AUTHOR=Macovei Anca , Faè Matteo , Biggiogera Marco , de Sousa Araújo Susana , Carbonera Daniela , Balestrazzi Alma 

TITLE=Ultrastructural and Molecular Analyses Reveal Enhanced Nucleolar Activity in Medicago truncatula Cells Overexpressing the MtTdp2α Gene

JOURNAL=Frontiers in Plant Science

VOLUME=9

YEAR=2018

URL=https://www.frontiersin.org/journals/plant-science/articles/10.3389/fpls.2018.00596

DOI=10.3389/fpls.2018.00596

ISSN=1664-462X

ABSTRACT=<p>The role of tyrosyl-DNA phosphodiesterase 2 (Tdp2) involved in the repair of 5′-end-blocking DNA lesions is still poorly explored in plants. To gain novel insights, <italic>Medicago truncatula</italic> suspension cultures overexpressing the <italic>MtTdp2α</italic> gene (Tdp2α-13C and Tdp2α-28 lines, respectively) and a control (CTRL) line carrying the empty vector were investigated. Transmission electron microscopy (TEM) revealed enlarged nucleoli (up to 44% expansion of the area, compared to CTRL), the presence of nucleolar vacuoles, increased frequency of multinucleolate cells (up to 4.3-fold compared to CTRL) and reduced number of ring-shaped nucleoli in Tdp2α-13C and Tdp2α-28 lines. Ultrastructural data suggesting for enhanced nucleolar activity in <italic>MtTdp2α</italic>-overexpressing lines were integrated with results from bromouridine incorporation. The latter revealed an increase of labeled transcripts in both Tdp2α-13C and Tdp2α-28 cells, within the nucleolus and in the extra-nucleolar region. <italic>MtTdp2α</italic>-overexpressing cells showed tolerance to etoposide, a selective inhibitor of DNA topoisomerase II, as evidenced by DNA diffusion assay. TEM analysis revealed etoposide-induced rearrangements within the nucleolus, resembling the nucleolar caps observed in animal cells under transcription impairment. Based on these findings it is evident that <italic>MtTdp2α</italic>-overexpression enhances nucleolar activity in plant cells.</p>