Poster abstracts

Poster number 48 submitted by Laura Ritchey

Hidden Breaks in Rice Chloroplast rRNA Revealed by Structure-seq2

Kaylin M. Gaudette (Chemistry Department, University of Pittsburgh at Johnstown), Colby K. Hillegass (Chemistry Department, University of Pittsburgh at Johnstown), Cooper J. Halliday (Biology Department, University of Pittsburgh at Johnstown), Laura E. Ritchey (Chemistry Department, University of Pittsburgh at Johnstown)

Abstract:
Within plants, chloroplasts are light-harvesting organelles that retain their own limited genomes. Part of this genome encodes chloroplast ribosomal RNA (rRNA) that is used to translate chloroplast-specific genes. Interestingly, the large subunit (23S) of the chloroplast rRNA harbors hidden breaks. (1-3) These covalent breaks in the backbone of the RNA are thought to be caused by endonucleases during or after assembly of the ribosome such that the ribosome structure remains intact. It has been proposed that hidden breaks are necessary for efficient translation in the chloroplast, and they have been identified in rRNAs of a select few organisms. By analyzing control Structure-seq2 data, (4-6) we were able to identify the presence of these hidden breaks in rice chloroplast rRNA and bacterial rRNA. We have also found evidence that the extent that the break occurs may be affected by stress conditions. Future plans are to study the extent of these hidden breaks in the chloroplast, mitochondria, and/or bacterial rRNA of various organisms to identify any trends in either sequence or structure. We will also study how stress affects the extent of breakage to begin to elucidate how these breaks affect translation. Currently, initial results in stress-treated rice, coffee, and spinach using reverse transcription and PCR spanning the hidden break will be presented. Preliminary results indicate that these hidden breaks are not found in every rRNA molecule. Advice on additional appropriate assays to perform at a primarily undergraduate institute with limited resources would be much appreciated.

References:
1. Nishimura, K., et al. Plant J 63 (5), 766-777. (2010)
2. Liu, J., et al. Plant Physiol 168 (1), 205-221, (2015).
3. Bieri, P., et al. EMBO J 36 (4), 475-486, (2017).
4. Ritchey, L.E. et al. Nucleic Acids Res 45 (14), e135 (2017).
5. Su, Z,... Ritchey, L.E. et al. PNAS.115 (48), 12170-12175, (2018)
6. Ritchey, L.E., et al. RNA 26 (10), 1431-1447, (2020)

Keywords: RNA structure, rRNA, chloroplast