Unraveling non-canonical facets of telomere biology in Arabidopsis thaliana
Abstract
Telomeres protect the chromosome ends from nucleolytic attack and facilitate complete replication of DNA. Telomerase is an essential enzyme composed of catalytic subunit Telomerase Reverse Transcriptase (TERT) and a long non-coding RNA (lncRNA), Telomerase RNA (TR). Telomerase, plays an indispensable role in telomere length homeostasis and a multitude of pathways have been implicated in regulating telomeres and telomerase. For example, in Arabidopsis thaliana a novel lncRNA named TER2 was previously identified as a negative regulator of telomerase in response to DNA damage.
In this dissertation, the role of TER2 in plant telomere biology was reevaluated. TER2 was originally shown to partially overlap with the 5' UTR of tRNA Adenosine Deaminase (TAD3) gene on the complementary strand. However, updated genome annotation revealed that TER2 was wholly embedded within the TAD3 5' UTR, raising the possibility that phenotypes ascribed to TER2 could be instead derived from the TER2/TAD3 locus. Based on the results from strand-specific qRT-PCR and RNA-Seq experiments, TER2 is not a stable lncRNA, and instead appears to be a PCR artifact emanating from the TAD3 5' UTR. Further, telomerase activity assays with hypomorphic tad3 mutant revealed that the TER2/TAD3 locus is non-responsive to DNA damage. However, the tad3 mutants failed to maintain proper telomere length, despite the presence of a wild type level of telomerase activity and a wild type terminal chromosome architecture. Additional genetic analysis confirmed TAD3 contributed to telomere maintenance via a telomerase-independent mechanism. Loss of TAD3 impacts several pathways that impinge on cell cycle, metabolism, and hormone signaling, implying that TAD3 affects telomere length maintenance indirectly by influencing cell cycle and/or metabolism-related pathways in A. thaliana.
I uncovered a second non-canonical mechanism in telomere biology by demonstrating that Protection Of Telomeres 1b, one of two POT1 paralogs in A. thaliana, is required for chromatin compaction and chromosome segregation. This function appears to be related to a role for POT1b in regulating reactive oxygen species. Overall the work presented in this dissertation provides new insights into the various non-canonical pathways involved in telomere maintenance and non-canonical functions of telomere associated protein. It also expands on understanding of the interplay between telomeres and cellular metabolism.
Citation
Bose, Sreyashree (2020). Unraveling non-canonical facets of telomere biology in Arabidopsis thaliana. Doctoral dissertation, Texas A&M University. Available electronically from https : / /hdl .handle .net /1969 .1 /192712.