Search

Purpose: Androgen receptor abundance and androgen receptor–regulated gene expression in castration-recurrent prostate cancer are indicative of androgen receptor activation in the absence of testicular androgen. Androgen receptor transactivation of target genes in castration-recurrent prostate cancer occurs in part through mitogen signaling that amplifies the actions of androgen receptor and its coregulators. Herein we report on the role of 14-3-3η in androgen receptor action.Experimental Design and Results: Androgen receptor and 14-3-3η colocalized in COS cell nuclei with and without androgen, and 14-3-3η promoted androgen receptor nuclear localization in the absence of androgen. 14-3-3η interacted with androgen receptor in cell-free binding and coimmunoprecipitation assays. In the recurrent human prostate cancer cell line, CWR-R1, native endogenous androgen receptor transcriptional activation was stimulated by 14-3-3η at low dihydrotestosterone concentrations and was increased by epidermal growth factor. Moreover, the dihydrotestosterone- and epidermal growth factor–dependent increase in androgen receptor transactivation was inhibited by a dominant negative 14-3-3η. In the CWR22 prostate cancer xenograft model, 14-3-3η expression was increased by androgen, suggesting a feed-forward mechanism that potentiates both 14-3-3η and androgen receptor actions. 14-3-3η mRNA and protein decreased following castration of tumor-bearing mice and increased in tumors of castrate mice after treatment with testosterone. CWR22 tumors that recurred 5 months after castration contained 14-3-3η levels similar to the androgen-stimulated tumors removed before castration. In a human prostate tissue microarray of clinical specimens, 14-3-3η localized with androgen receptor in nuclei, and the similar amounts expressed in castration-recurrent prostate cancer, androgen-stimulated prostate cancer, and benign prostatic hyperplasia were consistent with androgen receptor activation in recurrent prostate cancer.Conclusion: 14-3-3η enhances androgen- and mitogen-induced androgen receptor transcriptional activity in castration-recurrent prostate cancer. (Clin Cancer Res 2009;15(24):7571–81)

A graph is denoted by G with the vertex set V(G) and the edge set E(G). A path P = 〈v0, v1, … , vm〉 is a sequence of adjacent vertices. Two paths with equal length P1 = 〈 u1, u2, … , um〉 and P2 = 〈 v1, v2, … , vm〉 from a to b are independent if u1 = v1 = a, um = vm = b, and ui ≠ vi for 2 ⩽ i ⩽ m − 1. Paths with equal length { P i } i = 1 n from a to b are mutually independent if they are pairwisely independent. Let u and v be two distinct vertices of a bipartite graph G, and let l be a positive integer length, dG(u, v) ⩽ l ⩽ ∣V(G) − 1∣ with (l − dG(u, v)) being even. We say that the pair of vertices u, v is (m, l)-mutually independent bipanconnected if there exist m mutually independent paths P i l i = 1 m with length l from u to v. In this paper, we explore yet another strong property of the hypercubes. We prove that every pair of vertices u and v in the n-dimensional hypercube, with d Q n ( u , v ) ⩾ n - 1 , is (n − 1, l)-mutually independent bipanconnected for every l , d Q n ( u , v ) ⩽ l ⩽ | V ( Q n ) - 1 | with ( l - d Q n ( u , v ) ) being even. As for d Q n ( u , v ) ⩽ n - 2 , it is also (n − 1, l)-mutually independent bipanconnected if l ⩾ d Q n ( u , v ) + 2 , and is only (l, l)-mutually independent bipanconnected if l = d Q n ( u , v ) .

Purpose: Androgen receptor abundance and androgen receptor–regulated gene expression in castration-recurrent prostate cancer are indicative of androgen receptor activation in the absence of testicular androgen. Androgen receptor transactivation of target genes in castration-recurrent prostate cancer occurs in part through mitogen signaling that amplifies the actions of androgen receptor and its coregulators. Herein we report on the role of 14-3-3η in androgen receptor action. Experimental Design and Results: Androgen receptor and 14-3-3η colocalized in COS cell nuclei with and without androgen, and 14-3-3η promoted androgen receptor nuclear localization in the absence of androgen. 14-3-3η interacted with androgen receptor in cell-free binding and coimmunoprecipitation assays. In the recurrent human prostate cancer cell line, CWR-R1, native endogenous androgen receptor transcriptional activation was stimulated by 14-3-3η at low dihydrotestosterone concentrations and was increased by epidermal growth factor. Moreover, the dihydrotestosterone- and epidermal growth factor–dependent increase in androgen receptor transactivation was inhibited by a dominant negative 14-3-3η. In the CWR22 prostate cancer xenograft model, 14-3-3η expression was increased by androgen, suggesting a feed-forward mechanism that potentiates both 14-3-3η and androgen receptor actions. 14-3-3η mRNA and protein decreased following castration of tumor-bearing mice and increased in tumors of castrate mice after treatment with testosterone. CWR22 tumors that recurred 5 months after castration contained 14-3-3η levels similar to the androgen-stimulated tumors removed before castration. In a human prostate tissue microarray of clinical specimens, 14-3-3η localized with androgen receptor in nuclei, and the similar amounts expressed in castration-recurrent prostate cancer, androgen-stimulated prostate cancer, and benign prostatic hyperplasia were consistent with androgen receptor activation in recurrent prostate cancer. Conclusion: 14-3-3η enhances androgen- and mitogen-induced androgen receptor transcriptional activity in castration-recurrent prostate cancer. (Clin Cancer Res 2009;15(24):7571–81)

Ag raphG is hamiltonian connected if there exists a hamiltonian path joining any two distinct nodes of G. Two hamiltonian paths P1 = u1, u2 ,..., uν(G) and P2 = v1, v2 ,..., vν(G) of G from u to v are independent if u = u1 = v1, v = uν(G) = vν(G) ,a nduivi for every 1 < i < ν(G). A set of hamiltonian paths, {P1, P2 ,..., Pk} ,o fG from u to v are mutually independent if any two different hamiltonian paths are independent from u to v .A graph isk mutually independent hamiltonian connected if for any two distinct nodes u and v ,t here arek mutually independent hamiltonian paths from u to v. The mutually independent hamiltonian connectivity of a graph G, IHP(G), is the maximum integer k such that G is k mutually inde- pendent hamiltonian connected. Let n and k be any two distinct positive integers with n−k ≥ 2. We use Sn, k to denote the (n, k)-star graph. In this paper, we prove that IHP(Sn, k )= n − 2e x- cept for S4, 2 such that IHP(S4, 2 )= 1.

It is important for a network to tolerate as many faults as possible. With the graph representation of an interconnection network, a k -regular hamiltonian and hamiltonian connected network is super fault-tolerant hamiltonian if it remains hamiltonian after removing up to k − 2 vertices and/or edges and remains hamiltonian connected after removing up to k − 3 vertices and/or edges. Super fault-tolerant hamiltonian networks have an optimal flavor with regard to the fault-tolerant hamiltonicity and fault-tolerant hamiltonian connectivity. For this reason, a cycle composition framework was proposed to construct a ( k + 2)-regular super fault-tolerant hamiltonian network based on a collection of n k -regular super fault-tolerant hamiltonian networks containing the same number of vertices for n ⩾ 3 and k ⩾ 5. This paper is aimed to emphasize that the cycle composition framework can be still applied even when k = 4.

Partial androgen insensitivity with sex phenotype variation in two unrelated families was associated with missense mutations in the androgen receptor (AR) gene that disrupted the AR NH2-terminal/carboxy terminal interaction. Each mutation caused a single amino acid change within the region of the ligand-binding domain that forms activation function 2 (AF2). In one family, the mutation I737T was in alpha helix 4 and in the other F725L was between helices 3 and 4. Neither mutation altered androgen binding as determined by assays of mutant AR in the patient's cultured genital skin fibroblasts or of recombinant mutant receptors transfected into COS cells. In transient cotransfection assays in CV1 cells, transactivation with the AR mutants at low concentrations of DHT was reduced several fold compared with wild-type AR but increased at higher concentrations. Defects in NH2-terminal/carboxy terminal interactions were identified in mammalian two hybrid assays. In similar assays, there was reduced binding of the p160 coactivators TIF2/SRC2 and SRC1 to the mutant AR ligand binding domains (LBD). In the family with AR I737T, sex phenotype varied from severely defective masculinization in the proband to a maternal great uncle whose only manifestation of AIS was severe gynecomastia. He was fertile and passed the mutation to two daughters. The proband of the F725L family was also incompletely masculinized but was raised as a male while his half-sibling by a different father was affected more severely and reared as a female. These studies indicate that the function of an AR AF2 mutant in male development can vary greatly depending on the genetic background.

The transcription factor coregulator Casein kinase IIβ-binding protein 2 or CR6-interacting factor 1 (CKβBP2/CRIF1) binds the androgen receptor (AR) in prostate cancer cells and in response to dihydrotestosterone localizes with AR on the prostate-specific antigen gene enhancer, but does not bind DNA suggesting CKβBP2/CRIF1 localization in chromatin is determined by AR. In this study we show also that CKβBP2/CRIF1 inhibits wild-type AR and AR N-terminal transcriptional activity, binds to the AR C-terminal region, inhibits interaction of the AR N- and C-terminal domains (N/C interaction) and competes with p160 coactivator binding to the AR C-terminal domain, suggesting CKβBP2/CRIF1 interferes with AR activation functions 1 and 2. CKβBP2/CRIF1 is expressed mainly in stromal cells of benign prostatic hyperplasia and in stroma and epithelium of prostate cancer. CKβBP2/CRIF1 protein is increased in epithelium of androgen-dependent prostate cancer compared to benign prostatic hyperplasia and decreased slightly in castration recurrent epithelium compared to androgen-dependent prostate cancer. The multifunctional CKβBP2/CRIF1 is a STAT3 interacting protein and reported to be a coactivator of STAT3. CKβBP2/CRIF1 is expressed with STAT3 in prostate cancer where STAT3 may help to offset the AR repressor effect of CKβBP2/CRIF1 and allow AR regulation of prostate cancer growth.

An androgen receptor (AR) interacting protein was isolated from a HeLa cell cDNA library by two-hybrid screening in yeast using the AR DNA+ligand binding domains as bait. The protein has sequence identity with human protein inhibitor of activated signal transducer and activator of transcription (PIAS1) and human Gu RNA helicase II binding protein (GBP). Binding of PIAS1 to human AR DNA+ligand binding domains was androgen dependent in the yeast liquid beta-galactosidase assay. Activation of binding by dihydrotestosterone was greater than testosteroneestradiolprogesterone. PIAS1 binding to full-length human AR in a reversed yeast two hybrid system was also androgen dependent. [35S] PIAS1 bound a glutathione S-transferase-AR-DNA binding domain (amino acids 544-634) fusion protein in affinity matrix assays. In transient cotransfection assays using CV1 cells with full-length human AR and a mouse mammary tumor virus luciferase reporter vector, there was an androgen-dependent 3- to 5-fold greater increase in luciferase activity with PIAS1 over that obtained with an equal amount of control antisense cDNA or mutant PIAS1. Constitutive transcriptional activity of the AR N-terminal+DNA binding domain was increased 6-fold by PIAS1. PIAS1 also enhanced glucocorticoid receptor transactivation in response to dexamethasone but inhibited progesterone-induced progesterone receptor transactivation in the same assay system. mRNA for PIAS1 was highly expressed in testis of human, monkey, rat, and mouse. In rat testis the onset of PIAS1 mRNA expression coincided with the initiation of spermatogenesis between 25-30 days of age. Immunostaining of human and mouse testis with PIAS1-specific antiserum demonstrated coexpression of PIAS1 with AR in Sertoli cells and Leydig cells. In addition, PIAS1 was expressed in spermatogenic cells. The results suggest that PIAS1 functions in testis as a nuclear receptor transcriptional coregulator and may have a role in AR initiation and maintenance of spermatogenesis.

An androgen receptor (AR) gene mutation identified in the androgen-dependent human prostate cancer xenograft, CWR22, changed codon 874 in the ligand-binding domain (exon H) from CAT for histidine to TAT for tyrosine and abolished a restriction site for the endonuclease SfaNI. SfaNI digestion of AR exon H DNA from normal but not from prostate cancer tissue indicated H874Y is a somatic mutation that occurred before the initial tumor transplant. CWR22, an epithelial cell tumor, expresses a 9.6-kb AR mRNA similar in size to the AR mRNA in human benign prostatic hyperplasia. AR protein is present in cell nuclei by immunostaining as in other androgen-responsive tissues. Transcriptional activity of recombinant H874Y transiently expressed in CV1 cells in the presence of testosterone or dihydrotestosterone was similar to that of wild type AR. With dihydrotestosterone at a near physiological concentration (0.01 nM), H874Y and wild type AR induced 2-fold greater luciferase activity than did the LNCaP mutant AR T877A. The adrenal androgen, dehydroepiandrosterone (10 and 100 nM) with H874Y stimulated a 3- to 8-fold greater response than with wild type AR and at 100 nM the response was similar with the LNCaP mutant. H874Y, like the LNCaP cell mutant, was more responsive to estradiol and progesterone than was wild type AR. The antiandrogen hydroxyflutamide (10 nM) had greater agonist activity (4- to 7-fold) with both mutant ARs than with wild type AR. AR mutations that alter ligand specificity may influence tumor progression subsequent to androgen withdrawal by making the AR more responsive to adrenal androgens or antiandrogens.

New techniques for handling multicriteria fuzzy decision-making problems based on vague set theory are presented. The proposed techniques allow the degrees of satisfiability and non-satisfiability of each alternative with respect to a set of criteria to be presented by vague values. Furthermore, the proposed techniques allow the decision-maker to assign a different degree of importance to each criteria. The techniques proposed in this paper can provide a useful way to efficiently help the decision-maker to make his decisions.

The androgen-regulated 20-kDa protein gene consists of four exons that code for a major secretory protein of rat ventral prostate. Analysis of its potential cis-acting transcriptional regulatory elements revealed that a large intron 1 region (In-1) had stronger androgen response element (ARE) activity than did the 5'-flanking DNA. In cotransfected CV1 cells, In-1 and its most active subfragment In-1c functioned as AREs but not glucocorticoid response elements (GRE). Nevertheless several ARE/GRE-like partial palindromic sequences are present in In-1c, and it bound both androgen receptors and glucocorticoid receptors in mobility shift assays. A cluster of three ARE/GRE-like sequences contained within a 39-base pair sequence of In-1c had both ARE and GRE activities when analyzed as an isolated oligonucleotide, suggesting that other elements within In-1c determined its ARE specificity. In addition to ARE/GRE-like sequences, In-1c contains putative response elements for the transcription factors AP1, CREB, AP2, OCT-1, C/EBP, and a number of inverted and direct repeats. The ARE specificity of In-1c observed in CV1 cells was diminished in PC3 and HeLa cells transiently cotransfected with an androgen receptor or glucocorticoid receptor expression vector together with an In-1c reporter vector; however, the ARE activity of In-1c was greater than its GRE activity in these cell lines. Interestingly, a 131-base pair subfragment of In-1c retained ARE specificity in all three cell lines.

Intron and 5'-flanking regions of the androgen-regulated C3 subunit gene contain potential cis-acting transcription control sequences including several 15-base pair (bp) partial palindromes resembling response elements for glucocorticoid (GRE) and progesterone (PRE) receptors. Specific DNA binding of the androgen receptor (AR) and androgen-dependent activation of transcription indicate that some of these GRE/PRE-like sequences are capable of functioning as androgen response elements (ARE). A 0.3-kilobase pair (kbp) 5'-flanking fragment including the promoter region contains one such sequence (element A) and a 0.5-kbp region of the first intron contains two sequences (elements B and C). Androgen-dependent enhancement of transcription was assayed by cotransfection of CV1 cells with a rat AR expression vector, pCMVrAR, and C3 genomic fragments or synthetic elements cloned into the reporter vector ptkCAT. Enhancement of chloramphenicol acetyltransferase activity with the 0.5-kbp first intron fragment was 16 +/- 4-fold, while with the 0.3-kbp 5'-flanking fragment no response was detected and element C alone was greater than B or A. Binding of AR in the mobility shift assay correlated with androgen-dependent enhancement of chloramphenicol acetyltransferase activity. The intensity of transcriptional enhancement with the 0.5-kbp intron fragment suggested that other regulatory sequences within this intron region potentiated the ARE activities of elements B and C. ARE activity of the strongest C3 gene response element (C) was similar to that of a potent GRE (element M) of the mouse mammary tumor virus gene.

Human sperm-associated antigen 11 (SPAG11) is closely related to beta-defensins in structure, expression, and function. Like the beta-defensins, SPAG11 proteins are predominantly expressed in the male reproductive tract, where their best-known major roles are in innate host defense and reproduction. Although several hypotheses have emerged to describe the evolution of beta-defensin and SPAG11 multifunctionality, few describe these multiple functions in terms of defensin interactions with specific proteins. To gain insight into the protein interaction potentials of SPAG11 and the signaling pathways that SPAG11 may influence, we used a yeast two-hybrid screening of a human testis-epididymis library. The results reveal human SPAG11B isoform D (SPAG11B/D) interactions with tryptase alpha/beta 1 (TPSAB1), tetraspanin 7 (TSPAN7), and attractin (ATRN). These interactions were confirmed by coimmunoprecipitation and glutathione S-transferase affinity matrix binding. SPAG11B/D and the three interacting proteins are expressed in the proximal epididymis, and all function in immunity and fertility pathways. We analyzed the functional consequences of SPAG11B/D interaction with TPSAB1 and showed that SPAG11B/D is both a substrate and a potent inhibitor of TPSAB1 activity. Furthermore, we show that (like SPAG11B/D) TSPAN7 and ATRN are associated with spermatozoa.

Human sperm-associated antigen 11 (SPAG11) is closely related to beta-defensins in structure, expression, and function. Like the beta-defensins, SPAG11 proteins are predominantly expressed in the male reproductive tract, where their best-known major roles are in innate host defense and reproduction. Although several hypotheses have emerged to describe the evolution of beta-defensin and SPAG11 multifunctionality, few describe these multiple functions in terms of defensin interactions with specific proteins. To gain insight into the protein interaction potentials of SPAG11 and the signaling pathways that SPAG11 may influence, we used a yeast two-hybrid screening of a human testis-epididymis library. The results reveal human SPAG11B isoform D (SPAG11B/D) interactions with tryptase alpha/beta 1 (TPSAB1), tetraspanin 7 (TSPAN7), and attractin (ATRN). These interactions were confirmed by coimmunoprecipitation and glutathione S-transferase affinity matrix binding. SPAG11B/D and the three interacting proteins are expressed in the proximal epididymis, and all function in immunity and fertility pathways. We analyzed the functional consequences of SPAG11B/D interaction with TPSAB1 and showed that SPAG11B/D is both a substrate and a potent inhibitor of TPSAB1 activity. Furthermore, we show that (like SPAG11B/D) TSPAN7 and ATRN are associated with spermatozoa.

Protein inhibitor of activated STAT1 (PIAS1) functions as a nuclear receptor coregulator and is expressed in several cell types of human testis. However, the mechanism of PIAS1 coregulation is unknown. We report here that PIAS1 has characteristics of a scaffold attachment protein. PIAS1 localized in nuclei in a speckled pattern and bound A-T-rich double-stranded DNA, a function of scaffold attachment proteins in chromatin regions of active transcription. DNA binding was dependent on a 35-amino acid sequence conserved among members of the PIAS family and in scaffold attachment proteins. The PIAS family also bound the androgen receptor DNA binding domain, and binding required the second zinc finger of this domain. PIAS1 contained an intrinsic activation domain but had bi-directional effects on androgen receptor transactivation; lower expression levels inhibited and higher levels increased transactivation in CV1 cells. Other PIAS family members also had dose-dependent effects on transactivation, but they were in a direction opposite to those of PIAS1. When coexpressed with PIAS1, other PIAS family members counteracted PIAS1 coregulation of androgen receptor transactivation. The interaction of PIAS1 with other members of the PIAS family suggests a transcription coregulatory mechanism involving a multicomponent PIAS nuclear scaffold.

Androgen signaling influences the development and growth of prostate carcinoma. The transcriptional activity of androgen receptor (AR) is regulated by positive or negative transcriptional cofactors. We report here that PIAS1, PIAS3, and PIASy of the protein inhibitor of activated STAT (PIAS) family, which are expressed in human prostate, display distinct effects on AR-mediated gene activation in prostate cancer cells. While PIAS1 and PIAS3 enhance the transcriptional activity of AR, PIASy acts as a potent inhibitor of AR in prostate cancer cells. The effects of PIAS proteins on AR are competitive. PIASy binds to AR but does not affect the DNA binding activity of AR. An NH2-terminal LXXLL signature motif of PIASy, although not required for PIASy-AR interaction, is essential for the transrepression activity of PIASy. Our results identify PIASy as a transcriptional corepressor of AR and suggest that different PIAS proteins have distinct effects on AR signaling in prostate cancer cells.

An androgen receptor (AR) interacting protein was isolated from a HeLa cell complementary DNA library by two-hybrid screening in yeast using the AR DNA and ligand binding domains [amino acids (aa) 481-919] as bait. AR binding of the protein in yeast was dependent on the presence of testosterone or dihydrotestosterone (DHT). The isolated protein is identical to thyroid receptor activator molecule TRAM-1 but lacking aa 1-458. TRAM-1 is a steroid receptor coactivator-3 (SRC-3) subtype. In affinity matrix assays, 35S-labeled TRAM-1 bound the GST-AR ligand binding domain (aa 624-919) and GST-AR N-terminal and DNA binding domains (aa 1-660), but not the GST-AR DNA binding domain (aa 544-634) alone. Coexpression of TRAM-1 increased DHT-dependent AR transactivation 5-fold and constitutive activity of AR (aa 1-660) N-terminal and DNA-binding domains increased 9-fold. Full-length TRAM-1 (aa 1-1424) and the partial (aa 459-1424) were AR and GR coactivators as was SRC-1. In human testis, immunostaining of SRC-3 colocalized with AR in nuclei of Sertoli cells and peritubular myoid cells, indicating it could function as an AR coactivator in these cells. SRC-3 was also present in nuclei of spermatogenic cells where AR was not expressed, suggesting it might also be a coactivator with other nuclear receptors that regulate spermatogenesis.

Androgen (AR) and glucocorticoid (GR) receptors recognize a family of 15 base pair partial palindromic hormone response elements (HRE). We have studied receptor interactions with several HREs from androgen regulated genes to determine their potential to mediate a selective androgen response. Synthetic oligonucleotides corresponding to the elements were analysed for receptor binding and steroid dependent transcriptional enhancer activities. Each HRE contained the 3′ half-site sequence (5′-TGTNCT-3′) of the glucocorticoid response element (GRE) consensus sequence. HREs that countained the 5′ half-site GRE consensus sequence (5′-A/GGNACA/G-3′) had the strongest and-rogen response element (ARE) and GRE activities. In methylation interference assays, AR and GR interacted with identical base contact sites in the response elements. Two elements that deviated from the GRE consensus sequence by a single optimal base in the 5′ half, had reduced ARE activity with no significant change in GRE activity and displayed lower binding of AR than GR in mobility shift assays using purified DNA binding domain peptides. Transfections with AR/GR and GR/AR chimeras containing the N-terminal domain of one receptor linked to the DNA-binding and C-terminal domains of the other suggested that N-terminal domain functions of GR also contributed to the greater GRE than ARE activities of the response elements.

Publisher Summary This chapter presents studies of the androgen insensitivity syndrome (AIS). AIS is also referred to as testicular feminization (Tfm) in rats, mice, and humans. Studies have established that this disorder is linked to the X chromosome. Androgen insensitivity is characterized by lack of a target cell response to androgen. To localize the androgen receptor (AR) gene, a series of Tfm mouse–human cell hybrids containing X chromosome are produced. Expression of AR androgen binding indicated that either the AR locus or a factor controlling AR expression is located on the human X chromosome near the centromere. Cloning the AR enables a demonstration that it is the AR structural gene that occupies this locus on the X chromosome and establishes that a mutation in the AR gene can result in AIS. AR belongs to the subfamily of steroid hormone receptors within a larger family of nuclear proteins that likely evolved from a common ancestral gene. Each contains an amino-terminal region, which is variable in length and could have a role in transcriptional activation, a central cysteine-rich DNA-binding domain, and a carboxyl-terminal ligand-binding domain. The chapter presents a comparison of human and rat AR cDNAs—complementary DNAs.

A composite androgen receptor DNA sequence 4,181 base pairs in length was determined from three cDNA clones isolated from a rat epididymal bacteriophage lambda gt11 library. An open reading frame of 902 amino acids encodes a protein of 98,227 mol wt. Structural domains characteristic of the steroid receptor family include an amino-terminal region with five repeated amino acid motifs, a central DNA-binding domain homologous with other steroid receptors, and a carboxyl-terminal steroid-binding region. A receptor cDNA probe used in Northern blot analysis hybridized with a predominant 10-kilobase androgen receptor mRNA in male reproductive tissues of the rat. Autoregulation of androgen receptor mRNA was indicated in rat ventral prostate by an increase in the level of 10-kilobase mRNA after castration and suppression of receptor mRNA upon androgen restimulation. A 15 amino acid peptide with sequence derived from the deduced androgen receptor sequence was synthesized and used as immunogen in raising receptor antibodies in rabbits. Antisera reacted with high titer against the synthetic peptide by enzyme-linked immunosorbent assay and against the native [3H]dihydrotestosterone-labeled androgen receptor as evidenced by an increase in receptor sedimentation rate determined by sucrose gradient centrifugation. Immunocytochemical staining localized the androgen receptor to epithelial cell nuclei in rat ventral prostate.

Androgenic hormones mediate their effects on male sex differentiation and development through a high affinity receptor protein. We report here cloning of the complete coding sequence of the human androgen receptor (hAR). By sequence homology hAR is a member of the nuclear receptor family, with closest sequence identity to the progesterone, mineralocorticoid, and glucocorticoid receptors. Regions of highest homology include the DNA-binding domain and a small region within the hydrophobic ligand-binding domain. Comparison of the deduced 919 amino acid sequence of hAR (98,999 mol wt) to the 902 amino acid sequence of rat AR (98,227 mol wt) reveals identical sequences in the DNA- and hormone-binding domains, with an overall homology of 85%. In human prostate, the major androgen receptor mRNA species is 10 kilobases while a less abundant mRNA is approximately 7 kilobases. Rabbit polyclonal antibodies were raised against a synthetic peptide from the N-terminal region of hAR. Immunocytochemical analysis of human prostate tissue demonstrated that AR is localized predominantly in nuclei of glandular epithelial cells.