MetadataShow full item record
In this project rRNA maturation was investigated with the help of mathematical models of processing pathways from pre-rRNA to mature rRNA species. Previously described models were transferred from Excel to Mathematica. Additionally, two Mathematica based software applications were created, which help to analyse metabolic [3H]-Uracil labelling of pre-rRNA species. The first program, M Fit helps to visualize dependencies in the pre-rRNA processing. The other program S Fit tries to find a best fit of the model response to labelling time course data, hence optimizing parameter values. To validate the model anything that has an influence on the co-transcriptional cleavage is of interest, as these would have distinct effects on the 20S pre-rRNA labelling curve. A list of proteins which might play a role in A2 cleavage of the 35S was compiled and Rat1 was selected as the first candidate to investigate. All prerRNA species except the 35S pre-rRNA consist of two populations. One set created by nascent transcript cleavage (35S gets cleaved during transcription process) and one set created by released transcript cleavage (if a fully transcribed 35S pre-rRNA was released). These two species are not usually distinguishable on gels. However, with the help of the models the two different populations can be differentiated. This allows useful predictions to be made about [3H]-Uracil labelling courses in cases of high or low co-transcriptional cleavage. Experimental data for Rat1 depletion strains indeed showed an inhibition of co-transcriptional cleavage with a curve pattern as predicted by the models. Loss of another ribosome synthesis factor Srp40 was predicted to inhibit cotranscriptional pre-rRNA methylation. Of particular interest here was the effect on the 20S as this species supposed to be mostly methylated co-transcriptionally. Labelling with [3H] methionine the 20S curve for the Srp40 deletion mutant should have an earlier onset as compared to 20S curve from the corresponding wild type strain. A higher tritium response was shown for srp40Δ as compared to wild type; this might proof loss of co-transcriptional methylation.