The Direct Binding of Insulin-like Growth Factor-1 (IGF-1) to Integrin ��v��3 Is Involved in IGF-1 Signaling

It has been proposed that ligand occupancy of integrin αvβ3 with extracellular matrix ligands (e.g. vitronectin) plays a critical role in insulin-like growth factor-1 (IGF-1) signaling. We found that expression of αvβ3 enhanced IGF-1-induced proliferation of Chinese hamster ovary cells in serum-free conditions (in the absence of vitronectin). We hypothesized that the direct integrin binding to IGF-1 may play a role in IGF-1 signaling. We demonstrated that αvβ3 specifically and directly bound to IGF-1 in cell adhesion, enzyme-linked immunosorbent assay-type binding, and surface plasmon resonance studies. We localized the amino acid residues of IGF-1 that are critical for integrin binding by docking simulation and mutagenesis. We found that mutating two Arg residues at positions 36 and 37 in the C-domain of IGF-1 to Glu (the R36E/R37E mutation) effectively reduced integrin binding. Interestingly, although the mutant still bound to IGF1R, it was defective in inducing IGF1R phosphorylation, AKT and ERK1/2 activation, and cell proliferation. Furthermore wild type IGF-1 mediated co-precipitation of αvβ3 and IGF1R, whereas the R36E/R37E mutant did not, suggesting that IGF-1 mediates the interaction between αvβ3 and IGF1R. These results suggest that the direct binding to IGF-1 to integrin αvβ3 plays a role in IGF-1 signaling through ternary complex formation (αvβ3-IGF-IGF1R), and integrin-IGF-1 interaction is a novel target for drug discovery.Integrins are a family of cell adhesion receptors that mediate cell-extracellular matrix (ECM)³ interaction and cell-cell interaction (1). It has been proposed that signaling from inside the cells regulates the ligand binding affinity of integrins (inside-out signaling) (2). Each integrin is a heterodimer containing α and β subunits. At present 18 α and 8 β subunits have been identified that combine to form 24 integrins (3).It has been reported that integrin αvβ3 plays a role in cancer proliferation and invasiveness. High levels of integrin αvβ3 correlate with growth and/or progression of melanoma (4, 5), neuroblastoma (6), breast cancer (7, 8), colon cancer (9), ovarian cancer (10), and cervical cancer (11). Moreover, individuals homozygous for the β3L33P polymorphism that enhances the ligand binding affinity of β3 integrins have an increased risk to develop breast cancer, ovarian cancer, and melanoma (12). However, it remains unclear whether and how increased levels of αvβ3 on tumor cells contribute to cancer development.Insulin-like growth factor-1 (IGF-1) is a polypeptide hormone (75 kDa) that has a high degree of structural similarity to human proinsulin. IGF-1 acts through binding to the type I IGF receptor (IGF1R), a receptor tyrosine kinase. The IGF1R is a heterotetramer that consists of two α-subunits that contain the ligand-binding domains and two β-subunits that contain the tyrosine kinase activity. After ligand binding, the receptor undergoes a conformational change resulting in the activation of the tyrosine kinase, which results in transphosphorylation of the opposite β-subunit on specific tyrosine residues. These phosphotyrosines then bind to adapter molecules such as Shc and IRS-1. Phosphorylation of these proteins leads to activation of the phosphatidylinositol 3-kinase and mitogen-activated protein kinase (MAPK) signaling pathways (reviewed in Ref. 13).IGF-1 has been implicated in cancer progression (14). One of the major actions of IGF-1 is to inhibit apoptosis. IGF-1 confers resistance to chemotherapy and radiation therapy. IGF-1 expression levels are increased in breast, lung, prostate, and many other cancers. Several strategies to target IGF-1 signaling have been extensively studied, including small interfering RNA and monoclonal antibodies for IGF1R and kinase inhibitors to inhibit the enzymatic activity of the receptor. The IGF-1 system is a therapeutic target for cancer, and elucidation of the IGF-1 signaling pathway should have a major impact in designing new therapeutic strategies.It has been proposed that ligand occupancy of αvβ3 with ECM ligands such as vitronectin plays a critical role in enhancing IGF-1 signaling (14). It has been reported that inhibiting αvβ3-ECM interaction (“ligand occupancy”) of αvβ3 inhibited IGF-1 actions selectively in cell types that express αvβ3 (14). Inhibiting ligand occupancy of αvβ3 blocked IGF-1-induced cell migration (15), DNA synthesis, IRS-1 phosphorylation, and IGF1R-linked downstream signaling events, such as activation of phosphatidylinositol 3-kinase and ERK1/2 (16).In the present study, we demonstrated that expression of αvβ3 enhanced proliferation of ovarian cancer cells in the presence of fetal bovine serum (FBS) and in serum-free conditions if IGF-1 was present. This suggests that IGF-1 is involved in enhanced proliferation of αvβ3-expressing cells. We demonstrated that αvβ3 bound to IGF-1 in several different binding assays. We found that two Arg residues at positions 36 and 37 in the C-domain of IGF-1 are critical for integrin binding by docking simulation and mutagenesis. Mutation of these Arg residues to Glu (the R36E/R37E mutation) effectively reduced integrin binding. Interestingly, the R36E/R37E mutant was defective in inducing cell proliferation and IGF-1 intracellular signaling, although it still bound to IGF1R. We demonstrated that wild type IGF-1 mediated co-precipitation of αvβ3 and IGF1R, whereas the R36E/R37E mutant did not, suggesting that IGF-1 mediates the interaction between αvβ3 and IGF1R. These results suggest that the direct binding to IGF-1 plays a role in IGF-1 signaling.     

C-Nucleosides via Glycosyl Alkynyl Ketones. Synthesis of 5(3)-Phenyl-3(5)-(β-D-ribofuranosyl) Pyrazole

Abstract

A β-D-ribofuranosyl phenylethynyl ketone (3) has been synthesized and shown to be a suitable intermediate for heterocyclic elaboration to C-nucleosides. Cyclization of 3 with hydrazine hydrate produces the title C-nucleoside.     

Transforming Growth Factor-�� (TGF-��1) Activates TAK1 via TAB1-mediated Autophosphorylation, Independent of TGF-�� Receptor Kinase Activity in Mesangial Cells

Transforming growth factor-β1 (TGF-β1) is a multifunctional cytokine that signals through the interaction of type I (TβRI) and type II (TβRII) receptors to activate distinct intracellular pathways. TAK1 is a serine/threonine kinase that is rapidly activated by TGF-β1. However, the molecular mechanism of TAK1 activation is incompletely understood. Here, we propose a mechanism whereby TAK1 is activated by TGF-β1 in primary mouse mesangial cells. Under unstimulated conditions, endogenous TAK1 is stably associated with TβRI. TGF-β1 stimulation causes rapid dissociation from the receptor and induces TAK1 phosphorylation. Deletion mutant analysis indicates that the juxtamembrane region including the GS domain of TβRI is crucial for its interaction with TAK1. Both TβRI-mediated TAK1 phosphorylation and TGF-β1-induced TAK1 phosphorylation do not require kinase activity of TβRI. Moreover, TβRI-mediated TAK1 phosphorylation correlates with the degree of its association with TβRI and requires kinase activity of TAK1. TAB1 does not interact with TGF-β receptors, but TAB1 is indispensable for TGF-β1-induced TAK1 activation. We also show that TRAF6 and TAB2 are required for the interaction of TAK1 with TβRI and TGF-β1-induced TAK1 activation in mouse mesangial cells. Taken together, our data indicate that TGF-β1-induced interaction of TβRI and TβRII triggers dissociation of TAK1 from TβRI, and subsequently TAK1 is phosphorylated through TAB1-mediated autophosphorylation and not by the receptor kinase activity of TβRI.Members of the transforming growth factor-β (TGF-β)³ superfamily are key regulators of various biological processes such as cellular differentiation, proliferation, apoptosis, and wound healing (1, 2). TGF-β1, the prototype of TGF-β family, is a potent inducer of extracellular matrix synthesis and is well established as a central mediator in the final common pathway of fibrosis associated with progressive kidney diseases (3, 4). Upon ligand stimulation, TGF-β type I (TβRI) and type II (TβRII) receptors form heterotetrameric complexes, by which TβRI is phosphorylated in the GS domain and activated. Smad signaling pathway is well established as a canonical pathway induced by TGF-β1 (5, 6). Receptor-regulated Smads (Smad2 and Smad3) are recruited and activated by the activated TβRI. The phosphorylation in the GS domain (7) and L45 loop (8) of TβRI are crucial for its interaction with receptor-regulated Smads. After phosphorylation, receptor-regulated Smads are rapidly dissociated from TβRI and interact with common Smad (Smad4) followed by nuclear translocation. In addition to the Smad pathway, a recently emerging body of evidence has demonstrated that TGF-β1 also induces various Smad-independent signaling pathways (9–17) by which mitogen-activated protein kinases (MAPKs), c-Jun N-terminal kinase (JNK) (18, 19), p38 MAPK (20–22), and extracellular signal-regulated kinase 1/2 (23, 24) can be activated by TGF-β1.TAK1, initially identified as a MAPK kinase kinase 7 (MKKK7 or MAP3K7) in the TGF-β signaling pathway (11, 12), also can be activated by environmental stress (25), proinflammatory cytokines such as IL-1 and TNF-α (26, 27) and lipopolysaccharide (28). For TAK1 activation, phosphorylation at Thr-187 and Ser-192 in the activation loop of TAK1 is essentially required (29–31). TAK1 can transduce signals to several downstream signaling cascades, including the MAPK kinase (MKK) 4/7-JNK cascade, MKK3/6-p38 MAPK cascade, and nuclear factor κB (NF-κB)-inducing kinase-IκB kinase cascade (26–28). A recent report has shown that TAK1 is also activated by agonists of AMP-activated kinase (AMPK) and ischemia, which in turn activates the LKB1/AMPK pathway, a pivotal energy-sensor pathway (32). TAK1 is also involved in Wnt signaling (33). We and others have previously demonstrated that TAK1 is a major mediator of TGF-β1-induced type I collagen and fibronectin expression through activation of the MKK3-p38 MAPK and MKK4-JNK signaling cascades, respectively (34–37). Furthermore, increased expression and activation of TAK1 enhance p38 phosphorylation and promote interstitial fibrosis in the myocardium from 9-day-old TAK1 transgenic mice (37). These data implicate a crucial role of TAK1 in extracellular matrix production and tissue fibrosis. TAK1 is also implicated in regulation of cell cycle (38), cell apoptosis (39–41), and the Smad signaling pathway (42–44). Thus, TAK1 may function as an important regulator and mediator of TGF-β1-induced Smad-dependent and Smad-independent signaling pathways.It has been demonstrated that TAK1 can be activated by the interaction with TAK1-binding protein 1 (TAB1) by in vitro binding assays and in overexpression studies (29–31); however, it is not clear whether TAB1 plays a crucial role in ligand-induced TAK1 activation. In embryonic fibroblasts from TAB1 null mice, IL-1 and TNF-α could induce TAK1-mediated NF-κB and JNK activation (45). TAK1 activation induced by TNF-α, IL-1, and T-cell receptor requires TAB2 or its homologous protein TAB3 (46–50). Although many questions still remain, much progress has been made in understanding the activation mechanism of TAK1 by inflammatory cytokines (46, 47, 51–53). Ligand binding of IL-1 receptor (IL-1R) results in recruitment of MyD88, which serves as an adaptor for IL-1 receptor-associated kinase (IRAK) 1 and 4. Subsequently IRAK1 is hyperphosphorylated and induces interaction with TNF-α receptor-associated factor 6 (TRAF6), resulting in TRAF6 oligomerization. After oligomerization of TRAF6, IRAK1-TRAF6 complex is dissociated from the receptor and associated with TAK1, which is mediated by TAB2 (or TAB3). In this process polyubiquitination of TRAF6 by Ubc13/Uev1A is thought to be critical for the association with TAB2 (or TAB3), which links TAK1 activation (46, 54, 55). In the case of TNF-α stimulation, TNF-α receptors form trimers and recruit adaptor proteins, TRAF2/5, and receptor-interacting protein 1 on the membrane. Ubc13/Uev1A- and TRAF2-dependent polyubiquitination of receptor-interacting protein 1 induce association of TAB2 (or TAB3), which then activates TAK1. Thus, TAB2 is required for ubiquitin-dependent activation of TAK1 by TRAFs. On the other hand, it has been demonstrated that hematopoietic progenitor kinase 1 plays a role as an upstream mediator of TGF-β-induced TAK1 activation, which in turn activates the MKK4-JNK signaling cascade in 293T cells (56, 57). Besides hematopoietic progenitor kinase 1, it has been also suggested that X-linked inhibitor of apoptosis (XIAP) might link TAK1 to TGF-β/BMP receptors through the capability of XIAP to interact with TGF-β/BMP receptors and TAB1 (58). Thus, although various molecules participate in the activation of TAK1, the precise mechanism by which TGF-β1 induces TAK1 activation is incompletely understood. Here, we provide evidence that the association of TAK1 with TGF-β receptors is important for TGF-β1-induced activation of TAK1 in mouse mesangial cells. TGF-β1 stimulation induces interaction of TβRI and TβRII, triggering dissociation of TAK1 from TβRI, and subsequently TAK1 is phosphorylated through TAB1-mediated autophosphorylation, independent of receptor kinase activity of TβRI.     

Heterogeneous Nuclear Ribonucleoprotein C Proteins Interact with the Human Papillomavirus Type 16 (HPV16) Early 3′-Untranslated Region and Alleviate Suppression of HPV16 Late L1 mRNA Splicing

In order to identify cellular factors that regulate human papillomavirus type 16 (HPV16) gene expression, cervical cancer cells permissive for HPV16 late gene expression were identified and characterized. These cells either contained a novel spliced variant of the L1 mRNAs that bypassed the suppressed HPV16 late, 5′-splice site SD3632; produced elevated levels of RNA-binding proteins SRSF1 (ASF/SF2), SRSF9 (SRp30c), and HuR that are known to regulate HPV16 late gene expression; or were shown by a gene expression array analysis to overexpress the RALYL RNA-binding protein of the heterogeneous nuclear ribonucleoprotein C (hnRNP C) family. Overexpression of RALYL or hnRNP C1 induced HPV16 late gene expression from HPV16 subgenomic plasmids and from episomal forms of the full-length HPV16 genome. This induction was dependent on the HPV16 early untranslated region. Binding of hnRNP C1 to the HPV16 early, untranslated region activated HPV16 late 5′-splice site SD3632 and resulted in production of HPV16 L1 mRNAs. Our results suggested that hnRNP C1 controls HPV16 late gene expression.     

Inhibition of Human Steroid 5��-Reductase (AKR1D1) by Finasteride and Structure of the Enzyme-Inhibitor Complex

The Δ⁴-3-ketosteroid functionality is present in nearly all steroid hormones apart from estrogens. The first step in functionalization of the A-ring is mediated in humans by steroid 5α- or 5β-reductase. Finasteride is a mechanism-based inactivator of 5α-reductase type 2 with subnanomolar affinity and is widely used as a therapeutic for the treatment of benign prostatic hyperplasia. It is also used for androgen deprivation in hormone-de pend ent prostate carcinoma, and it has been examined as a chemopreventive agent in prostate cancer. The effect of finasteride on steroid 5β-reductase (AKR1D1) has not been previously reported. We show that finasteride competitively inhibits AKR1D1 with low micromolar affinity but does not act as a mechanism-based inactivator. The structure of the AKR1D1·NADP⁺·finasteride complex determined at 1.7 Å resolution shows that it is not possible for NADPH to reduce the Δ^1-2-ene of finasteride because the cofactor and steroid are not proximal to each other. The C3-ketone of finasteride accepts hydrogen bonds from the catalytic residues Tyr-58 and Glu-120 in the active site of AKR1D1, providing an explanation for the competitive inhibition observed. This is the first reported structure of finasteride bound to an enzyme involved in steroid hormone metabolism.The Δ⁴-3-ketosteroid functionality is present in many important steroid hormones, e.g. testosterone, cortisone, and progesterone. An initial step in steroid hormone metabolism is the reduction of the Δ⁴-ene, which in humans is mediated by steroid 5α-reductases (SRD5A1, SRD5A2) or steroid 5β-reductase (AKR1D1)³ to yield the corresponding 5α- or 5β-dihydrosteroids, respectively (1, 2). The products of these reactions are not always inactive. 5α-Reductase is responsible for the conversion of testosterone to 5α-dihydrotestosterone (5α-DHT), which is the most potent natural ligand for the androgen receptor. By contrast, in addition to being involved in bile acid biosynthesis, 5β-reductase is responsible for generating 5β-pregnanes, which are natural ligands for the pregnane-X receptor (PXR) in the liver (3, 4). PXR is involved in the induction of CYP3A4, which is responsible for the metabolism of a large proportion of drugs (5, 6). Thus both 5α-reductase and 5β-reductase are involved in the formation of potent ligands for nuclear receptors.Finasteride is a selective 5α-reductase type 2 inhibitor that reduces plasma 5α-dihydrotestosterone levels and shrinks the size of the prostate (7). It is a widely used therapeutic agent in the treatment of benign prostatic hyperplasia (8, 9), it is used in androgen deprivation therapy to treat prostate cancer (10), and it has been examined as a chemopreventive agent for hormone-dependent prostate cancer (11). Finasteride was originally thought to act as a competitive inhibitor with nanomolar affinity for 5α-reductase type 2 (12). More recently, it was found that finasteride acts as a mechanism-based inactivator of this enzyme (13). Subsequent to inhibitor binding, there is hydride transfer from the NADPH cofactor to the Δ^1-2-ene double bond of finasteride. The intermediate enolate tautomerizes at the enzyme active site to form a bisubstrate analogue in which dihydrofinasteride is covalently bound to NADP⁺ (13). The bisubstrate analogue has subnanomolar affinity for 5α-reductase type 2 (Fig. 1). No structural information exists for 5α-reductase type 1 or type 2; therefore, it is not possible to determine how finasteride would bind to the active site of a human steroid double bond reductase in the absence of an experimentally determined crystal structure.Open in a separate windowFIGURE 1.Mechanism-based inactivation of 5α-reductase type 2 by finasteride. Adapted from Bull et al. (13). R = −C(=O)-NH₂; PADPR = 2′-phosphoadenosine-5″-diphosphoribose.Human steroid 5β-reductase is a member of the aldo-keto reductase (AKR) superfamily and is formally designated (AKR1D1) (14). The AKRs are soluble NADP(H)-dependent oxidoreductases with monomeric molecular masses of 37 kDa. These enzymes are amenable to x-ray crystallography, and during the last year, we and others have reported crystal structures of ternary complexes of AKR1D1 (15–17). The ternary complexes containing steroid substrates include: AKR1D1·NADP⁺·testosterone (PDB: 3BUR), AKR1D1·NADP⁺·progesterone (PDB: 3COT), AKR1D1·NADP⁺·cortisone (PDB: 3CMF), and AKR1D1·NADP⁺·Δ⁴-androstene-3,17-dione (PDB: 3CAS) (17). In addition, ternary complexes containing the products 5β-dihydroprogesterone (PDB: 3CAV) and 5β-dihydrotestosterone (PDB: 3DOP) have also been described (16, 18).As part of an ongoing inhibitor screen of AKR1D1, we now report that finasteride acts as a competitive inhibitor with low micromolar affinity. Additionally, we report the x-ray crystal structure of the AKR1D1·NADP⁺·finasteride complex.     

Existence of Sixteen 2′→5′ Dinucleotides in Nuclease P1 (Penicillium Nuclease) Digest of Technical Grade Yeast RNA

Sixteen 2′→5′ dinucleotides; (2′–5′)pA-A, pA-G, pA-C, pA-U, pG-A, pG-G, pG-C, pG-U, pC-A, pC-G, pC-C, pC-U, pU-A, pU-G, pU-C, and pU-U were detected in nuclease P₁ digest of a technical grade yeast RNA by means of gel filtration on Sephadex G-10, DEAE-Sephadex A-25 column chromatography in the presence of 7 m urea, paper electrophoresis and paper chromatography. Content of each dinucleotide was about 0.1 to 0.6% of the digest. As the sixteen 2′→5′ dinucleotides were found in all of the digests of technical grade RNA preparations tested, each polynucleotide chain in the preparations may be concluded to contain several per cent of the 2′–5′ minor phosphodiester linkages in addition to the 3′–5′ major phosphodiester linkages.     

Structure and Conformation of the Branch Core Triribonucleotide Containing 2′-5′ and 3′-5′ Phosphodiester Linkages (A2′p5′G 3′p5′C) in Solution,Essential for Yeast mRNA Splicing,Deduced from 1H-NMR

Abstract

The non-exchangeable ¹H-NMR signals of the branch core trinucleotide of the lariat branch site (A^2′p5′G _3′p5′C), 1) and its derivatives 2 and 3 are completely assigned using one- and two- dimensional NMR techniques including NOE, COSY, NOESY, ¹H-¹HINADEQUATE and 2D-J-resolved spectroscopy. From the vicinal coupling constants in the individual ribose rings, NOE data and T₁ measurements, the following properties of the trimers are deduced.(i)The unique stacking behavior of the trimers is S^1′N _3′N, and the sugar rings exist predominantly in the N-conformation (3′-endo-2′-exo).(ii)The sugar-base orientations appear to be anti.(iii) The branched trimers exist in solution as single-stranded right-handed conformations resembling A-RNA with stacking between the adenine and guanine residues in aqueous solution at 21°C and pH 7.2.(iv) The calculated values for the torsion angles ε^t andγ⁺ for the trimers are 201–203° and 71–86%, respectively, while the percent β¹ values are higher for the guanine (87–92%) than the cytosine residues (73–77%). The computer generated depiction of the triribonucleotide 1 is also shown. These subtle structural features may act as recognition signals for this critical lariat branch site which is essential for the second step in yeast mRNA splicing.     

Discovery of potent and selective GIRK1/2 modulators via ‘molecular switches’ within a series of 1-(3-cyclopropyl-1-phenyl-1H-pyrazol-5-yl)ureas

This Letter describes the on-going SAR efforts based on ML297, a potent, efficacious and selective GIRK1/2 activator (∼10-fold vs GIRK1/4 and inactive on GIRK2/3) via an iterative parallel synthesis approach. The chemical optimization at the 3-position of pyrazole within ML297 indicated that various functionalized 3-cyclopropyl moieties modulated GIRK pharmacology between inhibitor/activator within a series of 1-(3-cyclopropyl-1-phenyl-1H-pyrazol-5-yl)ureas. Importantly, novel ‘molecular switches’ that modulated the mode of pharmacology from inhibitor to activator was discovered on both the 3-cyclopropyl and N-phenyl moiety of the pyrazole core, providing the first highly selective GIRK1/2 activator.     

Pseudouridine residues in the 5′-terminus of uridine-rich nuclear RNA I (U1 RNA)

The primary nucleotide sequence was reported earlier for U1 RNA (Reddy et al, (1974) J. Biol. Chem. $\text{249}$, 6486–6494), an snRNA implicated in splicing of HnRNAs. In view of the presence of homologous pseudouridine (ψ) residues in 5′-ends of several highly conserved U-snRNAs and the recent report of modified bases in the U1 RNA structure (Branlant et al, (1980) Nucleic Acids Res. $\text{8}$, 4143–4154) a study was made for the presence of ψ and other modified nucleotides in the 5′-end of the U1 RNA. Identification of ψ residues at positions 6 and 7, shows the 5′-sequence of U1 RNA is: m₃², 2,7 GpppAm-Um-A-C-ψ-ψ-A-C-C-U-G-G-C-A-G-G-G-G-A-G-A-U-A-C. The ψ residues in place of U at positions 6 and 7 may affect the binding of U1 RNA at intron-exon splice junctions.     

Protein arginine methyltransferase 5 (PRMT5) activates WNT/β-catenin signalling in breast cancer cells via epigenetic silencing of DKK1 and DKK3

Protein arginine methyltransferase 5 (PRMT5) activity is dysregulated in many aggressive cancers and its enhanced levels are associated with increased tumour growth and survival. However, the role of PRMT5 in breast cancer remains underexplored. In this study, we show that PRMT5 is overexpressed in breast cancer cell lines, and that it promotes WNT/β-CATENIN proliferative signalling through epigenetic silencing of pathway antagonists, DKK1 and DKK3, leading to enhanced expression of c-MYC, CYCLIN D1 and SURVIVIN. Through chromatin immunoprecipitation (ChIP) studies, we found that PRMT5 binds to the promoter region of WNT antagonists, DKK1 and DKK3, and induces symmetric methylation of H3R8 and H4R3 histones. Our findings also show that PRMT5 inhibition using a specific small molecule inhibitor, compound 5 (CMP5), reduces PRMT5 recruitment as well as methylation of H3R8 and H4R3 histones in the promoter regions of DKK1 and DKK3, which consequently results in reduced expression CYCLIN D1 and SURVIVIN. Furthermore, CMP5 treatment either alone or in combination with 5-Azacytidine and Trichostatin A restored expression of DKK1 and DKK3 in TNBCs. PRMT5 inhibition also altered the growth characteristics of breast cancer cells and induced their death. Collectively, these results show that PRMT5 controls breast cancer cell growth through epigenetic silencing of WNT/β-CATENIN pathway antagonists, DKK1 and DKK3, resulting in up-regulation of WNT/β-CATENIN proliferative signalling.     

Linkage of acid α-glucosidase (Gaa) and Thymidine kinase (Tk-1) to Esterase-3 (Es-3) on mouse Chromosome 11

Inheritance in recombinant inbred (RI) strains of restriction fragment length variants (RFLVs) detected by probes specific for Gaa and Tk-1 showed tight linkage of both to Es-3 on mouse Chromosome (Chr) 11. This result extends the region of homology between mouse Chr 11 and human chr 17q.