The Nrd1-Nab3-Sen1 (NNS) complex carries out the RNA polymerase II (Pol II) transcription termination of non-coding RNAs (ncRNAs) in yeast, although the detailed interactions among its subunits remain obscure. In this dissertation, we have identified three sequence motifs in Sen1 that mediate direct interactions with the Pol II CTD interaction domain (CID) of Nrd1, determined the crystal structures of these Nrd1 interaction motifs (NIMs) bound to the CID, which elucidated the molecular basis for their recognition by Nrd1 CID, and characterized the interactions in vitro and in yeast. Although the Sen1 NIMs are not essential for supporting viability from the in vivo studies, termination defects were observed from NIM deletions in a reporter assay. In addition, the conservation of Sen1 NIMs suggests these interactions are very likely to promote NNS function.
This dissertation also describes the structural studies of the flowering time control protein FPA in plants, which regulates the alternative 3’-end processing of the FLOWERING LOCUS C (FLC) antisense RNA. FPA belongs to the split ends (SPEN) family of proteins, which contain N-terminal RNA recognition motifs (RRMs) and a SPEN paralog and ortholog C-terminal (SPOC) domain. The SPOC domain is highly conserved among FPA homologs in plants, but the conservation with the domain in other SPEN proteins is much lower. We have determined the crystal structure of Arabidopsis thaliana FPA SPOC domain at 2.7 Å resolution. Structural and sequence analyses identify a surface patch that is conserved among plant FPA homologs. Mutations of two residues in this surface patch did not disrupt FPA functions, suggesting that either the SPOC domain is not required for the role of FPA in regulating RNA 3’-end formation or the functions of the FPA SPOC domain cannot be disrupted by this combination of mutations.
| Identifer | oai:union.ndltd.org:columbia.edu/oai:academiccommons.columbia.edu:10.7916/d8-n4dw-1684 |
| Date | January 2019 |
| Creators | Zhang, Yinglu |
| Source Sets | Columbia University |
| Language | English |
| Detected Language | English |
| Type | Theses |
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