Abundance and distribution of RNA polymerase II in Arabidopsis interphase nuclei.
RNA polymerase II (RNAPII) is responsible for the transcription of most eukaryotic protein-coding genes. Analysing the topological distribution and quantification of RNAPII can contribute to understanding its function in interphase nuclei. Previously it was shown that RNAPII molecules in plant nuclei form reticulate structures within euchromatin of differentiated Arabidopsis thaliana nuclei rather than being organized in distinct 'transcription factories' as observed in mammalian nuclei. Immunosignal intensity measurements based on specific antibody labelling in maximum intensity projections of image stacks acquired by structured illumination microscopy (SIM) suggested a relative proportional increase of RNAPII in endopolyploid plant nuclei. Here, photoactivated localization microscopy (PALM) was applied to determine the absolute number and distribution of active and inactive RNAPII molecules in differentiated A. thaliana nuclei. The proportional increase of RNAPII during endopolyploidization is confirmed, but it is also shown that PALM measurements are more reliable than those based on SIM in terms of quantification. The single molecule localization results show that, although RNAPII molecules are globally dispersed within plant euchromatin, they also aggregate within smaller distances as described for mammalian transcription factories.
Leibniz Institute of Plant Genetics and Crop Plant Research (IPK) Gatersleben, D-06466 Stadt Seeland, Germany schubertv@ipk-gatersleben.de.
Veit
V
Schubert
Carl Zeiss Microscopy GmbH, D-07743 Jena, Germany.
Klaus
K
Weisshart
D017360:Arabidopsis / Q000235:genetics* / Q000378:metabolism; D029681:Arabidopsis Proteins / Q000235:genetics* / Q000378:metabolism; D002467:Cell Nucleus / Q000378:metabolism; D007399:Interphase; D011123:Polyploidy; D012319:RNA Polymerase II / Q000235:genetics* / Q000378:metabolism
D029681:Arabidopsis Proteins; D012319:RNA Polymerase II
Arabidopsis thaliana; RNA polymerase II; endopolyploidy; photoactivated localization microscopy; structured illumination microscopy; transcription factory.