Hans Mehlin
Laboratory of Molecular Genetics, Department of Cell and Molecular Biology,
Karolinska Institutet, Box 60400, S-104 01 Stockholm

Premessenger RNA, synthesized in the nucleus, is complexed with proteins to form RNA- protein (RNP) particles and is actively transported to the cytoplasm through the nuclear pore complexes (NPCs). In the salivary gland cells of the dipteran Chironomus tentans it is possible to follow the assembly and transport of a specific RNP particle, synthesized on the Balbiani ring (BR) genes. The aim of this investigation has been to characterize the translocation of BR-RNP through the NPC.

The structure of the BR-RNP particle can be studied using electron microscope tomography, a general method for three dimensional reconstruction of aperiodic objects. The RNP particle in the nucleus is roughly spherical with a diameter of 50 nm. It can be described as an RNP ribbon bent into a ring-like configuration.

During translocation through the NPC, the RNP particle gradually becomes elongated and finally rod-shaped with a length of 135 nm and a width of 25 nm. The translocating RNP particle was shown to be a ribbon rather than a rod, and the structural change during translocation can be described as a relaxation of the bent ribbon. The RNP particle is translocated through the center of the NPC and the width of the particle corresponds to the maximum diameter of the NPC channel for active transport.

During an early stage of translocation, the nucleoplasmic part of the translocating RNP particle is still spherical, constituting an intact remainder of the spherical particle in the nucleoplasm. The different regions of the particle could be recognized and it was concluded that the 5' end leads the transport through the NPC. The 3' containing domain of the particle makes contact with the inner ring of the NPC. This contact varies however, due to rotation of the particle concomitant with the translocation process. The observation that the 5' end interacts with the center of the NPC, and that no stable contact is detected at the 3' end, suggests that the cap structure rather than the poly(A) region, is important at the early stage of translocation.

The RNP particle gradually unfolds during translocation and a thin RNP fiber can be seen entering the cytoplasm, first forming a zig-zag pattern and later straightening out and engaging in polysome formation.

A 7 nm fiber was identified as the basic structural element in the RNP particle. We propose that the fiber is built from a filamentous rod of polymerized proteins with the RNA being wound around it.

Keywords: RNA transport, translocation, RNP structure, nuclear pore complex, three-dimensional reconstruction, electron microscope tomography

ISBN 91-628-0744-7     Stockholm 1992

Vårboda Studio, Bäcktorpsvägen 23, 132 34 Saltsjö-Boo,