Repression of Gene Expression by an Exogenous Sequence Element Acting in Concert with a Heterogeneous Nuclear Ribonucleoprotein-Like Protein, Nrd1, and the Putative Helicase Sen1

Abstract

We have fortuitously identified a nucleotide sequence that decreases expression of a reporter gene in the yeast Saccharomyces cerevisiae 20-fold when inserted into an intron. The primary effect of the insertion is a decrease in pre-mRNA abundance accompanied by the appearance of 3′-truncated transcripts, consistent with premature transcriptional termination and/or pre-mRNA degradation. Point mutations in the cis element relieve the negative effect, demonstrating its sequence specificity. A novel yeast protein, named Nrd1, and a previously identified putative helicase, Sen1, help mediate the negative effect of the cis element. Sen1 is an essential nuclear protein that has been implicated in a variety of nuclear functions. Nrd1 has hallmarks of a heterogeneous nuclear ribonucleoprotein, including an RNA recognition motif, a region rich in RE and RS dipeptides, and a proline- and glutamine-rich domain. An N-terminal domain of Nrd1 may facilitate direct interaction with RNA polymerase II. Disruption of the NRD1 gene is lethal, yet C-terminal truncations that delete the RNA recognition motif and abrogate the negative effect of the cis element nevertheless support cell growth. Thus, expression of a gene containing the cis element could be regulated through modulation of the activity of Nrd1. The recent identification of Nrd1-related proteins in mammalian cells suggests that this potential regulatory pathway is widespread among eukaryotes.