Programmed Elimination of Internal Chromosomal Sequences and Fragmented Chromosomes in Tetrahymena thermophila

• Main Authors: Chih-Yi Lin, 林之儀
• Other Authors: Meng-Chao Yao
• Format: Others
• Language: en_US
• Published: 2017
• Online Access: http://ndltd.ncl.edu.tw/handle/v9q5d3

博士 === 國立臺灣大學 === 基因體與系統生物學學位學程 === 105 === Maintenance of chromosome integrity is crucial for genetic stability. However, programmed DNA rearrangements are known to occur in many organisms. In the ciliate Tetrahymena, thousands of deletion sequences are removed and the five germline chromosomes are fragmented into hundreds of minichromosomes during somatic nuclear differentiation. Although RNA is known to target the region for deletion, it is not clear how the precise boundaries are determined. Previous studies have shown that flanking cis-acting sequences determine the boundaries of some elements and have suggested that there are several cis-acting elements, each affecting a different group of internal elimination sequences (IESs). Here, we compare IESs and their boundary variations among three wild type Tetrahymena strains by genome sequencing. We show that the IESs are highly conserved among strains even though their boundaries are variable. Furthermore, we have identified several flanking inverted repeats (IRs) at both ends of several IES groups. Localization of these IRs was consistent in groups, suggesting that they could serve as the cis-acting elements. Recently a novel G-quadruplex-binding protein, Lia3, was found to determine the precise boundary of the M element and several IESs flanked by a cis-acting element, A5G5. To further investigate the relationship between cis-acting elements and their binding proteins, we compared the IES boundary variation between wild type strains and Lia3 progeny to identify the group of IES that is affected by Lia3. We found that the G-rich and C-rich IRs were highly correlated with the boundary determination of Lia3-affected IESs. More interestingly, the distances of these IRs to the IES boundaries were consistent even if the boundaries varied among different IES forms, suggesting that the cis-acting elements were the determining factor of IES boundary determinations. Next, we found that the fates of minichromosomes differ after chromosome breakage. Of the 326 somatic minichromosomes identified using genomic data, 50 were selectively eliminated from the mature somatic genome. Interestingly, many and probably most of these minichromosomes are eliminated during the growth period between 6 and 20 doublings right after conjugation. Genes with potential conjugation-specific functions were found in these minichromosomes. Our findings have revealed a new mode of programmed DNA elimination in ciliates similar to those observed in parasitic nematodes, which could play a role in developmental gene regulation. The results of this thesis provide a global picture of programmed DNA rearrangement and the mechanism of DNA diminution by internal sequence deletions and minichromosome eliminations in Tetrahymena.