University of Virginia Health System

The lac regulatory system relies on a line of mice constitutively and globally expressing the lac repressor protein. In order to allow for expression of the lac repressor from the mouse genome, the bacterial gene encoding lacI was reencoded with mammalian codons (synlacI). Although RNA expression was abundant in all tissues of the mouse, the original synlacI construct did not code for a functional protein. I mapped a region in the 5' end of the sequence that is necessary for appropriate splicing of the transcript, and a second region near the 3' end of the sequence that is necessary for protein expression. I constructed a modified lacI gene and established a line of transgenic mice in which expression of a functional lac repressor protein is high. The target transgene for my studies was a regulatable version of the tyrosinase gene, the defective enzyme in albinism. I inserted lac operator sequences into the tyrosinase promoter, and showed that this renders the promoter regulatable by the lac repressor. This Tyrosinase transgene rescued the coat color of an albino background strain, resulting in brown mice. When tyrosinase transgenic mice and lacI mice were crossed, the resultant double transgenic animals had an albino phenotype. When double transgenic mice were treated with the lactose analog IPTG, repression wasrelieved, and the mice were brown. Genes and Development 15:1506-1517.

Having established this "proof of principle" I then used these regulatable tyrosinase transgenic mice to investigate the developmental abnormalities in the albino visual system. Albinos have decreased numbers of ganglion cells and rod photoreceptors in their retinas, and have a misrouting of the projections of some retinal ganglion cells to the lateral geniculate nucleus. This leads to a disruption of the visual field maps in the lateral geniculate and in the visual cortex. By treating Tyrosinase, lacI double transgenic animal during discrete periods of visual system development, I could investigate when tyrosinase had to be present to allow for the correct development of these different components of the visual system. I was awarded a PhD for my work on tyrosonase in August, 2001.

C.A. Cronin. Tyrosinase (lacO)in development. PhD thesis, University of Virginia, 2001.

C.A. Cronin, W. Gluba, and H. Scrable (2001). The lac operator-repressor system is functional in the mouse. Genes & Development 15: 1506-1517.

C.A. Cronin, L. Abramova, L.A. Ligon, and H. Scrable (1999) Tight regulation of a mammalian promoter by bacterial lac operon elements. Amer. J. Human Genetics, 65 (4), A185.

C. A. Cronin and H. Scrable (1998) Differential splicing and translation of artificial genes caused by sequences linking the promoter to the coding region. Amer. J. Human Genetics, 63 (4), A37.

Rudolf, G.D., C.A. Cronin, G.B. Landwehrmeyer, D.G. Standaert, J.B. Penney, Jr., and A.B. Young (1996) Expression of N-methyl-D-aspartate glutamate receptor subunits in the prefrontal cortex of the rat. Neuroscience 73: 4