Senior Project Advisor
Large extrusion bodies in Piwi family protein knockout strains in Tetrahymena thermophila
The ciliate Tetrahymena thermophila encodes twelve distinct proteins of the PIWI family of small RNA binding proteins. Three of these Tetrahymena PIWI proteins (Twis) have previously been shown to be expressed in vegetative growth and bind predominantly to ~23-24 nucleotide (nt) small RNAs (sRNAs) (Couvillion et al., 2009). One of ~23-24 nt sRNA binding proteins, Twi2, is encoded by DNA sequence that has high sequence similarity to four other predicted Twi genes (Twis 3-6) which are normally not highly expressed. Using fluorescent microscopy imaging techniques, we examined mutant strains TWI8, TWI2-6Δ, TWI7Δ, TWI2-6/8Δ, and TWI7/8Δ for accumulation of large extrusion bodies (EBs). EBs are nonnuclear DNA containing structures which are likely akin to mammalian micronuclei; these are structures commonly found as markers of genomic instability. We found no significant EB size increase in the strains TWI2-6Δ and TWI7Δ and a moderately enlarged EB phenotype in TWI8 cells compared to the reference parental strain SB210. TWI2-6/8Δ cells had the most exaggerated enlarged EB phenotype, suggesting that at least one of the five Twi proteins encoded by the TWI2-6 gene cluster contributes to genome stability in addition to Twi8. TWI7/8Δ cells had highly variable EB area between replicates with the average EB being larger than that found in SB210, which may also implicate Twi7 in contributing to genome stability in absence of Twi8. Overall, these findings support a role for RNA interference (RNAi) in maintaining genome integrity in Tetrahymena that is dependent on Twi proteins.
Sasser, Jason, "Large extrusion bodies in Piwi family protein knockout strains in Tetrahymena thermophila" (2021). WWU Honors College Senior Projects. 524.
Subjects - Topical (LCSH)
RNA; Ciliata; Piwi genes; Nucleotide sequence
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