Wilhelm bernhard workshop on the cell nucleus

NUCLEAR MYOSIN I AND ACTIN ARE REQUIRED FOR RNA POLYMERASE I TRANSCRIPTION

¹Philimonenko V.V., ²Iben S., ¹Dingová H., ¹Kyselá K., ²Grummt I., ³de Lanerolle P., and ¹Hozák P.

¹Institute of Experimental Medicine, Academy of Sciences of the Czech Republic, Prague, Czech Republic; ²German Cancer Research Center, Heidelberg, Germany; ³University of Illinois at Chicago, USA
Recently, nuclear myosin I (NMI) was shown to be involved in RNA polymerase II transcription. We report for the first time that actin and NMI are required for rRNA synthesis. Electron microscopy reveals that both actin and NMI are localized within the nucleoli, and their distribution is highly specific. NMI colocalizes with nascent rRNA in the dense fibrillar component and its presence here is transcription-dependent. When nuclear extracts are fractionated, significant amounts of actin and NMI are detected in fractions containing partially purified Polymerase I (Pol I) and its transcription factors. Experiments of co-immuno-precipitation demonstrate association of actin with murine initiation-competent Pol I holoenzyme complexes. The rate of Pol I transcription in vitro is dramatically reduced by antibodies directed against NMI, while addition of purified NMI stimulates transcription from mouse rDNA promoter in a dose-dependent manner. Anti-actin antibodies specifically inhibit formation of full-length transcripts, but not ACU trimers in abortive initiation assay, indicating that actin plays a role in promoter clearance or transcript elongation. Microinjections of anti-actin and anti-NMI antibodies significantly reduce the level of nucleolar transcription in cultured HeLa cells, demonstrating the requirement for these proteins for transcription in vivo. We conclude that the acto-myosin molecular motor can be generally required for the movement of DNA relative to transcription complexes.