Jeung S. Yun

Metabolic control of gene expression: in vivo studies with transgenic mice

Transgenic animals provide a comprehensive model for investigating genes encoding inducible enzymes involved in metabolism, since the molecular mechanisms regulating gene transcription can be studied in the whole animal. Studies on the promoters of the genes encoding two key enzymes in the gluconeogenic and glycolytic pathways--phosphoenol-pyruvate carboxykinase and pyruvate kinase are described as examples of this approach. Work on the phosphoenolpyruvate carboxykinase promoter using transgenic mice has been particularly informative: the cis-acting elements involved in hormonal regulation, tissue specificity and developmental inhibition of gene expression have been identified and their function in vivo examined.

The Human Growth Hormone Transgene: Expression in Hemizygous and Homozygous Mice

Abstract Female transgenic mice carrying the mouse metallothionein-I/human growth hormone (hGH) fusion gene are sterile. Transmission of the transgene has been limited to the male germ line, resulting in the production of hemizygous (He) progeny containing only a single (paternal) copy of the gene. Using ovary transfer, we have developed procedures for producing homozygous (Ho) TG mice, viz., male TG mice were mated with control (non-TG) females carrying ovaries donated by female TG mice. In both He and Ho TG animals, serum levels of hGH were higher (1.5–fold) in males than in females, tended to decrease with age of the animal, and were increased (about 5-fold) by zinc induction. However, in comparison to He animals of the same sex, the Ho TG mice attained a greater body weight and had more than 2-fold higher levels of liver hGH-mRNA and serum hGH, both under basal conditions and in response to zinc induction. That is, the expression of the transgene was qualitatively similar in He and Ho TG mice, but the level of transgene activity was greater in the Ho animals. We interpret this to indicate that both copies (maternal and paternal) of the transgene were active and expressed additively (or cooperatively) in the Ho TG animal.

Intermediate filaments of bovine pulmonary artery smooth muscle: Distribution, isolation and polypeptide composition

1. The distribution of intermediate filaments in bovine pulmonary artery smooth muscle was studied. 2. These filaments contained at least two polypeptides of 43,000 and 55,000 daltons. 3. The 55,000 dalton polypeptide was purified by ion-exchange chromatography. 4. The amino acid compositions of the polypeptides were determined. 5. The 55,000 dalton polypeptide and a mixture of the 43,000 and 55,000 dalton polypeptide reassembled into structures similar to native intermediate filaments.

Stress-induced secretion of human growth hormone in transgenic mice.

Abstract The effect of stress on human growth hormone (hGH) secretion was studied in transgenic mice. Experiments were conducted on fourth, fifth, and sixth generation male mice carrying a fusion gene, consisting of the promoter sequence of the mouse metallothionein I gene ligated to the hGH structural gene (mMT-I/hGH). In animals adapted to a controlled photoperiod, basal (unstimulated) levels of plasma hGH exhibited a diurnal cycling, with peak values occurring during the later half of the light period (15.5 ± 1.0 vs 10.7 ± 0.9 ng/ml, mean ± SE, light versus dark, respectively). Food deprivation (5 days) led to elevated levels of plasma hGH (11.0 ± 0.7 vs 32.0 ± 4.2 ng/ml, preversus post-fast, respectively) accompanied by weight loss (49.5 ± 0.8 vs 34.3 ± 0.7 g), and hypoglycemia (7.8 ± 0.2 vs 5.0 ± 0.3 mM); glucose administration (5% drinking solution ad libitum) blocked the changes in levels of plasma hGH (12.2 ± 1.1 vs 13.8 ± 0.8 ng/ml) and plasma glucose (7.4 ± 0.3 vs 7.9 ± 0.5 mM), although the animals still sustained significant weight loss (44.9 ± 1.6 vs 35.2 ± 1.1 g). Vigorous exercise (swimming, 4 hr) produced a small but significant increase in plasma hGH, 12.1 ± 1.1 ng/ml (1 hr pre-swim) vs 16.7 ± 0.6 ng/ml (immediately post-swim). These findings indicate that the mMT-I/hGH transgene is responsive to the physiologic status of the host animal. Taken together with information regarding the heterologous components of the fusion gene, these data are consistent with the view that the hGH (structural) sequence may play a role in the response to stress.

HUMAN GROWTH HORMONE VARIANT (hGH-B) PROMOTES GROWTH IN TRANSGENIC MICE

The human growth hormone (hGH) gene family consists of 5 members linked on the long arm of chromosome 17. The hGH-A gene is expressed by the somatotrophs of the anterior pituitary. The hGH-B gene encodes a GH-like polypeptide of unknown function. To determine if hGH-B is capable of stimulating growth in animals, transgenic mice containing a metallothionine-I/hGH-B fusion gene were generated. The GH produced by these mice was compared with pituitary hGH-A, mouse GH, and hGH-A and hGH-B from transfected mouse L cell fibroblasts. The 3 transgenic male mice were larger at weaning (26-31 gms) than control males (20 ± 0.4 gms); by age 2 mos the transgenic mice were 40-90% larger than the controls. Serum from these mice contained large (17-30 μg/ml) amounts of hGH-immunoreactivity while control serum did not. The GH-immunoreactivity from the transgenic mice differed from hGH-A in a) cross-reactivity with a panel of monoclonal antibodies, b) behavior on gel filtration and c) pI.Conclusions: hGH-B is capable of stimulating growth in mice. The differences in amino acid sequence between hGH-A and hGH-B result in substantial differences in tertiary and quanternary structure.