BAG-1M: a potential specificity determinant of corticosteroid receptor action.

BAG-1M is a eukaryotic cochaperone that associates with several proteins, including the glucocorticoid receptor (GR). It down-regulates GR-mediated transactivation by a mechanism that requires its prior recruitment by the liganded receptor from cytoplasm into the nucleus. In the nucleus, it uses a repeated sequence motif ([EEX4]8) at its NH2 terminus to inhibit DNA binding, as well as transactivation functions of the receptor. The mineralocorticoid receptor (MR), a structural and functional homologue of the GR, is unable to translocate BAG-1M into the nucleus, and its transactivation function is also not affected by this protein. This differential regulation of GR and MR activity could be relevant in classic mineralocorticoid tissues such as the kidney in which GR activity needs to be repressed to allow the MR to exert its action. In in situ hybridization studies, we show that BAG-1M is expressed in the kidney. Its expression pattern, especially in the developing kidney, correlated well with that of the GR. We therefore postulate that BAG-1M may be a specificity determinant in GR and MR action, and may feature prominently in the control of GR activity in kidney development.

Isoform-specific expression of BAG-1 in mouse development.

BAG-1 is a family of proteins with diverse activities that range in cultured cells from protection against programmed cell death through to regulation of steroid hormone action. At least three proteins (BAG-1L, BAG-1M and BAG-1) are encoded by the Bag-1 mRNA through the use of alternative translation-initiation sites. To assess the in vivo function of these factors, we have used in situ hybridization and immunohistochemical techniques to determine the distribution of Bag-1 transcript and proteins during mouse development. Bag-1 mRNA was identified in several organs with cartilaginous tissues showing the highest expression levels. The level of expression at some of these sites was downregulated during the course of development. In the immunohistochemical studies, antibodies directed against the BAG-1 proteins stained all the sites identified in the in situ hybridization studies although isoform-specific differences were observed. BAG-1L specific antibody showed ubiquitous staining as early as day 10.5 post-coitum but there was a progressive restriction during subsequent stages of embryogenesis. On the contrary, an antibody that preferentially recognized the other isoforms only stained the mouse myocardium in the early developmental stages before finally recognizing additional organs later on in development. These results demonstrate a stage- and site-specific expression of the BAG-1 isoforms during mouse development.

The Wilms tumor suppressor gene wt1 is required for development of the spleen.

The Wilms tumor suppressor gene WT1 (wt1 in mouse) is unique among tumor suppressors because, in addition to its involvement in cancer [1] [2] and various other diseases [3] [4] [5] [6], it has an essential role in the development of certain organs. This is revealed by the phenotype of mice with inactivated wt1 alleles [7]. These animals exhibit a complete failure of kidney and gonad development as well as abnormalities of the heart and mesothelial structures. On a C57BL/6 genetic background, wt1(-/-) animals die between day 13.5 (E13.5) and 15.5 (E15.5) of embryonic development [7]. We report here that crossing of the wt1 mutation onto different mouse backgrounds delayed embryonic lethality until birth. In wt1(-/-) mice on these different genetic backgrounds, we observed a dramatic failure of spleen development, in addition to the well characterized phenotypic abnormalities. The spleen anlage formed at around E12 to E13 and involuted by the E15 stage, before the invasion of hematopoietic cells. The absence of proper spleen development in these wt1(-/-) embryos correlated with enhanced apoptosis in the primordial spleen cells. The expression of hox11, a gene that also controls development of the spleen [8] [9], was not altered by the inactivation of wt1. In situ hybridization revealed that the two genes are regulated independently. These findings demonstrate that the penetrance of the wt1(-/-) phenotype depends on the existence of one or more modifier gene(s) and that wt1 plays a pivotal role in the development of the spleen, thereby extending its role in organogenesis.

Expression of androgen receptor mRNA during mouse embryogenesis.

Androgen receptor (AR) is a member of the nuclear receptor superfamily which acts as a ligand-dependent transcription factor (Beato, M., Herrlich, P., Schütz, 1989. Steroid hormone receptors: many actors in search of a plot. Cell 83, 851-857). It plays a pivotal role in sexual development and reproduction (Wilson, J.D., Griffin, J.E., George, F.W., Leshin, M., 1981. The role of gonadal steroids in sexual differentiation. Rec. Prog. Horm. Res. 37, 1-39; Jost, A., 1990. Hormonal control of the masculinization of the body. In: Baulieu, E.E., Kelly, D.A., (Eds.), Hormones, from Molecules to Disease. Chapman and Hall, New York and London, pp. 439-442.). Mutations in the AR sequence cause a number of physiological disorders, such as partial and complete androgen insensitivity syndromes, that lead to abnormal sexual development (Patterson, M.N., McPhaul, M.J., Hughes, I.A., 1994. Androgen insensitivity syndrome. Ballière's Clin. Endocrinol. Metab. 8, 379-404.). There are indications that AR may also have other functions. For example, structural alterations of the AR sequence have been implicated in prostate cancer (Visakorpi, T., Huytinen, E., Koivisto, P., Tanner, M., Keinänen, R., Palmberg, C., Palotie, A., Tammela, T., Isola, J., Kallioniemi, O.-P., 1995. In vivo amplification of the androgen receptor gene and progression of human prostate cancer. Nature Genet. 9, 401-406.) and in the development of spinal and bulbar muscular atrophy, a neurodegenerative disease (Kennedy, W.R., Alter, M., Sung, J.H., 1968. Progressive proximal spinal and bulbar muscular atrophy of late onset: a sex-linked recessive trait. Neurology 18, 671-680.). Here, we have investigated the spatial and temporal expression of AR during mouse organogenesis by in situ hybridisation. We demonstrate that AR transcripts occur in the developing external genitalia, pituitary, adrenals, kidneys and musculus levator ani, in addition to the known expression sites in the Wolffian ducts and its derivatives and during development of the mammary glands.