FUNCTIONS OF MILK PROTEIN GENE 5' FLANKING REGIONS ON HUMAN GROWTH-HORMONE GENE

Fragments containing 5' flanking regions of four bovine milk protein genes-alpha lactalbumin (b alpha LA), alpha S-1 casein (b alpha S1CN), beta casein (b beta CN), kappa casein (b kappa CN)-and mouse whey acidic protein (mWAP) gene were prepared by PCR and ligated to human growth hormone (hGH) gene. These recombinant DNAs were microinjected into rat embryos to produce transgenic rats, and the functions of the 5' regions to direct secretion of hGH in the milk were tested. Although milk was obtained only in 5 of 19 mWAP/hGH rat lines, more than two-thirds of the rats carrying the other four DNAs produced milk. More than 80% of the lactated rats carrying b alpha LA/, b beta CN/, and mWAP/hGH, and 33% of the lactated b alpha S1CN/hGH rats secreted detectable amounts of hGH (> 0.05 mu g/ml) in the milk. In some rats, the hGH concentrations in the milk were comparable to or more than that of the corresponding milk protein in bovine milk. The ranges of hGH concentrations in the milk of b alpha lA/, b beta CN/, b alpha S1CN/, and mWAP/hGH rats were 1.13-4,360 mu g/ml, 0.11-10,900 mu g/ml, 86.8-6,480 mu g/ml, and 6.87-151 mu g/ml, respectively. HGH was also detected in the sera of these rats, and some abnormalities of growth and reproduction were observed. All but one virgin mWAP/hGH rat secreted up to 0.0722 mu g/ml of hGH in the serum, and more than half of them showed abnormal fat accumulations at their abdomen. None of the b kappa CN/hGH rats secreted detectable amount of hGH into their milk, whereas 8 of the 11 lines secreted hGH into their sera, For the production of hGH in transgenic rat milk, the 5' region of b alpha S1CN was shown most suitable, because the b alpha S1CN/hGH rat secreted > 6,000 mu g/ml of hGH into the milk and could be reproduced. (C) 1994 Wiley-Liss, Inc.