Regulation of the human neuropeptide Y gene in transgenic mice

Geoffrey Carl Waldbieser, Purdue University

Abstract

Neuropeptide Y (NPY) is a 36 residue peptide widespread in the mammalian nervous system, and is involved in the regulation of blood pressure, hormone release, and control of appetite. Current research at Purdue University involves the identification of factors which regulate transcriptional activity of the NPY gene. In order to identify regions of the NPY gene which confer tissue-specificity, fusion genes, consisting of human NPY 5$\sp\prime$ flanking DNA inserted upstream from the bacterial chloramphenicol acetyltransferase (CAT) gene, were microinjected into one-called mouse embryos to make transgenic mice. These gene constructs contained approximately 6500, 3800, 1080, 800, or 152 bp of NPY 5$\sp\prime$ flanking sequence. In order to establish the techniques necessary for the production of transgenic mice, gene constructs containing cholecystokinin and/or growth hormone sequences were injected into one-celled embryos. Transgenic mice were produced for only four of eight constructs, and the transgene integrated in 8-15% of the mice which developed from microinjected embryos. Six of seven founder animals used for breeding transmitted the transgene to offspring. Of the five NPY/CAT constructs injected into one pronucleus of one-celled mouse embryos, only two integrated into the mouse genome. Five transgenic mice were produced for the NPY 803/CAT construct, and one was produced for the NPY 152/CAT construct. Each founder mouse transmitted the transgene to offspring, and one NPY 803/CAT line demonstrated X-chromosome-linked inheritance of the transgene. Tissue extracts were prepared from NPY/CAT mice in order to determine levels of transgene activity. Four of the five lines of NPY 803/CAT mice were tested. Significant levels of CAT activity were detected in the brain, spinal cord, eyes, and adrenal gland, which contain moderate to high levels of endogenous neuropeptide Y. Tissue-specific patterns of expression varied between lines of transgenic mice. Mice from the NPY 152/CAT transgenic line expressed significant levels of the transgene product in the brain, spinal cord, and adrenal gland. However, significant levels of transgene activity were also detected in the lung and liver. These experiments demonstrated tissue-specific elements which exist between $-$803 and $-$152 bp of the neuropeptide Y gene in transgenic mice.

Degree

Ph.D.

Advisors

Chrisman, Purdue University.

Subject Area

Genetics|Molecular biology

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