Expression of I-CreI endonuclease generates deletions within the rDNA of Drosophila.
The rDNA arrays in Drosophila contain the cis-acting nucleolus organizer regions responsible for forming the nucleolus and the genes for the 28S, 18S, and 5.8S/2S RNA components of the ribosomes and so serve a central role in protein synthesis. Mutations or alterations that affect the nucleolus organizer region have pleiotropic effects on genome regulation and development and may play a role in genomewide phenomena such as aging and cancer. We demonstrate a method to create an allelic series of graded deletions in the Drosophila Y-linked rDNA of otherwise isogenic chromosomes, quantify the size of the deletions using real-time PCR, and monitor magnification of the rDNA arrays as their functions are restored. We use this series to define the thresholds of Y-linked rDNA required for sufficient protein translation, as well as establish the rate of Y-linked rDNA magnification in Drosophila. Finally, we show that I-CreI expression can revert rDNA deletion phenotypes, suggesting that double-strand breaks are sufficient to induce rDNA magnification.
Paredes
Silvana
S
Department of Biology, Texas A&M University, College Station, Texas 77801, USA.
Maggert
Keith A
KA
D000818:Animals; D002874:Chromosome Mapping; D003433:Crosses, Genetic; D004262:DNA Restriction Enzymes / Q000235:genetics* / Q000502:physiology; D004275:DNA, Ribosomal / Q000032:analysis / Q000235:genetics*; D004330:Drosophila / Q000235:genetics*; D005260:Female; D017353:Gene Deletion; D015971:Gene Expression Regulation, Enzymologic / Q000502:physiology; D008297:Male; D008954:Models, Biological; D008957:Models, Genetic; D020411:Oligonucleotide Array Sequence Analysis
D004275:DNA, Ribosomal; D004262:DNA Restriction Enzymes; C048901:endodeoxyribonuclease CreI