The goal of this research project is to understand the molecular basis of trinucleotide repeat instability. Expansions of trinucleotide repeats cause 13 inherited disorders in humans. The expanded trinucleotide repeats are genetically unstable, changing in length when transmitted from parents to offspring (intergenerational instability). Although various trinucleotide sequences are capable of expanding to cause disease, most trinucleotide repeat disorders involve CAG/CTG tracts. Transgenic mice carrying CAG/CTG repeat expansions that are markedly unstable in humans do not typically show intergenerational instability when introduced on cDNA constructs. We recently generated yeast artificial chromosome (YAC) transgenic mice that show intergenerational instability with CAG repeats from the human androgen receptor (AR) locus. These AR YAC transgenic mice are unique in that they show intergenerational instability with CAG repeats from the human androgen receptor (AR) locus. These AR YAC transgenic mice are unique in that they show intergenerational instability with only 45 CAG repeats. Repeat transgenic mice are unique in that they show intergenerational instability with only 45 CAG repeats. Repeat instability in the AR YAC transgenic mice suggests that cis-acting information at the human AR locus must be retained to obtain intergenerational changes in CAG/CTG tract length. The AR YAC transgenic mice shown an effect of parental sex and parental age upon repeat instability, suggesting that trans-acting factors regulate intergenerational repeat instability. To efficiently study repeat instability in mice, we have developed the capacity to sort single sperm cells and determine their CAG repeat lengths with the polymerase chain reaction (PCR). We now wish to examine whether cis- acting elements and trans-acting factors are involved in intergenerational repeat instability in the AR YAC transgenic mice. First, we propose to test the hypothesis that trinucleotide repeat instability in the AR YAC transgenic mice is due to human genomic DNA sequences (""""""""instability elements"""""""") instability in the AR YAC transgenic mice is due to human genomic DNA sequences (""""""""instability elements"""""""") flanking in the CAG repeat expansion. If we can confirm that repeat instability depends upon genomic context, then we plan to identify the cis-acting sequences at the human AR locus that permit trinucleotide repeat instability in mice. Second, we propose to test the hypothesis that trans-acting factors in the mouse regulated trinucleotide repeat instability the placing the unstable CAG/CTG tracts onto different murine strain backgrounds. If trinucleotide repeat instability is significantly altered in one or more of the congenic strains that we generate, than we plan to identify the genetic loci responsible for the observed alterations by carrying out directed breeding experiments. By studying the molecular basis of intergenerational repeat instability in the mouse, we hope to find the sequences, molecules and pathways that produce trinucleotide repeat instability in humans.
