Desmond J. Smith, Mary E. Stevens, Jan-Fang Cheng and Edward M. Rubin
Human Genome Center, Lawrence Berkeley National Laboratory, University of California, I Cyclotron Road, Berkeley, CA 94720
Moving from genomic sequence information to an understanding of biological function is a problem that is beginning to loom large as the genome project moves forward. As a pilot investigation of this problem, we have created a 2 Mb in vivo library from the Down syndrome region of human chromosome 2lq22.2 in transpolygenic mice in order to link defined DNA sequences and genes to biological function.1 Mice were created using 4 YACs (230E8, 141G6, 152F7 and 285E6, varying between 400-700 kb) and 4 P1s from human chromosome 21q22.2. The mice were screened for new genes based on the phenotypic effects on the host organism.
Our assays especially concentrate on behavioral traits such as learning and memory, since defects in these processes are the most important consequences of an extra copy of the Down syndrome region in humans. Learning and memory in the transpolygenic mice was evaluated using the Morris water maze test and the activity of the mice is being evaluated using an open field test. Mice bearing YACs 141G6 and 285E6 did not differ from non-transgenic controls. The two separate founder lines of mice created bearing YAC 152F7 display deficiencies in learning suggestive of specific hippocampal abnormalities and also show abnormalities on an open field test. Both lines of mice containing the YAC 152F7 also display a phenotype which we call "Clencher," clasping of all four limbs together when the animals are suspended by the tail. The process of microinjection in addition to producing mice which contain the intact YACs also produces mice that contain fragments of the YAC. We are now examining four different lines of mice containing different segments of YAC 152F7 and preliminary studies suggest that one of these lines shares the phenotype seen in mice containing the intact YAC while the others appear normal. We hope that molecular characterization of these different lines can be used to localize the gene on YAC 152F7 responsible for the phenotype seen in animals containing the intact YAC.
Mice harboring YAC 230E8 demonstrate very distinct abnormalities from that observed in the YAC 152F7 transgenics. The YAC 230E8 transgenics are profoundly deficient in their performance on the Morris maze, indicating functional abnormalities throughout the cerebral cortex. These studies thus demonstrate that two distinct segments of DNA from the Down syndrome region can cause distinct phenotypes of abnormal learning and memory when expressed in transgenic mice. The approaches described in this abstract can be used to investigate other regions of the human genome as a general strategy for linking gene discovery and phenotype.
[1] D.J. Smith et al., Genomics 27, 425-434 (1995).