This invention relates to the human thyroid stimulating hormone Beta chain or subunit (hTSH-xcex2), and the gene producing it. Further, the invention relates to applications of this gene.
The thyroid stimulating hormone (TSH) is a member of a family of glycoprotein hormones which includes the gonadotropins, luteinizing hormone, follicle stimulating hormone, and chorionic gonadotropin. See, e.g., Kourides et. al., Rec. Prog. Hormone Res. 40:79-120 (1984).
Each of the hormones listed supra has been found to consist of two dissimilar, noncovalently bound subunits, alpha and beta.
In an individual species, the alpha subunit for all of the hormones listed has been found to be identical, while the beta unit is different. It is the beta subunit that gives biologic and immunologic specificity to the hormones. Again, in the same species, there are areas of strong homology among the beta subunits.
Pierce, Endocrinology 89:1331 (1971), and Pierce et. al., Ann. Rev. Biochem. 50:465 (1981), show that any alpha subunit can be combined with a beta subunit to give a complete hormone. Shome, et. al., J. Clin. Endocrin. Metab. 36:618 (1983); Morgan, et. al., J. Biol. Chem. 250:5247 (1975); Birken, et. al., J. Biol. Chem. 252:5386 (1977) and Keutmann, et. al., J. Biol. Chem. 252:5393 (1977), and Biochem. Biophys. Res. Commun. 90:842 (1979), have shown that the beta subunits of chorionic gonadotropin and luteinizing hormone are most closely related, with amino acid sequence homology of 82%. Other beta subunits have lower amino acid sequence homolgies, in the range of 25-40%. Pierce, et. al., (1981) supra.
A single gene coding for the alpha subunit of human glycoprotein hormones has been isolated. Fiddes, et. al., J. Mol. Appl. Genet. 1:3 (1981); Boothby, et. al., J. Biol. Chem. 256:5121 (1981). Additionally, seven human chorionic gonadotropin beta subunit genes and one human luteinizing hormone beta subunit gene have been isolated. Talmadge, et. al., DNA 2:281 (1983); Policastro, et. al., J. Biol. Chem. 258:11492 (1983). These beta subunit genes are all highly homologous and are linked on a fragment of human chromosome 19, less than 50 kilobases long.
With respect to the beta subunit of human thyroid stimulating hormone, it has not been possible, until now, to obtain the gene expressing this subunit. This is in spite of the fact that mouse TSH-beta subunit cDNA has been synthesized and cloned and the mouse gene isolated. The gene obtained has been characterized following cross-species hybridization experiments. Gurr, et. al., Proc. Natl. Acad. Sci. 80:2122 (1983); Kourides, et. al., supra (1984). Rat and bovine TSH beta subunit cDNA have also been cloned. Croyle, et al., DNA 3:231 (1984); Maurer, et. al., J. Biol. Chem. 259:5024 (1984). Now, using mouse and bovine cDNA which have been cloned, the gene expressing human thyroid stimulating hormone beta chains has been obtained.
Usually, in obtaining a desired gene, the practice is to isolate the mRNA produced by transcription of the desired gene. Once this is obtained, cDNA can be synthesized and used as a hybridization probe to isolate the complementary gene. The methods for doing this are well known to the art. In the case of the beta subunit of human TSH, this method has proven to be unworkable. Undegraded mRNA has not been available from human pituitary glands, post mortem or post surgery.
The difficulties involved, however, have now been overcome. By relying on cDNA of different species, i.e., mouse and bovine, it has been and now is possible to obtain the gene expressing the beta subunit of human thyroid stimulating hormone.