alpha 2u-Globulin is a rat protein of as yet unknown function whose synthesis can be induced by glucocorticoids and several other hormones. Induction by glucocorticoids is a secondary response to the hormone: protein synthesis is required before the hormone can exert its stimulatory effect on alpha 2u-globulin transcription. We have used the linker-scanning mutagenesis procedure, followed by transfer of the mutant genes into mouse L-cells for analysis of their phenotype, to determine sequences within a cloned alpha 2u-globulin promoter that are required for its regulation by glucocorticoids. Mutations between positions -115 and -160 abolish or greatly reduce the inducibility of alpha 2u-globulin by the hormone. Mutations just upstream from this region, between positions -177 and -220, have an opposite effect; they increase induction two- to fourfold.

Expression of the rat 2u -globulin gene family is regulated in the adult male liver by a number of hormones, including growth hormone, thyroid hormone and several steroids. Upon injection into ovariectomized females, estrogens first induce 2u -globulin expression and then suppress this gene after several days of hormone administration. To study this phenomenon, we developed a mouse L-cell line that expressed the human estrogen receptor. High levels of rat 2u -globulin transcript were induced in stable transfectants of this line carrying a cloned 2u -globulin gene, following exposure to 17 -estradiol. Since this induction was inhibited by cycloheximide, the response to estrogen, as to other steroids, appears to be secondary.Using genes with variously deleted 5 -upstream regions, sequences responsible for this induction were located between 730 bp and 223 bp relative to the start of transcription. Examination of the DNA in this region revealed that an estrogen receptor element was located at 590 bp in an area that is highly conserved in most known 2u -globulin genes. Administration of both dexamethasone and estrogen produced a synergistic effect in this system. The induction of 2u -globulin RNA by estrogen in L-cells may re-capitulate the initial response to estrogen in vivo, and therefore represents a good model system to seek the identity of the other factors required to effect full induction.

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