Original Research ARTICLE
Quantitative proteomics reveals a role for SERINE/ARGININE-RICH 45 in regulating RNA metabolism and modulating transcriptional suppression via the ASAP complex in Arabidopsis thaliana
- 1Department of Bioinformatics, St. Bonaventure University, United States
- 2Department of Biochemistry, St. Bonaventure University, United States
- 3Soybean Genomics & Improvement Laboratory, Agricultural Research Service, United States Department of Agriculture, United States
- 4Animal Biosciences and Biotechnology Laboratory (USDA-ARS), United States
- 5Department of Biology, St. Bonaventure University, United States
Pre-mRNA alternative splicing is a conserved mechanism for eukaryotic cells to leverage existing genetic resources to create a diverse pool of protein products. It is regulated in coordination with other events in RNA metabolism such as transcription, polyadenylation, RNA transport, and nonsense-mediated decay via protein networks. SERINE/ARGININE-RICH 45 (SR45) is thought to be a neutral splicing regulator. It is orthologous to a component of the apoptosis and splicing-associated protein (ASAP) complex functioning to regulate RNA metabolism at multiple levels. Within this context, we try to understand why the sr45-1 mutant Arabidopsis has malformed flowers, delayed flowering time, and increased disease resistance. Prior studies revealed increased expression for some disease resistance genes and the flowering suppressor Flowering Locus C (FLC) in sr45-1 mutants and a physical association between SR45 and reproductive process-related RNAs. Here, we used Tandem Mass Tag-based quantitative mass spectrometry to compare the protein abundance from inflorescence between Arabidopsis wild-type (Col-0) and sr45-1 mutant plants. A total of 7,206 proteins were quantified, of which 227 proteins exhibited significantly different accumulation. Only a small percentage of these proteins overlapped with the dataset of RNAs with altered expression. The proteomics results revealed that the sr45-1 mutant had increased amounts of enzymes for glucosinolate biosynthesis which are important for disease resistance. Furthermore, the mutant inflorescence had a drastically reduced amount of the Sin3-associated protein 18 (SAP18), a second ASAP complex component, despite no significant reduction in SAP18 RNA. The third ASAP component protein, ACINUS, also had lower abundance without significant RNA changes in the sr45-1 mutant. To test the effect of SR45 on SAP18, a SAP18-GFP fusion protein was overproduced in transgenic Arabidopsis Col-0 and sr45-1 plants. SAP18-GFP has less accumulation in the nucleus, the site of activity for the ASAP complex, without SR45. Furthermore, transgenic sr45-1 mutants overproducing SAP18-GFP expressed even more FLC and had a more severe flowering delay than non-transgenic sr45-1 mutants. These results suggest that SR45 is required to maintain the wild-type level of SAP18 protein accumulation in the nucleus and that FLC-regulated flowering time is regulated by the correct expression and localization of the ASAP complex.
Keywords: Acinus, apoptosis and splicing-associated protein complex, Arabidopsis thaliana, Inflorescence, Quantitative Proteomics, RNA metabolism, Sin3-associated protein 18, Serine/arginine-rich 45
Received: 29 Mar 2019;
Accepted: 14 Aug 2019.
Copyright: © 2019 Chen, Rooney, Hu, Beard, Garrett, Mangalath, Powers, Cooper and Zhang. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
* Correspondence: Dr. Xiao-Ning Zhang, Department of Biochemistry, St. Bonaventure University, St. Bonaventure, United States, firstname.lastname@example.org