nuclear lamina and chromatin organizing proteins. It has been previously reported that BAF and emerin (its LEM-inner nuclear membrane binding protein) interact with key components of the UV-DNA-damage-binding protein complex which recognizes DNA-distorting lesions and mediate DNA-damage response. Considering that disorders with progeroid phenotype are frequently due to defects in the DNA-damage response, it is not accidental?? that a point mutation on the BAF codifying gene (BANF1 c.34G>A[p.Ala12Thr]) causes a rare progeroid syndrome named Nestor-Gullermo progeroid syndrome (NGPS). In order to improve our knowledge on BAF function during DNA-damage response, we evaluated the expression and localization of this protein during DNA-damage response due to the oxidative stress. To this aim, U2OS cells were subjected to an hydrogen peroxide (H2O2) in vivo treatment . BAF was evaluated by both biochemical and immunofluorescence techniques after acute treatment as well as during the oxidative stress recovery. Interestingly, we observed that in U2OS cells BAF, at the nuclear level, localized in the nucleoli where it colocalized with Werner protein (WRN). After the DNA-damage induction BAF protein level decreased and translocated from the nucleoli to the nucleoplasm. BAF recovered the nucleolar localization and the normal protein amount during the cell cycle arrest and during the DNA-damage marker ?-H2AX decrease. Lamin A/C was modulated during the oxidative stress response, in particular acute H2O2 treatment decreased lamin A/C protein amount while resolving of DNA damage induced lamin A/C increase and affected its intranuclear localization. Indeed, lamin A/C and BAF colocalized at the nucleoli of U2OS cells after recovering from 24/48 hours of oxidative stress. Emerin was normally located and expressed at the nuclear periphery in treated and recovered U2OS cells. PARP1, P-21 and WRN expression were also evaluated as marker of DNA-damage response. In this study we demonstrate a new BAF intranuclear localization affected by DNA-damage induction that strongly suggests a possible involvement of this DNA binding protein not only in UV-DNA-damage response but also in oxidative stress-DNA-damage response.