Antisense in Abundance: The Ribosome as a Vehicle for Antisense RNA

Abstract

We have developed in Tetrahymena thermophila a new vehicle for introducing antisense RNAs into cells, the “antisense ribosome”. This system allows antisense RNAs to be expressed and to function as part of a very stable and abundant RNA molecule, the large subunit ribosomal RNA (rRNA), without adversely affecting rRNA function (1). Unlike almost all other organisms, the ciliate T. thermophila contains a single copy of its rRNA genes (rDNA) in its silent germline genome (2). It has multiple copies in the form of short, linear chromosomes in its transcribed somatic genome (3, 4). This unique situation plus a transformation system that allows complete replacement of the somatic rDNA have provided convenient ways to study rDNA through traditional and modern genetic methods. The single germline copy of the rDNA makes classical genetic studies possible (5, 6). With the transformation system, rDNA can be altered as desired in vitro and used to transform cells. If functional, the transforming rDNA can totally replace the somatic rDNA in transformed lines (7, 8). A series of studies (8–11) have revealed interesting features of rRNA variable regions leading to the realization that rRNA can be exploited to serve as a carrier of RNA sequences, such as antisense sequences, that can be designed to exert specific effects in the cell. This method of presenting an antisense RNA to a cell may magnify its effects on gene expression since rRNA is very abundant and stable and is in close physical proximity to mRNAs. This article will summarize the relevant features of the rRNA variable regions and describe the creation of “antisense ribosomes” and their potential applications in T. thermophila and other organisms.