Variability of an artificial tandem repeat Ted Pak HURS 2007 - PowerPoint PPT Presentation

Variability of an artificial tandem repeat Ted Pak HURS 2007

Variability of an artificial tandem repeat Ted Pak HURS 2007

Variability of an artificial tandem repeat Ted Pak HURS 2007

Variability of an artificial tandem repeat Ted Pak HURS 2007

Tandem Repeats ª Consecutive iterations of short sequences of DNA ª “ Junk DNA ” : Previously ignored by genomic researchers ª Highly polymorphic: very prone to errors during DNA replication …ACACACACACAC… … C… …GCC CCTAGCC CCTAGCC CCTAGCC CCTA…

Abundance in genomes ª Tandem Repeats are very common in eukaryotic genomes! 3% 63% 54% 49% 51 51 % S. cerevisae D. rerio M. musculus H. sapiens

Highly polymorphic …ACACAC… C… +1 unit -1 unit …ACACACAC… C… …ACAC … C … ª Occurs 100-1000 times more frequently than other DNA replication errors ª Causes/mechanism for this is unknown. ª Repeat expansion causes a number of human diseases.

Artificial TR Construct Transformation! ª Artificial tandem repeat was inserted into a marker gene in S. cerevisae

Switching Mechanism

Measuring switching rate

First switching data: Long AC dinucleotide repeats …AC AC AC AC AC AC AC AC… ª We first tried a basic AC dinucleotide repeat ª Longer repeats change length faster!

Confirming the trend: Long 10mer repeats …GTACGTCACT GTACGTCACT GTACGTCACT GTACGTCACT GTACGTCACT… ª Same trend is observed with 10-mers

Other trends: Purity in AC dinucleotide repeats 60% 70% 80% 90% 100% 1e-4 …AC AG AC 1e-5 Switching Rate AC AC 1e-6 AT AC 1e-7 AC… 1e-8 Purity (% bp identity to repeat unit) ª Increasing purity of a dinucleotide tract increases its variability

Sequence properties alter variability ª Several trends have been observed: ª More repeat units = é variability ª Greater purity of repeat = é variability ª Longer repeat units = é variability so given the above… ª A model should be able to be constructed that predicts the variability of a repeat based on its DNA sequence.

Predictive Computational Model Genomic Data Experimental Data Purity (%) Purity (%) Unit number Unit number Purity (%) Purity (%) Unit number Unit number

Correlation between model and experimental data Matthieu Legendre, Nathalie Pochet, Theodore Pak, and Kevin J. Verstrepen. Sequence-based estimation of minisatellite and microsatellite repeat variability. Genome Res. doi:10.1101/gr.6554007

Switching Mechanism? Paques 2001

Recombination Proteins RAD genes, the MRE complex Krogh 2004

Future directions ª Delete genes in the recombination pathway and examine effects on variability OR ª Attempt to create a more sophisticated model for Tandem Repeat variability based on experimental data

Acknowledgements Kevin J. Verstrepen, PI Matthieu Legendre, Nathalie Pochet Marcelo Vinces, Chris Brown The Verstrepen Lab My labmates Chen, Bianca, Amy, Barry, Bena… HURS, HCRP, PRISE The FAS Center for Systems Biology

Mechanisms? Long dinucleotide repeats w/ deletions 1e+0 1e-1 Switching Rate 1e-2 1e-3 1e-4 1e-5 1e-6 1e-7 MLH1 MSH2 PMS1 RAD27 RAD50 RAD51 RAD52 RAD55 RAD57 CR Deletion AC Repeats (30) Deleting genes related to the recombination pathway appears to affect switching rates.

Unit number makes a difference 2.5 2 log(delta(Switching Rate)) 1.5 1 0.5 0 MLH1 MSH2 PMS1 RAD27 RAD50 RAD51 RAD52 RAD57 -0.5 -1 -1.5 -2 Twomers Deletion Tenmers Twentymers • Examine REC pathways, hypothesize why this might be the case Additional genes. THO complex - required for transcription of repeats? •