Transgenic Rescue of the PWS-IC Deletion Mouse

 Stormy Jo Chamberlain, James Resnick* and Camilynn I. Brannan.  Department of Molecular Genetics and Microbiology, University of Florida, Box 100266 JHMHC, Gainesville FL  32610  USA. 

             Prader-Willi syndrome (PWS) is a contiguous gene syndrome that results from the absence of two or more paternally expressed genes.  PWS patients show neonatal failure to thrive, hypotonia, and poor suckle, but later develop hyperphagia, obesity, moderate mental retardation, and small stature.  We have taken advantage of a mouse model for PWS in which paternal transmission of a PWS Imprinting Center (PWS-IC) deletion results in complete loss of local paternal gene expression.  This targeted mutation accurately models the both the molecular and phenotypic aspects of PWS.  Specifically, mice which inherit the PWS-IC deletion show silencing of all of the paternally expressed genes in the region, including Mkrn3, Magel2, Ndn, Snrpn, and at least 5 snoRNA species.  On the (C57BL/6 X 129/Sv) F1 strain background these mice exhibit failure to thrive, poor suckle, and ultimately die by 7 days of age.

We sought to correlate specific attributes of the PWS phenotype to specific gene products.  Our strategy was to make transgenic mice harboring each of the paternally expressed genes, mate them to PWS-IC deletion carrier males, and ascertain transgenic rescue in the pups that inherit both the transgene and the PWS-IC deletion allele.  We screened a BAC library and identified three clones that were used for production of transgenic mice.  One clone carried the Mkrn3, Magel2, and Ndn genes, another carried Snrpn and 2 single-copy snoRNA species, and the final clone carried Snrpn, 4 single-copy snoRNA species, and one multi-copy snoRNA species.  Multiple transgenic lines were obtained with each clone and the lines were analyzed for transgene copy number and integrity.  Single and multiple copy lines were found and several of these were intact.  Furthermore, the lines were also tested for expression of the paternally expressed genes in the region. 

When transgenic mice were mated to PWS-IC deletion mice, we found that the PWS-IC deletion mice on the (FVB X C57BL/6) F1 strain background could survive until adulthood if the wild-type littermates were removed.  The surviving mice were smaller than wild-type littermates, but never got obese. Secondly, we found that one transgenic line exhibiting expression of a single snoRNA cluster rescues the small stature phenotype of PWS mice, while three other genes were found to have no obvious affect on the PWS phenotype in mice.

In total, we have found that the PWS phenotype is less severe on certain strain backgrounds, suggesting the presence of modifier genes that may ameliorate the PWS phenotype.  We have also determined that Mkrn3, Magel2, Ndn, Snrpn, and 5 species of snoRNAs are not solely responsible for neonatal failure to thrive.  Finally, we demonstrated that the small stature phenotype of PWS-IC deletion mice could be rescued by a single transgenic line that is the only one showing expression of one particular snoRNA species, suggesting that this tandemly repeated snoRNA cluster is involved in the regulation of growth of PWS-IC deletion mice, and most likely PWS individuals. 

July 2003