"Direct Observation of DNA Rotation During Transcription by Escherichia coli RNA Polymerase".

Yoshie Harada ^{1, 2, 6}, Osamu Ohara ³, Akira Takatsuki ¹, Hiroyasu Itoh ^{2, 4}, Nobuo Shimamoto ⁵, and Kazuhiko Kinosita Jr. ^{1, 2}

¹ Department of Physics, Faculty of Science and Technology, Keio University, Hiyoshi 3-14-1, Kohoku-ku, Yokohama 223-8522, Japan.
² CREST (Core Research for Evolutional Science and Technology), "Genetic Programming" Team 13, Nogawa 907, Miyamae-ku, Kawasaki 216-0001, Japan.
³ Kazusa DNA Research Institute, Yana 1532-3, Kisarazu 292-0812, Japan.
⁴ Tsukuba Research Laboratory, Hamamatsu Photonics KK, Tokodai, Tsukuba 300-2635, Japan.
⁵ Structural Biology Center, National Institute of Genetics, Mishima 411-8540, Japan.

⁶ Correspondence and requests for materials should be addressed to Y.H. at present address:
Department of Molecular Physiology, The Tokyo Metropolitan Institute of Medical Science, 18-22, Honkomagome 3-chome, Bunkyo-ku, Tokyo 113-8613, Japan.
E-mail:  yharada@rinshoken.or.jp

Helical filaments driven by linear molecular motors are anticipated to rotate around their axis, but rotation consistent with the helical pitch has not been observed. 14S dynein [1] and non-claret disjunctional protein (ncd) [2] rotated a microtubule more efficiently than expected for its helical pitch, and myosin rotated an actin filament only poorly [3]. For DNA-based motors such as RNA polymerase, transcription-induced supercoiling of DNA [4] supports the general picture of tracking along the DNA helix [5]. Here we report direct and real-time optical microscopy measurements of rotation rate that are consistent with high-fidelity tracking. Single RNA polymerase molecules attached to a glass surface rotated DNA for >100 revolutions around the right-handed screw axis of the double helix with a rotary torque of >5 pN nm. This real-time observation of rotation opens the possibility of resolving individual transcription steps.

1. Vale RD, and Toyoshima YY, "Rotation and translocation of microtubules in vitro induced by dyneins from Tetrahymena cilia". Cell 52: 459-469 (1988).

2. Walker RA, Salmon ED, and Endow SA, "The Drosophila claret segregation protein is a minus-end directed motor molecule", Nature 347: 780-782 (1990).

3. Sase I, Miyata H, Ishiwata S, and Kinosita K, Jr., "Axial rotation of sliding actin filaments revealed by single-fluorophore imaging", Proc. Natl. Acad. Sci. USA 94: 5646-5650 (1997).

4. Wang JC, and Lynch AS, "Transcription and DNA supercoiling", Curr. Opin. Genet. Dev. 3: 764-768 (1993).

5. Cook PR, "The organization of replication and transcription". Science 284: 1790-1795 (1999).

Additional References:

1. Frenster JH, "Selective Control of DNA Helix Openings During Gene Regulation", Cancer Res. 36: 3394-3398 (1976).