Poster abstracts

Poster number 49 submitted by Kathryn Holzmacher

Stress-Induced Increase in Small ncRNAs Generated from Regulatory Regions of the Mitochondrial Genome

Kathryn E. Holzmacher (Department of Biochemistry at University at Buffalo), Jonathan E. Bard (Center of Excellence in Bioinformatics and Life Sciences, and Department of Biochemistry at University at Buffalo), Theodore L. Mathuram (Department of Biochemistry at University at Buffalo), Natalie A. Lamb (Center of Excellence in Bioinformatics and Life Sciences at the University at Buffalo), Norma J. Nowak (Center of Excellence in Bioinformatics and Life Sciences, and Department of Biochemistry at University at Buffalo), A. Blumental-Perry (Department of Biochemistry at University at Buffalo)

Abstract:
Rationale: Alveolar Epithelial Type (AET) 2 cells serve as progenitors for the repair of destructed alveoli. We identified AET2-specific mitochondria-associated retrograde signaling via mtDNA-encoded non-coding (nc) RNA, mito-ncR-805. mito-ncR-805 levels are increased in the mitochondria and nucleus during stress, and positively regulate mitochondrial metabolism and respiration. We propose that mito-ncR-805 represents a subtype of mitochondrial small RNAs that protect cells under stress. To identify transcripts with similar functions we compared mitochondrial transcriptomes of unstressed and stressed cells.
Methods: CS-exposures of MLE12 cells were used to study adaptational transcriptomes. Stress signaling was evaluated by Western blot. Mitochondrial small RNA transcriptome was evaluated by constructing and analyzing strand-specific RNAseq libraries of transcripts 100 bp and shorter. A bioinformatic pipeline was specifically developed for the analysis of mitochondrial transcripts. Identified transcripts were validated by Northern blot and qPCR analyses.
Results: CS exposure increased the generation of 3 small RNAs. The transcripts had a specific processing pattern and originated from the control regions of the mitochondrial genome (D-loop and Rnr genes). The presence of the transcripts in cells and their elevation during smoke exposure was confirmed by Northern. Mitochondrial origin was validated in either mitochondrial-depleted cells (grown in EtBr), or in cells where mitochondrial transcription was inhibited by 6G-rhodamine. Cellular localization of Rnr-derived transcripts is currently being evaluated.
Conclusion: We demonstrate the stress-dependent generation of 4 distinct transcripts that originate from control regions of the mitochondrial genome. This suggests a possible regulatory function in coordinating the activities in both mitochondrial and nuclear genomes. We propose that those small RNAs aid in signaling which aim to restore mitochondrial homeostasis.

References:
Blumental-Perry A, Jobava R, Bederman I, Degar AJ, Kenche H, Guan BJ, Pandit K, Perry NA, Molyneaux ND, Wu J, Prendergas E, Ye ZW, Zhang J, Nelson CE, Ahangari F, Krokowski D, Guttentag SH, Linden PA, Townsend DM, Miron A, Kang MJ, Kaminski N, Perry Y, Hatzoglou M. Retrograde signaling by a mtDNA-encoded non-coding RNA preserves mitochondrial bioenergetics. Commun Biol. 2020 Oct 30;3(1):626. doi: 10.1038/s42003-020-01322-4. PMID: 33127975; PMCID: PMC7603330.
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Keywords: Alveolar Epithelial Type 2 Cells, Mitochondrial Stress Signaling , Small non-coding RNA