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!

51 51 %

49% 54% 63% 3%

• S. cerevisae
• D. rerio
• M. musculus
• H. sapiens

Highly polymorphic

…ACACAC… C… …ACACACAC… C… …ACAC … C …

+1 unit

• 1 unit

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

ª Artificial tandem repeat

was inserted into a marker gene in S. cerevisae

Transformation!

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

ª Increasing purity of a dinucleotide tract

increases its variability

…AC AG AC AC AC AT AC AC…

1e-8 1e-7 1e-6 1e-5 1e-4 60% 70% 80% 90% 100%

Purity (% bp identity to repeat unit) Switching Rate

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

Unit number Unit number Unit number Unit number Purity (%) Purity (%) Purity (%) Purity (%)

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

Deleting genes related to the recombination pathway appears to affect switching rates.

1e-7 1e-6 1e-5 1e-4 1e-3 1e-2 1e-1 1e+0 MLH1 MSH2 PMS1 RAD27 RAD50 RAD51 RAD52 RAD55 RAD57 CR

Deletion Switching Rate AC Repeats (30)

Unit number makes a difference

• 2
• 1.5
• 1
• 0.5

0.5 1 1.5 2 2.5 MLH1 MSH2 PMS1 RAD27 RAD50 RAD51 RAD52 RAD57

Deletion log(delta(Switching Rate))

Twomers Tenmers Twentymers

• Examine REC pathways, hypothesize why this might be the case
• Additional genes. THO complex - required for transcription of repeats?