bluuebunny/crossref_metadata_2025_split huggingface dataset:
Defective Activation of Phospholipase Cγ2 by Collagen in Platelets Lacking the Semaphorin Family Member, Sema4D.
Authors: Zhu Li , Jiang Hong , Kumanogoh Atsushi , Kikutani Hitoshi , Brass Lawrence F. 
Published: October 2019

Abstract
Semaphorins are a large family of cell surface molecules best known for their ability to mediate communication between cells during neural development. We have recently shown that human platelets express the semaphorin family member, sema4D, and both of its known receptors, CD72 and plexin-B1 (Zhu, et al, PNAS, 2007). We have also shown that sema4D(−/−) mice have an impaired response to arterial injury in vivo and a selective defect in collagen- and convulxin-induced platelet aggregation in vitro. In the present studies we have sought the molecular basis for these defects, focusing on events downstream of glycoprotein VI (GPVI), which serves as a receptor for both collagen and convulxin. In normal platelets, GPVI signaling leads to the phosphorylation and activation of phospholipase Cγ2 (PLCγ2) through the formation of a signaling complex that includes SLP-76 and LAT. This complex is activated when GPVI-associated FcRγ is phosphorylated, allowing the tyrosine kinase, Syk, to bind. PLCγ2 activation results in phosphoinositide hydrolysis, an IP3-mediated increase in cytosolic Ca++, and activation of additional kinases, such as Akt. In theory, the absence of sema4D could affect any of these steps and by doing so impair collagen-induced platelet aggregation. Working backwards through the GPVI pathway, our results showed that compared to platelets from matched WT mice, sema4D(−/−) platelets have 1) a rightward-shift in the dose/response curve for collagen-induced Akt phosphorylation, 2) a 37% smaller increase in cytosolic Ca++, and 3) a 43% smaller increase in PLCγ2 phosphorylation. However, we found no defect in collagen-induced FcRγ phosphorylation, which is the earliest event in GPVI signaling. The defect in PLCγ2 phosphorylation was not limited to mouse platelets, but was also observed when human platelets were stimulated with collagen in the presence of an antibody directed towards the sema4D extracellular domain. Taken together, these results show that sema4D is needed for optimal activation of PLCγ2 by collagen downstream of the GPVI/FcRγ complex. Sema4D is believed to act in part through contact-dependent binding of sema4D to its receptors, CD72 and plexin-B1. Since these studies were performed under conditions in which platelet:platelet contacts can occur, the observed defect in collagen and convulxin responses could be due to impaired signaling by either of these receptors or, in theory, by retrograde signaling via sema4D. One candidate mechanism involves a regulatory complex between CD72 and the tyrosine phosphatase, SHP-1, which we have shown to occur in resting human platelets and to be lost when platelets are activated by agonists or stimulated by soluble sema4D. In theory, sema4D-dependent loss of the CD72/SHP-1 complex allows SHP-1 to relax into an inactive conformation, promoting protein tyrosine phosphorylation, which would not occur when sema4D is absent or blocked.
Stat5a Is Epigenetically Silenced by Oncogenic Tyrosine Kinase NPM/ALK and Acts as Tumor Suppressor by Reciprocally Inhibiting Expression of NPM/ALK.
Authors: Zhang Qian , Wang Hong Yi , Liu XiaoBin , Wasik Mariusz A. 
Published: October 2019

Abstract
Although Stat5a and Stat5b exhibit some non-redundant functional properties, their distinct contributions to carcinogenesis are not clearly defined. Here we report that Stat5a expression is selectively inhibited by DNA methylation of the Stat5a gene promoter region in cells expressing an oncogenic tyrosine kinase NPM/ALK. The DNA methylation is induced by NPM/ALK itself by employing its key effector Stat3, and is associated with binding to the promoter of MeCP2 capping protein and lack of binding of the Stat5a transcription activator Sp-1. Reversal of the methylation by DNA methyltransferase inhibitor, 5′-aza-2-deoxycytidine, restores the Sp-1 binding and Stat5a expression. Strikingly, the induced or exogenously expressed Stat5a binds to enhancer and exon 14 of the NPM/ALK gene and triggers selective suppression of NPM/ALK expression. These results demonstrate that NPM/ALK induces epigenetic silencing of Stat5a and that Stat5a can act as key tumor suppressor by reciprocally inhibiting expression of NPM/ALK.
A Study of Multiple Drug Resistance Mechanisms Improved Against Bortezomib on Multiple Myeloma Cell Lines In Vitro.
Authors: Uyuklu Tolga , Ural A. Ugur , Sarper Meral , Avcu Ferit , Baran Yusuf , Elci Pinar , Akar Nejat 
Published: October 2019

Abstract
The most important problem in the treatment of Multiple Myeloma (MM) is the multi drug resistance (MDR) observed before and after the treatment. For this reason in MM cases an early resistance to treatment can be developed or the disease can relapsed in early period. Yet, there has been no improved drug resistance against proteazom inhibitor Bortezomib (Bor), which is used alone or with other chemotherapeutic agents in resistant or relapsed MM cases. In this study, bortezomib resistant human MM cell lines; RPMI-8226, secreting lambda light chain, and ARH-77, secreting IgG, were developed and responsible resistance mechanisms were investigated. For this purpose, by exposing to the cells to sequentially gradual doses of Bor in vitro conditions, resistant cell lines were acquired throughout one year. The IC50 values for Bor were determined after 48 hour incubation by MTT cytotoxicity assay (IC50:1,16nM for RPMI-8226 and IC80:0,6nM for ARH-77) against wild type cells. Throughout one year some cell lines resistant to 1,3nM Bor were acquired by performing Bor to both cell lines in gradual doses. In resistant cell lines IC50:18,07 for RPMI-8226 and IC50:97,56 nM for ARH-77 were determined by MTT assay. In parallel of the gradual increase in drug concentration; the expression changes of the genes of ATP binding cassette protein; MDR1 (Multi Drug Resistance Protein), MRP1 (Multi Drug Resistance Associated Protein), BCRP (Breast Cancer Resistance Protein); and LRP (Lung Resistant Protein) which is responsible for accumulation of the drug in cytoplasm with the aid of nuclear membrane were determined with Reverse Transcriptase Polymerase Chain Reaction (RT-PCR) and densitometric analysis. In resistant cells, high expression of MDR1, MRP1, BRCP and LRP genes showed that; pumping the drug out of the cell membrane and decrease in accumulation of the drug in the cytoplasm had effects on the resistant mechanisms against Bor. Furthermore, expression changes of an important sing of apoptosis ‘caspase-3’, pro-apoptotic ‘bax’ and an anti-apoptotic ‘bcl-2’ genes were examined by RT-PCR and we could come to a point that when compared the sensitive cells to resistant cells, expression of caspase-3 gene and pro-apoptotic bax protein decreased but bcl-2 gene expression increased in resistant cell lines. Finally, we concluded that resistant cell lines acquired resistance against apoptosis by means of mitochondria. By means of this project, the genes which are responsible for secondary drug resistance in ARH-77 and RPMI-8226 MM cell lines in vitro conditions against Bor were determined. Also resistance mechanisms against apoptosis were demonstrated. Cross resistance to different chemotherapeutic agents mechanisms are still continuing.
Identification of LIV1, a Zinc Transporter Gene, Responsible for HDACI-Induced Apoptosis, Using a Genetic Approach.
Authors: Zhou Jianfeng , Wang Beibei , Chen Pingbo , Wang Jintao , Zhang Yicheng , Ma Ding , Ma Xiaoli 
Published: October 2019

Abstract
Histone deacetylase inhibitors (HDACI) show promise as chemotherapeutic agents and have been demonstrated to induce apoptosis of tumor cells. To gain a better understanding of the molecular basis for HDACI-induced apoptosis and directly identify the gene products which are essential for mediating the apoptosis-regulatory functions of HDACI, we employed the suppression of mortality by antisense rescue technique (SMART). This approach permits the isolation of apoptosis-associated genes based on their selective inactivation by overexpression of antisense cDNAs. Because the antisense mRNA inactivates gene expression of apoptosis-specific genes, transfected cells survive in the presence of apoptosis inducers. We isolated several genes associated with HDACI-induced mortality (GHIM). Here we characterized one of the GHIM cDNAs, GHIM-27. Sequence analysis suggests that the GHIM-27 product is identical to human zinc transporter LIV1. Further researches indicate that LIV1 is significantly induced by HDACI treatment in various cancer cell lines. Down-regulation of LIV1 confers a strong resistance against HDACI-induced apoptosis, which is caused at least in part by disregulation of intracellular zinc homeostasis. Together, these studies identify a zinc transporter gene LIV1 which is responsible for HDACI-induced apoptosis by using a genetic approach. This may provide a potentially important target for anticancer therapy.
Exogenous Nucleic Acids - A Potential Source of Resistance to Nucleoside Analogues in Cancer and Antiviral Therapy.
Authors: Buhmann Raymund , Yang Ting , Obermeier Martin , Jaeger Gundula , Kolb Hans-Jochem 
Published: October 2019

Abstract
The metabolism of exogenous nucleic acids is not well defined. Moreover, there is little information whether there might be interference with chemically and structurally related drugs, e.g. nucleoside analogues (NA) widely used in cancer or in antiviral therapy. In the present report we provide evidence, that nucleic-acid based drugs might antagonize fludarabine or acyclovir. In vitro, fludarabine treated lymphocytes or myeloid blasts where rescued from apoptosis when incubated with defibrotide (DF), a polydisperse mixture of single-stranded oligodeoxyribonucleotides (15 to 30 kD) or singular deoxynucleotides. Thereby deoxycytidine (dCTP) turned out to be the key substrate competing with fludarabine for phosphorylation by deoxycitidine kinase (dCK) and suggested interference with nucleic acid metabolism rather than direct competition with the drug. To further prove this hypothesis the influence of defibrotide on HSV replication was evaluated. In standard drug resistance assays performed with acyclovir sensitive herpes simplex virus (HSV) strains (V0631508) 4 mM of DF restored viral replication in presence of 50 mM acyclovir. This was confirmed by quantitative PCR of viral DNA. Moreover, in this case deoxythymidine (dTTP) turned out to be the competitor for intracellular phosphorylation mediated by virus thymidine kinase. We conclude that treatment with DF and other nucleic-acid-based drugs interfere with the efficacy of NA used for cancer and antiviral therapy. Prospective clinical trials are required to confirm these in vitro findings.
Mutations of the Genes for Ribosomal Proteins L5 and L11 Are a Common Cause of Diamond-Blackfan Anemia.
Authors: Gazda Hanna T. , Sheen Mee Rie , Darras Natasha , Shneider Hal , Sieff Colin A. , Ball Sarah E. , Niewiadomska Edyta , Newburger Peter E. , Atsidaftos Eva , Vlachos Adrianna , Lipton Jeffrey M. , Beggs Alan H. 
Published: October 2019

Abstract
Diamond-Blackfan anemia (DBA), a form of congenital red cell aplasia with marked clinical heterogeneity and increased risk of malignancy, has been associated with mutations in ribosomal protein (RP) gene RPS19 in 25% of probands and in RPS24 or RPS17 in ∼2% of patients. Thus, DBA appears to be a disorder of ribosome synthesis. To test the hypothesis that mutations in other RP genes may also cause DBA, we carried out direct sequencing of candidate RP genes. Genomic DNA samples from 96 unrelated DBA probands (14 familial and 82 sporadic cases) without RPS19 or RPS24 mutations were screened for mutations in RPS3a, RPS13, and RPS16 (previous studies revealed that RPs S19, S24, S3a, S13, and S16 are involved in binding of eIF-2 to the 40S subunit); RP genes L18, L13A, L36, L28, L18A, L40, S5, S9, S11, and S28 (located on chromosome 19); and RP genes, L5, L11, L22, S8, and S27 (on chromosome 1). PCR primers were designed to amplify the coding exons and intron/exon boundaries. We found multiple mutations in two RP genes, L5 and L11. Subsequently we sequenced these two genes in 42 additional DNA samples from DBA probands. In total, we screened 5′UTR, promoter and coding regions, and exon/intron boundaries of RPL5 and RPL11 in 138 DBA unrelated probands. We identified 14 mutations in RPL5 in 138 probands (∼10%), 13 of which are nonsense mutations, deletions or insertions of 1–5 nucleotides causing frameshift and premature termination. One missense mutation, 418G>A, results in a G140S substitution. We found nine mutations in RPL11 in138 DBA probands (6.5%), including five acceptor or donor splice site mutations (introns 1–4) and four deletions or insertions of 1–4 nucleotides causing frameshifts (codons 32-120). None of these sequence changes were found on the NCBI (http://www.ncbi.nlm.nih.gov/SNP/) or the HapMap (http://www.hapmap.org/) SNP lists. Both genes, as well as RPL23 have recently been demonstrated by others to activate the p53 tumor suppressor protein by inhibiting MDM2-mediated p53 ubiquitination and degradation. Moreover, knockdown of any of these genes by siRNAs markedly reduced p53 induction by the ribosomal biogenesis stressor, actinomycin-D. These findings suggest that DBA patients with mutated L5 and L11 proteins may have inadequate p53 pathway activation and (consistent with clinical observations) be at increased risk for neoplasia. We are currently investigating the role of RPL5 and RPL11 mutations in ribosomal biogenesis and in the p53-mediated cell cycle arrest and apoptosis in DBA patients.
Novel Heritable Mutation of the Transcription Factor RUNX1 as a Cause of Autosomal Dominant Familial Platelet Disorder with Predisposition to Acute Myeloid Leukemia (FPD/AML).
Authors: Patton William N. , Suthers Graeme , Altree Meryl , Carmichael Catherine , Wilkins Ella , Carroll Jacqueline , Rawlings Lesley , Scott Hamish 
Published: October 2019

Abstract
Aim To identify the causative heritable mutation in a family with autosomal dominant familial platelet disorder with predisposition to acute myeloid leukemia (FPD/AML).
Method Confirmation of family pedigree, enrolment into ethics committee approved Australian Familial Hematological Cancer Study, procurement of genomic DNA from pedigree members and genetic analysis by sequencing of RUNX1 and CEBPA genes.
Results The proband intially presented aged 50 with mild thrombocytopenia initially diagnosed as idiopathic thrombocytopenic purpura when bone marrow examination (including cytogenetics analysis) was normal. Three years later she was referred with severe progressive thrombocytopenia unresponsive to high dose steroids and intravenous immunoglobulin together with mild anemia and neutropenia. Marrow examination revealed subtle dysplasia with monosomy 7 in 12/20 metaphases. A diagnosis of myelodysplastic syndrome (MDS) was made. Three months later (Feb 2007), she progressed to acute myeloid leukemia (AML). The proband’s mother had had mild thrombocytopenia with subsequent MDS (aged 70) and died of AML two years later. The proband’s only sibling and nephew have mild thrombocytopenia without features of MDS. The proband entered cytogenetic remission following one course of AML induction therapy but continued to show dysplastic features. She then received 2 cycles of consolidation therapy and has undergone unrelated donor allogeneic stem cell transplant (July 2007). Sequencing of PCR products from exons of the RUNX1 gene identified a novel heterozygous mutation in the proband’s constitutional DNA: c.958C>T in exon 7, in the transactivation domain, causing a nonsense mutation, p.Arg293X (sequence variation for RUNX1 classified according to GenBank Accession No. NC_000021). The sibling has the same mutation and studies in other relatives are underway.
Conclusion This study has identified a novel RUNX1 mutation responsible for FPD/AML in this family. Clinicians should be aware of this rare inherited disorder and the availability of genetic testing. People with mutations may be asymptomatic and genetic testing in such families is essential before considering bone marrow transplantation from a living related donor.
FBW7 E3-Ligase Mutations Are Associated with T Cell Leukemia.
Authors: Buonamici Silvia , Thompson Benjamin J. , Sulis Maria L. , Palomero Teresa , Ferrando Adolfo , Aifantis Iannis 
Published: October 2019

Abstract
Recent studies have shown that activating mutations of NOTCH1 are responsible for the majority of T cell acute lymphoblastic leukemia (T-ALL) cases. Most of these mutations truncate its C-terminal domain, a region that is important for the NOTCH1 proteasome-mediated degradation. We report that the E3 ligase FBW7 targets NOTCH1 for ubiquitination and degradation. Our studies map in detail the amino acid degron sequence required for NOTCH1-FBW7 interaction. Furthermore, we identify inactivating FBW7 mutations in a large fraction of human T-ALL lines and primary leukemias. These mutations abrogate the binding of FBW7 not only to NOTCH1 but also to the two other characterized targets, c-Myc and cyclin E. The majority of the FBW7 mutations were present during relapse, and they were associated with NOTCH1 HD mutations. Interestingly, most of the T-ALL lines harboring FBW7 mutations were resistant to γ-secretase inhibitor treatment and this resistance appeared to be related to the stabilization of the c-Myc protein. Our data suggest that FBW7 is a novel tumor suppressor in T cell leukemia, and implicate the loss of FBW7 function as a potential mechanism of drug resistance in T-ALL.
Treatment with Valproic Acid Restores Differentiation in AML1/ETO-Positive Leukaemic Cells.
Authors: Zapotocky Michal , Starkova Julia , Mejstrikova Ester , Smetana Karel , Trka Jan 
Published: October 2019

Abstract
In t (8, 21) acute myeloid leukaemia (AML), the leukaemogenesis is supposed to be promoted by interference with expression of AML1 target genes. Repressor complex associated with AML1/ETO fusion protein recruits class I histone deacetylases (HDAC). Valproic acid (VPA) was found to have an extensive effect on AML blasts, via inhibition of class I HDAC. It was shown previously that VPA treatment disrupts the AML1/ETO-HDAC1 complex from AML1 promoter thus leading to apoptosis at different cell lines. However, there is still lack of in-depth morphological and immunophenotypical proof of the hypothesized restoration of differentiation after treatment with VPA. We aimed to characterize the differentiation effect of VPA on AML1/ETO-positive leukaemic cells. Kasumi-1 (AML1/ETO-positive) cell line and MV4-11 (MLL/AF4-positive) cells were treated with VPA and Trichostatin-A (TSA) another HDAC inhibitor. The effect of VPA on cell cycle and differentiation was examined by flowcytometry. DNA, RNA and AgNORs staining of nucleus and nucleolus were used for morphological evaluation. VPA treatment of Kasumi-1 cells resulted in decreased expression of early myeloid progenitor antigens [CD33 (p<0.001), CD34 (p=0.003), CD117 (p<0.001)] and increased expression of antigens typical for more differentiated myeloid cells [CD11a (p=0.001), CD11b (p=0.007)]. Morphological evaluation revealed nucleus fragmentation without chromatin condensation, which evoked monocytoid look of the nucleus in VPA-treated Kasumi cells. The nucleolar morphology and cytochemistry indicated that these cells entered the ageing process and decreased proliferation activity. Large nucleoli characterised by a reduced number of silver-binding nucleolar organizer regions (AgNORs) as well as by their translocation to the nucleolar periphery represented a base for such interpretation. VPA-treated Kasumi cells exhibited accumulation in G1/G0 phase and decreased proliferation (VPA 0.5mM - G1/G0 70%, S/G2/M 16%; control cells - G1/G0 62%, S/G2/M 28%). Conversely, MV4-11 cell line (representing myeloid leukaemic cells of non-CBF origin) did not show any statistically significant differences in imunophenotype after VPA treatment (CD15, CD11b, CD117, CD45, HLA DR, CD14). VPA treated MV4-11 cells did not show accumulation in G1/G0 phase (VPA 0.5mM G1/G0 63%, S/G2/M 13%; control G1/G0 70%, S/G2/M 22%). We show that treatment with VPA restores differentiation of AML1/ETO positive cells as presented by both changes in immunophenotype and morphological analysis. This observed differentiation correlates with decrease in cell cycle. We demonstrate that the effect on differentiation is specific for AML1/ETO-positive cells as it is not noticeable in leukaemic cells of non-CBF origin.
Identification of Differentially Expressed Proteins from the Mitochondria Enriched Cytoplasmic Fractions in Primary AML Cells.
Authors: Shin Myung-Geun , Kim Hye-Ran , Kim Hyeoung-Joon , Lee Il-Kwon , Kim Mi-Ji , Shin Jong-Hee , Suh Soon-Pal , Ryang Dong-Wook 
Published: October 2019

Abstract
Mitochondrial dysfunction has been associated with chronic degenerative diseases, aging and cancer. Mitochondrial proteins are interesting targets for the study of apoptosis in leukemia and other malignancies. Modern mass-spectrometry instrumentation has achieved excellent performance in terms of sensitivity, resolution and mass accuracy; however, so far, the contribution of proteomics to the care of patients with AML is virtually zero. Therefore, we set up strategies for the proteomic analysis of the mitochondria enriched cytoplasmic fractions in primary AML cells. Primary AML cells and normal blood cells were derived from bone marrow aspirates of a group of five patients with AML (M1 and M2) at the time of initial diagnosis before chemotherapy and three healthy controls. Peptide mass spectra were obtained on a MALDI-TOF-TOF mass spectrometer. Protein identification was processed and analyzed by searching the Swiss-Prot protein database using the MASCOT search engine of Matrix Science that integrated in the Global ProteinServer Workstation. Tandem electrospray ionization mass spectrometry (ESI-MS/MS) experiments were performed on a quadrupole (Q)-time of flight 2 (Q-TOF2) hybrid quadrupole/TOF mass spectrometry. We identified 48 spots corresponding to 38 proteins in primary AML cells, the expressions of which were altered significantly compared with normal blood cells. Out of these deregulated proteins, totally 12 and 20 proteins were observed in up- or down-regulated spots, respectively. Interestingly, prohibitin (gi4505773) was highly expressed in primary AML cells. Prohibitin is known to induce growth suppression and repress E2F-mediated transcription. But it has recently found that prohibitin protects cells from apoptosis by down-regulating E2F activity when Rb family members are inactive (Oncogene2002;21:4539–48). The current proteomic analysis for primary AML cells supports that prohibitin not only acts as a repressor of E2F activity during normal cellular conditions, but may also aid in the decision between survival and death during times of apoptotic stimulation. Our ongoing studies will try to elucidate the precise role and mechanism in primary AML cells. Six proteins (ADP-ribosylation factor guanine nucleotide-exchange factor 2, cytokeratin-9, vimentin, chain A, kinesin family member 1C and gi34533624 unnamed protein product) were exclusively identified in primary cells. In conclusion, this mitochondrial proteomic analyses enable us to define the deregulated mitochondria-related proteins important for the initiation and the progression of AML. Also, these analyses may promote the identification of new targets for specific treatment approaches in AML.