Transcription termination by the eukaryotic RNA polymerase III

Abstract

RNA polymerase (pol) III transcribes a multitude of tRNA and 5S rRNA genes as well as other small RNA genes distributed through the genome. By being sequence-specific, precise and efficient, transcription termination by pol III not only defines the 3' end of the nascent RNA which directs subsequent association with the stabilizing La protein, it also prevents transcription into downstream DNA and promotes efficient recycling. Each of the RNA polymerases appears to have evolved unique mechanisms to initiate the process of termination in response to different types of termination signals. However, in eukaryotes much less is known about the final stage of termination, destabilization of the elongation complex with release of the RNA and DNA from the polymerase active center. By comparison to pols I and II, pol III exhibits the most direct coupling of the initial and final stages of termination, both of which occur at a short oligo(dT) tract on the non-template strand (dA on the template) of the DNA. While pol III termination is autonomous involving the core subunits C2 and probably C1, it also involves subunits C11, C37 and C53, which act on the pol III catalytic center and exhibit homology to the pol II elongation factor TFIIS and TFIIFα/β respectively. Here we compile knowledge of pol III termination and associate mutations that affect this process with structural elements of the polymerase that illustrate the importance of C53/37 both at its docking site on the pol III lobe and in the active center. The models suggest that some of these features may apply to the other eukaryotic pols. This article is part of a Special Issue entitled: Transcription by Odd Pols.

Figure 1. Schematic of termination mechanisms by multisubunit RNA polymerases

The mechanisms reveal three themes common to more than one pol, i) 5′-3′ exonuclease digestion of a downstream fragment of the nascent transcript attached to the polymerase, ii) weakly base pairing oligo(rU:dA) hybrid and iii) involvement of a helicase, Rho or Sen1. A) Rho mediated termination by bacterial RNA polymerase. Rho helicase binds to the nascent transcript at a C rich region and travels along the RNA in a 5′ to 3′ direction toward the polymerase to induce destabilization and termination. B) Intrinsic termination by bacterial RNA polymerase. A hairpin formed by the nascent RNA followed by a T-rich stretch on non-template strand comprise the intrinsic termination signal. Transcript release occurs within the T stretch. C) Pol II termination mechanism for poly(A)-containing mRNA-coding genes. Transcript is cleaved downstream of the poly(A) addition site by cleavage factors associated with the pol II CTD. The polymerase-attached RNA fragment is digested by Rat1/Xrn2 in 5′-3′ direction. Sen1 helicase also binds to pol II and is required for termination. This figure is over simplified to emphasize similarities with other pols. Also, there are different pathways for termination at different classes of genes (see text). D) Eukaryotic pol I termination mechanism. Most of the transcripts are terminated upstream of the Nsi1 (Ydr026C) binding site within the T stretch. Cotranscriptional cleavage of the transcript followed by processive 5′-3′ cleavage by Rat1 is also involved in termination. E) Pol III termination mechanism. A stretch of Ts on the non-template strand is sufficient to direct termination.

Figure 2. RNA polymerase III subunits and structural elements that affect termination

A) Comparison of subunit compositions of pols II and III. The termination subcomplex that shows homology to TFIIFα/β subunits, the initiation subcomplex that shows similarity to TFIIEα/β, and C11 which is homologous to RPB9 at its N terminus and to the C-terminal motif of TFIIS at its C terminus, are highlighted. B) Cartoon of surface features of pol III derived from the electron microscopic structure of pol III (EMD-1802) deposited by [91]. Blue color reflects the proposed position of the C53/37 dimerization domains and pink shows the initiation subcomplex [91]. The cartoon of the RNA:DNA template hybrid was obtained from pol II elongation structure (PDB ID: 3HOV). The non-template strand is not shown. Yellow color reflects localization of the N-terminal, Rpb9-homologous domain of C11 only. Other features discussed in the text and/or elsewhere in the figure are indicated. For other structural representations, refer to [91]; for additional models see figures 3 & 5 in [94]. C) Cartoon depicting the structural elements that are involved in transcription elongation and termination. The catalytic site along with the hybrid binding region and other elements such as the for loops, the trigger loop and bridge helix are considered as part of a larger, extended active center.