Research

We study the mechanisms of two essential nanomachines of gene expression, RNA polymerase II (RNAP II) and the spliceosome, using brewer’s yeast as a model system.

Our work on RNAP II focuses on transcription termination as a mechanism for gene regulation. In the 1990’s we discovered a pathway which uses two RNA-binding proteins, Nrd1 and Nab3, and the RNA/DNA helicase Sen1 to terminate RNAP II transcription within ~500 base pairs from the transcription start site (see figure). The NNS pathway produces short non-coding RNAs, such as snRNAs and snoRNAs, and also regulates mRNA synthesis by attenuation. More recently, we showed that another RNA-binding protein, Hrp1, can contribute to “NNS” termination. We use genetics, genomics, cell biology and biochemistry to define the mechanism and targets of the NNS pathway.

When elongating RNAP II (gray) produces a transcript (red) that contains NNS terminator sequences, binding of these sequences by the Nrd1-Nab3-Sen1 proteins results in termination of transcription. Mutations in these proteins, and in Hrp1 and the Rpb3 subunit of RNAP II, can cause transcription to continue through NNS terminators.

Our studies on the spliceosome focus on U6 snRNA and the dynamic RNA-RNA and RNA-protein interactions required for its pairing first with U4 snRNA and then with U2 snRNA and the intron during activation of the spliceosome for intron excision (Figure A). U2 and U6 snRNAs make up the catalytic core of the spliceosome and align the intron sequences for the first step of splicing. An internal stem-loop in U6, the ISL, is unwound for pairing with U4 and reforms upon pairing with U2. In the 1990’s we showed that stabilization of the ISL at the position indicated with an asterisk creates a cold-sensitive block to U4/U6 snRNP formation. Recently, a mutation in human U6 snRNA that is nearly identical to our earlier mutation (Figure B) was associated with the degenerative eye disease retinitis pigmentosa. Our studies suggest a possible therapeutic strategy for individuals with this mutation, or related mutations in U4 snRNA or tri-snRNP proteins. (See Brow 2026).