The Hawaiian Islands form as the Pacific Plate moves over a 'hot spot' in the earth's mantle where magma extrudes through the crust to build huge shield volcanos. The islands subside and erode as the plate carries them to the north-west, eventually to become coral atolls and seamounts. Thus islands are ordered linearly by age, with the oldest islands in the north-west (e.g. Kauai at 5.1 Ma) and the youngest in the south-east (e.g. Hawaii at 0.43 Ma). K-Ar estimates of the date of an island's formation provide a maximum age for the taxa inhabiting the island. These ages can be used to calibrate rates of molecular change under the following assumptions: (i) K-Ar dates are accurate; (ii) tree topologies show that derivation of taxa parallels the timing of island formation; (iii) populations do not colonize long after island emergence; (iv) the coalescent point for sister taxa does not greatly predate the formation of the colonized younger island; (v) saturation effects and (vi) among-lineage rate variation are minimal or correctable; and (vii) unbiased standard errors of distances and regressions can be estimated from multiple pairwise comparisons. We use the approach to obtain overall corrected rate calibrations for: (i) part of the mitochondrial cytochrome b gene in Hawaiian drepanidines (0.016 sequence divergence/Myr); (ii) the Yp1 gene in Hawaiian Drosophila (0.019/Myr Kambysellis et al. 1995); and (iii) parts of the mitochondrial 12S and 16S rRNA and tRNAval in Laupala crickets (0.024-0.102/Myr, Shaw 1996). We discuss the reliability of the estimates given the assumptions (i-vii) above and contrast the results with previous calibrations of Adh in Hawaiian Drosophila and chloroplast DNA in lobeliods.

Transcription of the prostate-specific antigen (PSA) gene escapes regulation by androgens in advanced prostate cancer. To determine the molecular mechanism(s) of androgen-independent regulation of the PSA gene, the possibility that the androgen receptor (AR) is activated in the absence of androgen by stimulation of protein kinase A (PKA) was investigated. Activation of PKA by forskolin resulted in elevated expression of the PSA gene in androgen-depleted LNCaP cells, an effect that was blocked by the antiandrogen, bicalutamide. Further evidence that induction of PSA gene expression was dependent on AR was obtained from experiments using PC3 cells devoid of AR. Neither PSA, PB, nor ARR3 androgen-responsive reporters could be induced by activation of PKA in the absence of transfected AR. In addition, when nuclear AR from forskolin-treated LNCaP cells was incubated with oligonucleotides encoding an androgen response element of the PSA promoter and examined by electromobility shift assay, an increase in AR-androgen response element complex formation was observed. Lastly, cotransfection of an expression vector for a chimeric protein encoding the amino-terminal domain of the human AR linked to Gal4 and a 5xGal4UAS reporter gene construct resulted in activation of the amino-terminal domain of the AR by stimulation of PKA activity. These results demonstrate androgen-independent induction of PSA gene expression in prostate cancer cells by an AR-dependent pathway.

Cone photoreceptor disorders form a clinical spectrum of diseases that include progressive cone dystrophy (CD) and complete and incomplete achromatopsia (ACHM). The underlying disease mechanisms of autosomal recessive (ar)CD are largely unknown. Our aim was to identify causative genes for these disorders by genome-wide homozygosity mapping. We investigated 75 ACHM, 97 arCD, and 20 early-onset arCD probands and excluded the involvement of known genes for ACHM and arCD. Subsequently, we performed high-resolution SNP analysis and identified large homozygous regions spanning the PDE6C gene in one sibling pair with early-onset arCD and one sibling pair with incomplete ACHM. The PDE6C gene encodes the cone alpha subunit of cyclic guanosine monophosphate (cGMP) phosphodiesterase, which converts cGMP to 5'-GMP, and thereby plays an essential role in cone phototransduction. Sequence analysis of the coding region of PDE6C revealed homozygous missense mutations (p.R29W, p.Y323N) in both sibling pairs. Sequence analysis of 104 probands with arCD and 10 probands with ACHM revealed compound heterozygous PDE6C mutations in three complete ACHM patients from two families. One patient had a frameshift mutation and a splice defect; the other two had a splice defect and a missense variant (p.M455V). Cross-sectional retinal imaging via optical coherence tomography revealed a more pronounced absence of cone photoreceptors in patients with ACHM compared to patients with early-onset arCD. Our findings identify PDE6C as a gene for cone photoreceptor disorders and show that arCD and ACHM constitute genetically and clinically overlapping phenotypes.

Diabetic neuropathy is the most common peripheral neuropathy in western countries. Although every effort has been made to clarify the pathogenic mechanism of diabetic neuropathy, thereby devising its ideal therapeutic drugs, neither convinced hypotheses nor unequivocally effective drugs have been established. In view of the pathologic basis for the treatment of diabetic neuropathy, it is important to enhance nerve regeneration as well as prevent nerve degeneration. Nerve regeneration or sprouting in diabetes may occur not only in the nerve trunk but also in the dermis and around dorsal root ganglion neurons, thereby being implicated in the generation of pain sensation. Thus, inadequate nerve regeneration unequivocally contributes to the pathophysiologic mechanism of diabetic neuropathy. In this context, the research on nerve regeneration in diabetes should be more accelerated. Indeed, nerve regenerative capacity has been shown to be decreased in diabetic patients as well as in diabetic animals. Disturbed nerve regeneration in diabetes has been ascribed at least in part to all or some of decreased levels of neurotrophic factors, decreased expression of their receptors, altered cellular signal pathways and/or abnormal expression of cell adhesion molecules, although the mechanisms of their changes remain almost unclear. In addition to their steady-state changes in diabetes, nerve injury induces injury-specific changes in individual neurotrophic factors, their receptors and their intracellular signal pathways, which are closely linked with altered neuronal function, varying from neuronal survival and neurite extension/nerve regeneration to apoptosis. Although it is essential to clarify those changes for understanding the mechanism of disturbed nerve regeneration in diabetes, very few data are now available. Rationally accepted replacement therapy with neurotrophic factors has not provided any success in treating diabetic neuropathy. Aside from adverse effects of those factors, more rigorous consideration for their delivery system may be needed for any possible success. Although conventional therapeutic drugs like aldose reductase (AR) inhibitors and vasodilators have been shown to enhance nerve regeneration, their efficacy should be strictly evaluated with respect to nerve regenerative capacity. For this purpose, especially clinically, skin biopsy, by which cutaneous nerve pathology including nerve regeneration can be morphometrically evaluated, might be a safe and useful examination.

Taxol chemotherapy is one of the few therapeutic options for men with castration-resistant prostate cancer (CRPC). However, the working mechanisms for Taxol are not fully understood. Here, we showed that treatment of 22Rv1, a PTEN-positive CRPC cell line, with paclitaxel and its semisynthetic analogue docetaxel decreases expression of the androgen receptor (AR)-activated genes prostate-specific antigen (PSA) and Nkx3.1 but increases expression of the AR repression gene maspin, suggesting that Taxol treatment inhibits AR activity. This was further supported by the observation that the activity of AR luciferase reporter genes was inhibited by paclitaxel. In contrast, paclitaxel treatment failed to inhibit AR activity in the PTEN-null CRPC cell line C4-2. However, pretreatment of C4-2 cells with the phosphoinositide 3-kinase inhibitor LY294002 restored paclitaxel inhibition of the AR. Treatment of 22Rv1 xenografts in mice with docetaxel induced mitotic arrest and a decrease in PSA expression in tumor cells adjacent to vascular vessels. We further showed that paclitaxel induces nuclear accumulation of FOXO1, a known AR suppressive nuclear factor, and increases the association of FOXO1 with AR proteins in the nucleus. FOXO1 knockdown with small interfering RNA attenuated the inhibitory effect of paclitaxel on AR transcriptional activity, expression of PSA and Nkx3.1, and cell survival. These data reveal a previously uncharacterized, FOXO1-mediated, AR-inhibitory effect of Taxol in CRPC cells that may play an important role in Taxol-mediated inhibition of CRPC growth.

Inhibitors of histone deacetylases have been approved for clinical application in cancer treatment. On the other hand, histone acetyltransferase (HAT) inhibitors have been less extensively investigated for their potential use in cancer therapy. In prostate cancer, the HATs and coactivators p300 and CBP are upregulated and may induce transcription of androgen receptor (AR)-responsive genes, even in the absence or presence of low levels of AR. To discover a potential anticancer effect of p300/CBP inhibition, we used two different approaches: (i) downregulation of p300 and CBP by specific short interfering RNA (siRNA) and (ii) chemical inhibition of the acetyltransferase activity by a newly developed small molecule, C646. Knockdown of p300 by specific siRNA, but surprisingly not of CBP, led to an increase of caspase-dependent apoptosis involving both extrinsic and intrinsic cell death pathways in androgen-dependent and castration-resistant prostate cancer cells. Induction of apoptosis was mediated by several pathways including inhibition of AR function and decrease of the nuclear factor kappa B (NF-κB) subunit p65. Furthermore, cell invasion was decreased upon p300, but not CBP, depletion and was accompanied by lower matrix metalloproteinase (MMP)-2 and MMP-9 transcriptions. Thus, p300 and CBP have differential roles in the processes of survival and invasion of prostate cancer cells. Induction of apoptosis in prostate cancer cells was confirmed by the use of C646. This was substantiated by a decrease of AR function and downregulation of p65 impairing several NF-κB target genes. Taken together, these results suggest that p300 inhibition may be a promising approach for the development of new anticancer therapies.

Quantitative and structural genetic alterations cause the development and progression of prostate cancer. A number of genes have been implicated in prostate cancer by genetic alterations and functional consequences of the genetic alterations. These include the ELAC2 (HPC2), MSR1, and RNASEL (HPC1) genes that have germline mutations in familial prostate cancer; AR, ATBF1, EPHB2 (ERK), KLF6, mitochondria DNA, p53, PTEN, and RAS that have somatic mutations in sporadic prostate cancer; AR, BRCA1, BRCA2, CHEK2 (RAD53), CYP17, CYP1B1, CYP3A4, GSTM1, GSTP1, GSTT1, PON1, SRD5A2, and VDR that have germline genetic variants associated with either hereditary and/or sporadic prostate cancer; and ANXA7 (ANX7), KLF5, NKX3-1 (NKX3.1), CDKN1B (p27), and MYC that have genomic copy number changes affecting gene function. More genes relevant to prostate cancer remain to be identified in each of these gene groups. For the genes that have been identified, most need additional genetic, functional, and/or biochemical examination. Identification and characterization of these genes will be a key step for improving the detection and treatment of prostate cancer.

Glomerular targets of autoimmunity in human membranous nephropathy are poorly understood. Here, we used a combined proteomic approach to identify specific antibodies against podocyte proteins in both serum and glomeruli of patients with membranous nephropathy (MN). We detected specific anti-aldose reductase (AR) and anti-manganese superoxide dismutase (SOD2) IgG(4) in sera of patients with MN. We also eluted high titers of anti-AR and anti-SOD2 IgG(4) from microdissected glomeruli of three biopsies of MN kidneys but not from biopsies of other glomerulonephritides characterized by IgG deposition (five lupus nephritis and two membranoproliferative glomerulonephritis). We identified both antigens in MN biopsies but not in other renal pathologies or normal kidney. Confocal and immunoelectron microscopy (IEM) showed co-localization of anti-AR and anti-SOD2 with IgG(4) and C5b-9 in electron-dense podocyte immune deposits. Preliminary in vitro experiments showed an increase of SOD2 expression on podocyte plasma membrane after treatment with hydrogen peroxide. In conclusion, our data support AR and SOD2 as renal antigens of human MN and suggest that oxidative stress may drive glomerular SOD2 expression.

Primary immunodeficiencies (PIDs) were long thought to be exclusively recessive traits -- autosomal recessive (AR) in most cases, with a few X-linked recessive (XR) diseases. In recent years, autosomal dominant (AD), mitochondrial, polygenic, and even somatic PIDs have been described. However, AR remains the most frequent inheritance pattern among recently described PIDs. Some PIDs have been shown to be genetically heterogeneous. Mendelian susceptibility to mycobacterial diseases (MSMD) displays a high level of genetic heterogeneity. There are 6 MSMD-causing genes, including 1 X-linked gene (nuclear factor-kappaB-essential modulator [NEMO]) and 5 autosomal genes (IFN-gamma receptor 1 [IFNGR1], IFN-gamma receptor 2 [IFNGR2], signal transducer and activator of transcription 1 [STAT1], IL-12 p40 subunit [IL12P40], and IL-12 receptor beta-subunit [IL12RB1]). The X-linked trait is XR; STAT1 deficiency is AD; the IFNGR2, IL12P40 subunit, and IL12RB1 deficiencies are AR; and IFNGR1 deficiency may be AD or AR. Two of the AR traits (IFNGR1, IFNGR2) may be subdivided into complete and partial deficiencies, and 3 AR complete deficiencies (IFNGR1, IFNGR2, IL12RB1) may be subdivided into disorders with and without cell surface expression. Finally, there are 2 types of AD STAT1 deficiency, depending on whether the mutation impairs phosphorylation or DNA binding. Thirteen genetic disorders conferring MSMD have been described, involving 1 XR, 3 AD (2 genes), and 9 AR traits (4 genes). However, no genetic etiology has yet been identified for about half of all patients with MSMD. We expect to identify new XR and AD causes of MSMD, but new AR etiologies of MSMD are also likely to be discovered. The investigation of children from areas in which consanguineous marriages are common will probably facilitate the description of many more AR traits.

The androgen-signaling pathway plays an important role in the development and hormonal progression of prostate cancer to the castrate-resistant stage (also called androgen-independent or hormone refractory). The Wnt pathway and beta-catenin contribute to prostate biology and pathology. Here application of Affymetrix GeneChip analysis revealed the genomic similarity of the LNCaP hollow fiber model to clinical samples and identified genes with differential expression during hormonal progression. The fiber model samples clustered according to the expression profile of androgen-regulated genes to provide genomic evidence for the reactivation of the AR signaling pathway in castrate-resistant prostate cancer. Pathway-based characterization of gene expression identified activation of the Wnt pathway. Together with the increased expression of AR and beta-catenin, there was increased nuclear colocalization and interaction of endogenous AR and beta-catenin in castrate-resistant prostate cancer from castrated mice. Surprisingly, no interaction or colocalization of AR and beta-catenin could be detected in xenografts from noncastrated mice. These studies provide the first in vivo evidence to support aberrant activation of the AR through the Wnt/beta-catenin signaling pathway during progression of prostate cancer to the terminal castrate-resistant stage.

Androgens are essential for the development, growth, and maintenance of the prostate. They exert their effects via the intracellular androgen receptor (AR), which is a ligand-dependent transcription activator. As is the case with normal prostate development, primary prostatic cancers are largely dependent on androgens for growth and survival. Most patients respond favorably to androgen ablation and antiandrogen therapy, which has become a standard treatment of metastatic disease. However, virtually all patients will relapse with clinically defined androgen-independent cancer. This phenomenon raises the question of how cancer cells survive and grow in the low androgen environment? Two of the routes cells can take to adapt are (1) bypassing and (2) sensitizing the AR pathway. The vast numbers of AR abnormalities observed in prostate tumors from patients treated with hormonal therapy suggest that many cells sensitize or change the AR pathway. To continue to activate this pathway in a low androgen environment, cells can (1) mutate the AR to become promiscuously activated by different steroids, (2) amplify the AR, (3) activate the AR in a ligand-independent manner by growth factors and cytokines, or (4) amplify coactivators. Alternatively, prostate cancer cells that have lost AR expression must have bypassed the AR pathway. Activation of oncogenes and autocrine growth factor stimulation are two mechanisms that likely contribute to becoming completely androgen-independent. From all the studies on AR function in prostate cancer, it is clear that the AR plays an important role in cancer development and progression. Moreover, the AR pathway remains important in most cells from patients with clinically defined androgen-independent prostate cancer.

OBJECTIVE

Progression to the castration-resistant state is the incurable and lethal end stage of prostate cancer, and there is strong evidence that androgen receptor (AR) still plays a central role in this process. We hypothesize that knocking down AR will have a major effect on inhibiting growth of castration-resistant tumors.

METHODS

Castration-resistant C4-2 human prostate cancer cells stably expressing a tetracycline-inducible AR-targeted short hairpin RNA (shRNA) were generated to directly test the effects of AR knockdown in C4-2 human prostate cancer cells and tumors.

RESULTS

In vitro expression of AR shRNA resulted in decreased levels of AR mRNA and protein, decreased expression of prostate-specific antigen (PSA), reduced activation of the PSA-luciferase reporter, and growth inhibition of C4-2 cells. Gene microarray analyses revealed that AR knockdown under hormone-deprived conditions resulted in activation of genes involved in apoptosis, cell cycle regulation, protein synthesis, and tumorigenesis. To ensure that tumors were truly castration-resistant in vivo, inducible AR shRNA expressing C4-2 tumors were grown in castrated mice to an average volume of 450 mm(3). In all of the animals, serum PSA decreased, and in 50% of them, there was complete tumor regression and disappearance of serum PSA.

CONCLUSIONS

Whereas castration is ineffective in castration-resistant prostate tumors, knockdown of AR can decrease serum PSA, inhibit tumor growth, and frequently cause tumor regression. This study is the first direct evidence that knockdown of AR is a viable therapeutic strategy for treatment of prostate tumors that have already progressed to the castration-resistant state.

OBJECTIVE

This study was undertaken for the comparison of local resection for early rectal carcinomas using transanal endoscopic microsurgery or anterior resection.

METHODS

Data from 50 of 52 patients with proven adenocarcinoma (GI/II) and intraluminal ultrasound with Stage uT1 N negative (uTNM) were evaluated in a prospective randomized study with two therapeutic arms: transanal endoscopic microsurgery (TEM; n = 24) or anterior resection (AR; n = 26), performed under general anesthesia.

RESULTS

Patients' ages and rectal tumor locations showed insignificant differences of distribution in comparison of TEM with AR (Welsh's alternate t-test; t-test). Local recurrence (4.2 percent) and five-year survival rates (96 percent) differed insignificantly (log-rank test). Early postoperative mortality was zero. Significant differences were found comparing time of hospitalization, loss of blood, operation time, and opiate analgesia (Welsh's alternate t-test; Wilcoxon's test; each P < 0.05). Early and late morbidity differed considerably.

CONCLUSIONS

Lower morbidity, similar local recurrence, and survival rates favor the TEM technique. Comparable results in survival rate to the gold standard (AR) are objective arguments for choosing the adequate surgical procedure. For early rectal cancer, the minimum invasive TEM technique should be preferred because of superior overview during operation with safer suturing after meticulous full wall thickness excision.

Resistance to antiandrogen therapy in patients with metastatic prostate cancer poses a major challenge, which, if overcome, may lead to significant advances in the treatment of these patients. Hormone resistance of prostate cancer develops, in part, from upregulation of antiapoptotic genes after androgen deprivation. Given the accumulating evidence that Survivin, a new member of the inhibitor of apoptosis (IAP) family, is associated with both cancer progression and drug resistance, we hypothesized that Survivin plays a potentially important role in hormone therapy resistance, and that targeting of Survivin may enhance sensitivity to antiandrogen therapy in prostate cancer. Patterns of Survivin expression were assessed in three prostate cancer cell lines LNCaP, PC-3, and DU-145 using quantitative Western analysis. All three cell lines were found to strongly express Survivin. In LNCaP cells with intact androgen receptors (ARs), it was observed that androgen stimulation with 5alpha-dihydrotestosterone (DHT) increased Survivin expression. Conversely, treatment with Flutamide decreased Survivin expression in LNCaP cells. We next studied the functional effect of Survivin on sensitivity to Flutamide. LNCaP cells were infected with replication-deficient adenoviruses encoding either wild-type Survivin pAd-S(WT) or a phosphorylation-defective Survivin Thr34 ->> Ala dominant-negative mutant pAd-S(T34A), and then treated with Flutamide. Cell viability and apoptosis were assessed in vitro and in vivo. It was determined that Survivin can mediate resistance to such antiandrogen therapies based on our assays. Direct androgen stimulation resulted in pan-cell cycle expression of Survivin, which was found to be mediated by AKT, as it was determined that exogenous insulin-like growth factor-1 (IGF-1), a known activator of AKT signaling, could increase Survivin expression and result in pan-cell cycle expression even in AR-negative prostate cancer cell lines PC-3 and DU-145. Given this alternative mechanism of Survivin expression and our findings that Survivin can mediate resistance to Flutamide treatment, we further investigated whether IGF-1-mediated activation of Survivin via AKT could mediate resistance to antiandrogen therapy. Both in vitro and in vivo, this was found to be the case, supporting a novel mechanism of resistance to antiandrogen therapy. Our study indicates that upregulation of Survivin via IGF-1 signaling confers resistance to Flutamide in prostate cancer cells. Targeted inhibition of Survivin appears to enhance the therapeutic effects of Flutamide in vitro and in vivo, revealing a novel strategy to enhance sensitivity to androgen ablation therapy.

The CAG repeat expansions that occur in translated regions of specific genes can cause human genetic disorders known as polyglutamine (poly-Q)-triggered diseases. Huntington's disease and spinobulbar muscular atrophy (SBMA) are examples of these diseases in which underlying mutations are localized near other trinucleotide repeats in the huntingtin (HTT) and androgen receptor (AR) genes, respectively. Mutant proteins that contain expanded polyglutamine tracts are well-known triggers of pathogenesis in poly-Q diseases, but a toxic role for mutant transcripts has also been proposed. To gain insight into the structural features of complex triplet repeats of HTT and AR transcripts, we determined their structures in vitro and showed the contribution of neighboring repeats to CAG repeat hairpin formation. We also demonstrated that the expanded transcript is retained in the nucleus of human HD fibroblasts and is colocalized with the MBNL1 protein. This suggests that the CAG repeats in the HTT mRNA adopt ds-like RNA conformations in vivo. The intracellular structure of the CAG repeat region of mutant HTT transcripts was not sufficiently stable to be protected from cleavage by an siRNA targeting the repeats and the silencing efficiency was higher for the mutant transcript than for its normal counterpart.

Premature ovarian failure (POF) syndrome, an early decline of ovarian function in women, is frequently associated with X chromosome abnormalities ranging from various Xq deletions to complete loss of one of the X chromosomes. However, the genetic locus responsible for the POF remains unknown, and no candidate gene has been identified. Using the Cre/LoxP system, we have disrupted the mouse X chromosome androgen receptor (Ar) gene. Female AR(-/-) mice appeared normal but developed the POF phenotype with aberrant ovarian gene expression. Eight-week-old female AR(-/-) mice are fertile, but they have lower follicle numbers and impaired mammary development, and they produce only half of the normal number of pups per litter. Forty-week-old AR(-/-) mice are infertile because of complete loss of follicles. Genome-wide microarray analysis of mRNA from AR(-/-) ovaries revealed that a number of major regulators of folliculogenesis were under transcriptional control by AR. Our findings suggest that AR function is required for normal female reproduction, particularly folliculogenesis, and that AR is a potential therapeutic target in POF syndrome.

Weill-Marchesani syndrome (WMS) is a connective tissue disorder characterised by short stature, brachydactyly, joint stiffness, and characteristic eye anomalies including microspherophakia, ectopia of the lenses, severe myopia, and glaucoma. Both autosomal recessive (AR) and autosomal dominant (AD) modes of inheritance have been described and a gene for AR WMS has recently been mapped to chromosome 19p13.3-p13.2. Here, we report on the exclusion of chromosome 19p13.3-p13.2 in a large AD WMS family and show that, despite clinical homogeneity, AD and AR WMS are genetically heterogeneous entities. Because two AD WMS families were consistent with linkage to chromosome 15q21.1, the fibrillin-1 gene was sequenced and a 24 nt in frame deletion within a latent transforming growth factor-beta1 binding protein (LTBP) motif of the fibrillin-1 gene was found in a AD WMS family (exon 41, 5074_5097del). This in frame deletion cosegregated with the disease and was not found in 186 controls. This study strongly suggests that AD WMS and Marfan syndrome are allelic conditions at the fibrillin-1 locus and adds to the remarkable clinical heterogeneity of type I fibrillinopathies.

Dandy-Walker malformation (DWM), the most common human cerebellar malformation, has only one characterized associated locus. Here we characterize a second DWM-linked locus on 6p25.3, showing that deletions or duplications encompassing FOXC1 are associated with cerebellar and posterior fossa malformations including cerebellar vermis hypoplasia (CVH), mega-cisterna magna (MCM) and DWM. Foxc1-null mice have embryonic abnormalities of the rhombic lip due to loss of mesenchyme-secreted signaling molecules with subsequent loss of Atoh1 expression in vermis. Foxc1 homozygous hypomorphs have CVH with medial fusion and foliation defects. Human FOXC1 heterozygous mutations are known to affect eye development, causing a spectrum of glaucoma-associated anomalies (Axenfeld-Rieger syndrome, ARS; MIM no. 601631). We report the first brain imaging data from humans with FOXC1 mutations and show that these individuals also have CVH. We conclude that alteration of FOXC1 function alone causes CVH and contributes to MCM and DWM. Our results highlight a previously unrecognized role for mesenchyme-neuroepithelium interactions in the mid-hindbrain during early embryogenesis.

Tobacco smoke contains multiple classes of established carcinogens including benzo(a)pyrenes, polycyclic aromatic hydrocarbons, and tobacco-specific nitrosamines. Most of these compounds exert their genotoxic effects by forming DNA adducts and generation of reactive oxygen species, causing mutations in vital genes such as K-Ras and p53. In addition, tobacco-specific nitrosamines can activate nicotinic acetylcholine receptors (nAChR) and to a certain extent β-adrenergic receptors (β-AR), promoting cell proliferation. Furthermore, it has been demonstrated that nicotine, the major addictive component of tobacco smoke, can induce cell-cycle progression, angiogenesis, and metastasis of lung and pancreatic cancers. These effects occur mainly through the α7-nAChRs, with possible contribution from the β-ARs and/or epidermal growth factor receptors. This review article will discuss the molecular mechanisms by which nicotine and its oncogenic derivatives such as 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone and N-nitrosonornicotine induce cell-cycle progression and promote tumor growth. A variety of signaling cascades are induced by nicotine through nAChRs, including the mitogen-activated protein kinase/extracellular signal-regulated kinase pathway, phosphoinositide 3-kinase/AKT pathway, and janus-activated kinase/STAT signaling. In addition, studies have shown that nAChR activation induces Src kinase in a β-arrestin-1-dependent manner, leading to the inactivation of Rb protein and resulting in the expression of E2F1-regulated proliferative genes. Such nAChR-mediated signaling events enhance the proliferation of cells and render them resistant to apoptosis induced by various agents. These observations highlight the role of nAChRs in promoting the growth and metastasis of tumors and raise the possibility of targeting them for cancer therapy.

In the present study we characterize the properties of the potent MCT1 (monocarboxylate transporter 1) inhibitor AR-C155858. Inhibitor titrations of L-lactate transport by MCT1 in rat erythrocytes were used to determine the Ki value and number of AR-C155858-binding sites (Et) on MCT1 and the turnover number of the transporter (kcat). Derived values were 2.3+/-1.4 nM, 1.29+/-0.09 nmol per ml of packed cells and 12.2+/-1.1 s-1 respectively. When expressed in Xenopus laevis oocytes, MCT1 and MCT2 were potently inhibited by AR-C155858, whereas MCT4 was not. Inhibition of MCT1 was shown to be time-dependent, and the compound was also active when microinjected, suggesting that AR-C155858 probably enters the cell before binding to an intracellular site on MCT1. Measurement of the inhibitor sensitivity of several chimaeric transporters combining different domains of MCT1 and MCT4 revealed that the binding site for AR-C155858 is contained within the C-terminal half of MCT1, and involves TM (transmembrane) domains 7-10. This is consistent with previous data identifying Phe360 (in TM10) and Asp302 plus Arg306 (TM8) as key residues in substrate binding and translocation by MCT1. Measurement of the Km values of the chimaeras for L-lactate and pyruvate demonstrate that both the C- and N-terminal halves of the molecule influence transport kinetics consistent with our proposed molecular model of MCT1 and its translocation mechanism that requires Lys38 in TM1 in addition to Asp302 and Arg306 in TM8 [Wilson, Meredith, Bunnun, Sessions and Halestrap (2009) J. Biol. Chem. 284, 20011-20021].

Androgens play pivotal roles in the regulation of male development and physiological processes, particularly in the male reproductive system. Most biological effects of androgens are mediated by the action of nuclear androgen receptor (AR). AR acts as a master regulator of downstream androgen-dependent signaling pathway networks. This ligand-dependent transcriptional factor modulates gene expression through the recruitment of various coregulator complexes, the induction of chromatin reorganization, and epigenetic histone modifications at target genomic loci. Dysregulation of androgen/AR signaling perturbs normal reproductive development and accounts for a wide range of pathological conditions such as androgen-insensitive syndrome, prostate cancer, and spinal bulbar muscular atrophy. In this review we summarize recent advances in understanding of the epigenetic mechanisms of AR action as well as newly recognized aspects of AR-mediated androgen signaling in both men and women. In addition, we offer a perspective on the use of animal genetic model systems aimed at eventually developing novel therapeutic AR ligands.

Prostate cancer is the most commonly diagnosed malignancy among men in industrialized countries, accounting for the second leading cause of cancer-related deaths. Although we now know that the androgen receptor (AR) is important for progression to the deadly advanced stages of the disease, it is poorly understood what AR-regulated processes drive this pathology. Here we demonstrate that AR regulates prostate cancer cell growth via the metabolic sensor 5'-AMP-activated protein kinase (AMPK), a kinase that classically regulates cellular energy homeostasis. In patients, activation of AMPK correlated with prostate cancer progression. Using a combination of radiolabeled assays and emerging metabolomic approaches, we also show that prostate cancer cells respond to androgen treatment by increasing not only rates of glycolysis, as is commonly seen in many cancers, but also glucose and fatty acid oxidation. Importantly, this effect was dependent on androgen-mediated AMPK activity. Our results further indicate that the AMPK-mediated metabolic changes increased intracellular ATP levels and peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α)-mediated mitochondrial biogenesis, affording distinct growth advantages to the prostate cancer cells. Correspondingly, we used outlier analysis to determine that PGC-1α is overexpressed in a subpopulation of clinical cancer samples. This was in contrast to what was observed in immortalized benign human prostate cells and a testosterone-induced rat model of benign prostatic hyperplasia. Taken together, our findings converge to demonstrate that androgens can co-opt the AMPK-PGC-1α signaling cascade, a known homeostatic mechanism, to increase prostate cancer cell growth. The current study points to the potential utility of developing metabolic-targeted therapies directed toward the AMPK-PGC-1α signaling axis for the treatment of prostate cancer.

After-ripening (AR) is a time and environment regulated process occurring in the dry seed, which determines the germination potential of seeds. Both metabolism and perception of the phytohormone abscisic acid (ABA) are important in the initiation and maintenance of dormancy. However, molecular mechanisms that regulate the capacity for dormancy or germination through AR are unknown. To understand the relationship between ABA and AR, we analysed genome expression in Arabidopsis thaliana mutants defective in seed ABA synthesis (aba1-1) or perception (abi1-1). Even though imbibed mutant seeds showed no dormancy, they exhibited changes in global gene expression resulting from dry AR that were comparable with changes occurring in wild-type (WT) seeds. Core gene sets were identified that were positively or negatively regulated by dry seed storage. Each set included a gene encoding repression or activation of ABA function (LPP2 and ABA1, respectively), thereby suggesting a mechanism through which dry AR may modulate subsequent germination potential in WT seeds. Application of exogenous ABA to after-ripened WT seeds did not reimpose characteristics of freshly harvested seeds on imbibed seed gene expression patterns. It was shown that secondary dormancy states reinstate AR status-specific gene expression patterns. A model is presented that separates the action of ABA in seed dormancy from AR and dry storage regulated gene expression. These results have major implications for the study of genetic mechanisms altered in seeds as a result of crop domestication into agriculture, and for seed behaviour during dormancy cycling in natural ecosystems.

Medicinal chemical approaches have been applied to all four of the adenosine receptor (AR) subtypes (A(1), A(2A), A(2B), and A(3)) to create selective agonists and antagonists for each. The most recent class of selective AR ligands to be reported is the class of A(2B)AR agonists. The availability of these selective ligands has facilitated research on therapeutic applications of modulating the ARs and in some cases has provided clinical candidates. Prodrug approaches have been developed which improve the bioavailability of the drugs, reduce side-effects, and/or may lead to site-selective effects. The A(2A) agonist regadenoson (Lexiscan®), a diagnostic drug for myocardial perfusion imaging, is the first selective AR agonist to be approved. Other selective agonists and antagonists are or were undergoing clinical trials for a broad range of indications, including capadenoson and tecadenoson (A(1) agonists) for atrial fibrillation, or paroxysmal supraventricular tachycardia, respectively, apadenoson and binodenoson (A(2A) agonists) for myocardial perfusion imaging, preladenant (A(2A) antagonist) for the treatment of Parkinson's disease, and CF101 and CF102 (A(3) agonists) for inflammatory diseases and cancer, respectively.