Role for the Related Poly(ADP-Ribose) Polymerases Tankyrase 1 and 2 at Human Telomeres

ABSTRACT Telomere maintenance is essential for the continuous growth of tumor cells. In most human tumors telomeres are maintained by telomerase, a specialized reverse transcriptase. Tankyrase 1, a human telomeric poly(ADP-ribose) polymerase (PARP), positively regulates telomere length through its interaction with TRF1, a telomeric DNA-binding protein. Tankyrase 1 ADP-ribosylates TRF1, inhibiting its binding to telomeric DNA. Overexpression of tankyrase 1 in the nucleus promotes telomere elongation, suggesting that tankyrase 1 regulates access of telomerase to the telomeric complex. The recent identification of a closely related homolog of tankyrase 1, tankyrase 2, opens the possibility for a second PARP at telomeres. We therefore sought to establish the role of tankyrase 1 at telomeres and to determine if tankyrase 2 might have a telomeric function. We show that endogenous tankyrase 1 is a component of the human telomeric complex. We demonstrate that telomere elongation by tankyrase 1 requires the catalytic activity of the PARP domain and does not occur in telomerase-negative primary human cells. To investigate a potential role for tankyrase 2 at telomeres, recombinant tankyrase 2 was subjected to an in vitro PARP assay. Tankyrase 2 poly(ADP-ribosyl)ated itself and TRF1. Overexpression of tankyrase 2 in the nucleus released endogenous TRF1 from telomeres. These findings establish tankyrase 2 as a bona fide PARP, with itself and TRF1 as acceptors of ADP-ribosylation, and suggest the possibility of a role for tankyrase 2 at telomeres.

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Rescue of an hTERT Mutant Defective in Telomere Elongation by Fusion with hPot1

Molecular and Cellular Biology ◽

10.1128/mcb.24.8.3552-3561.2004 ◽

2004 ◽

Vol 24 (8) ◽

pp. 3552-3561 ◽

Cited By ~ 55

Author(s):

Blaine N. Armbruster ◽

Corinne M. Linardic ◽

Tim Veldman ◽

Niharika P. Bansal ◽

Diane L. Downie ◽

...

Keyword(s):

Binding Proteins ◽

Telomeric Dna ◽

Telomere Elongation ◽

Telomere Binding Protein ◽

Catalytically Active ◽

Bona Fide ◽

Per Se ◽

Telomeric Protein

ABSTRACT The protein hPot1 shares homology with telomere-binding proteins in lower eukaryotes and associates with single-stranded telomeric DNA in vitro as well as colocalizing with telomere-binding proteins in vivo. We now show that hPot1 is coimmunoprecipitated with telomeric DNA and that stable expression of this protein in telomerase-positive cells results in telomere elongation, supporting the idea that hPot1 is a bona fide mammalian telomere-binding protein. We previously found that mutations in the N-terminal DAT domain of the hTERT catalytic subunit of telomerase rendered the enzyme catalytically active but unable to elongate telomeres in vivo. This phenotype could be partially rescued by fusion with the double-stranded telomeric protein hTRF2. Given that hPot1 binds to single-stranded DNA in vitro (at the same site that hTERT binds to in vivo), we addressed whether fusion of hPot1 can rescue the DAT mutations more efficiently than that of hTRF2. We now report that a DAT mutant of hTERT is indeed efficiently rescued upon fusion to hPot1. However, this rescue depended on the ability of hPot1 to localize to telomeres rather than binding to DNA per se. These data support a model whereby the DAT domain of hTERT is implicated in telomere-telomerase associations.

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Specificity of the HIV-1 Protease on Substrates Representing the Cleavage Site in the Proximal Zinc-Finger of HIV-1 Nucleocapsid Protein

Viruses ◽

10.3390/v13061092 ◽

2021 ◽

Vol 13 (6) ◽

pp. 1092

Author(s):

János András Mótyán ◽

Márió Miczi ◽

Stephen Oroszlan ◽

József Tőzsér

Keyword(s):

Amino Acid ◽

Zinc Finger ◽

Cleavage Site ◽

Potential Role ◽

Binding Mode ◽

Interaction Energies ◽

Vitro Assay ◽

Hiv 1

To explore the sequence context-dependent nature of the human immunodeficiency virus type 1 (HIV-1) protease’s specificity and to provide a rationale for viral mutagenesis to study the potential role of the nucleocapsid (NC) processing in HIV-1 replication, synthetic oligopeptide substrates representing the wild-type and modified versions of the proximal cleavage site of HIV-1 NC were assayed as substrates of the HIV-1 protease (PR). The S1′ substrate binding site of HIV-1 PR was studied by an in vitro assay using KIVKCF↓NCGK decapeptides having amino acid substitutions of N17 residue of the cleavage site of the first zinc-finger domain, and in silico calculations were also performed to investigate amino acid preferences of S1′ site. Second site substitutions have also been designed to produce “revertant” substrates and convert a non-hydrolysable sequence (having glycine in place of N17) to a substrate. The specificity constants obtained for peptides containing non-charged P1′ substitutions correlated well with the residue volume, while the correlation with the calculated interaction energies showed the importance of hydrophobicity: interaction energies with polar residues were related to substantially lower specificity constants. Cleavable “revertants” showed one residue shift of cleavage position due to an alternative productive binding mode, and surprisingly, a double cleavage of a substrate was also observed. The results revealed the importance of alternative binding possibilities of substrates into the HIV-1 PR. The introduction of the “revertant” mutations into infectious virus clones may provide further insights into the potential role of NC processing in the early phase of the viral life-cycle.

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Telomerase RNA Template Mutations Reveal Sequence-Specific Requirements for the Activation and Repression of Telomerase Action at Telomeres

Molecular and Cellular Biology ◽

10.1128/mcb.20.8.2941-2948.2000 ◽

2000 ◽

Vol 20 (8) ◽

pp. 2941-2948 ◽

Cited By ~ 33

Author(s):

John C. Prescott ◽

Elizabeth H. Blackburn

Keyword(s):

Enzymatic Activity ◽

Activity Levels ◽

Telomeric Dna ◽

Telomere Shortening ◽

Telomere Elongation ◽

Telomere Binding Protein ◽

Telomere Function ◽

Double Base ◽

Telomere Lengthening

ABSTRACT Telomeric DNA is maintained within a length range characteristic of an organism or cell type. Significant deviations outside this range are associated with altered telomere function. The yeast telomere-binding protein Rap1p negatively regulates telomere length. Telomere elongation is responsive to both the number of Rap1p molecules bound to a telomere and the Rap1p-centered DNA-protein complex at the extreme telomeric end. Previously, we showed that a specific trinucleotide substitution in the Saccharomyces cerevisiae telomerase gene (TLC1) RNA template abolished the enzymatic activity of telomerase, causing the same cell senescence and telomere shortening phenotypes as a complete tlc1 deletion. Here we analyze effects of six single- and double-base changes within these same three positions. All six mutant telomerases had in vitro enzymatic activity levels similar to the wild-type levels. The base changes predicted from the mutations all disrupted Rap1p binding in vitro to the corresponding duplex DNAs. However, they caused two classes of effects on telomere homeostasis: (i) rapid, RAD52-independent telomere lengthening and poor length regulation, whose severity correlated with the decrease in in vitro Rap1p binding affinity (this is consistent with loss of negative regulation of telomerase action at these telomeres; and (ii) telomere shortening that, depending on the template mutation, either established a new short telomere set length with normal cell growth or was progressive and led to cellular senescence. Hence, disrupting Rap1p binding at the telomeric terminus is not sufficient to deregulate telomere elongation. This provides further evidence that both positive and negativecis-acting regulators of telomerase act at telomeres.

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Interaction between Poly(ADP-ribose) and NuMA Contributes to Mitotic Spindle Pole Assembly

Molecular Biology of the Cell ◽

10.1091/mbc.e09-06-0477 ◽

2009 ◽

Vol 20 (21) ◽

pp. 4575-4585 ◽

Cited By ~ 58

Author(s):

Paul Chang ◽

Margaret Coughlin ◽

Timothy J. Mitchison

Keyword(s):

Binding Proteins ◽

Magnetic Beads ◽

Spindle Pole ◽

Covalent Modification ◽

Mitotic Spindles ◽

Spindle Poles ◽

Tankyrase 1 ◽

Mitotic Spindle Pole

Poly(ADP-ribose) (pADPr), made by PARP-5a/tankyrase-1, localizes to the poles of mitotic spindles and is required for bipolar spindle assembly, but its molecular function in the spindle is poorly understood. To investigate this, we localized pADPr at spindle poles by immuno-EM. We then developed a concentrated mitotic lysate system from HeLa cells to probe spindle pole assembly in vitro. Microtubule asters assembled in response to centrosomes and Ran-GTP in this system. Magnetic beads coated with pADPr, extended from PARP-5a, also triggered aster assembly, suggesting a functional role of the pADPr in spindle pole assembly. We found that PARP-5a is much more active in mitosis than interphase. We used mitotic PARP-5a, self-modified with pADPr chains, to capture mitosis-specific pADPr-binding proteins. Candidate binding proteins included the spindle pole protein NuMA previously shown to bind to PARP-5a directly. The rod domain of NuMA, expressed in bacteria, bound directly to pADPr. We propose that pADPr provides a dynamic cross-linking function at spindle poles by extending from covalent modification sites on PARP-5a and NuMA and binding noncovalently to NuMA and that this function helps promote assembly of exactly two poles.

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High Expression of GSDMC Is Associated with Poor Survival in Kidney Clear Cell Cancer

BioMed Research International ◽

10.1155/2021/5282894 ◽

2021 ◽

Vol 2021 ◽

pp. 1-10

Author(s):

Yun-Qian Cui ◽

Fei Meng ◽

Wen-Li Zhan ◽

Zhou-Tong Dai ◽

Xinghua Liao

Keyword(s):

Potential Role ◽

Clear Cell ◽

Cell Cancer ◽

Progression Free Survival ◽

Renal Clear Cell Carcinoma ◽

High Expression ◽

Poor Survival ◽

Free Survival

This study is aimed at exploring the potential role of GSDMC in kidney renal clear cell carcinoma (KIRC). We analyzed the expression of GSDMC in 33 types of cancers in TCGA database. The results showed that the expression of GSDMC was upregulated in most cancers. We found a significant association between high expression of GSDMC and shortened patient overall survival, progression-free survival, and disease-specific survival. In vitro experiments have shown that the expression of GSDMC was significantly elevated in KIRC cell lines. Moreover, decreased expression of GSDMC was significantly associated with decreased cell proliferation. In summary, we believe that this study provides valuable data supporting future clinical treatment.

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Ozone Therapy as Adjuvant for Cancer Treatment: Is Further Research Warranted?

Evidence-based Complementary and Alternative Medicine ◽

10.1155/2018/7931849 ◽

2018 ◽

Vol 2018 ◽

pp. 1-11 ◽

Cited By ~ 11

Author(s):

Bernardino Clavo ◽

Norberto Santana-Rodríguez ◽

Pedro Llontop ◽

Dominga Gutiérrez ◽

Gerardo Suárez ◽

...

Keyword(s):

Cancer Treatment ◽

Immune Modulation ◽

Animal Studies ◽

Potential Role ◽

Randomized Clinical Trials ◽

Tumor Hypoxia ◽

Tumor Resection ◽

Ozone Therapy

Introduction. This article provides an overview of the potential use of ozone as an adjuvant during cancer treatment.Methods. We summarize the findings of the most relevant publications focused on this goal, and we include our related clinical experience.Results. Over several decades, prestigious journals have publishedin vitrostudies on the capacity of ozone to induce direct damage on tumor cells and, as well, to enhance the effects of radiotherapy and chemotherapy. Indirect effects have been demonstrated in animal models: immune modulation by ozone alone and sensitizing effect of radiotherapy by concurrent ozone administration. The effects of ozone in modifying hemoglobin dissociation curve, 2,3-diphosphoglycerate levels, locoregional blood flow, and tumor hypoxia provide additional support for potential beneficial effects during cancer treatment. Unfortunately, only a few clinical studies are available. Finally, we describe some works and our experience supporting the potential role of local ozone therapy in treating delayed healing after tumor resection, to avoid delays in commencing radiotherapy and chemotherapy.Conclusions.In vitroand animal studies, as well as isolated clinical reports, suggest the potential role of ozone as an adjuvant during radiotherapy and/or chemotherapy. However, further research, such as randomized clinical trials, is required to demonstrate its potential usefulness as an adjuvant therapeutic tool.

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The conserved structure of plant telomerase RNA provides the missing link for an evolutionary pathway from ciliates to humans

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1915312116 ◽

2019 ◽

Vol 116 (49) ◽

pp. 24542-24550 ◽

Cited By ~ 8

Author(s):

Jiarui Song ◽

Dhenugen Logeswaran ◽

Claudia Castillo-González ◽

Yang Li ◽

Sreyashree Bose ◽

...

Keyword(s):

Telomerase Activity ◽

Telomere Maintenance ◽

Telomerase Rna ◽

Telomeric Dna ◽

Structural Element ◽

Stem Loop ◽

Evolutionary Pathway ◽

Pol Iii

Telomerase is essential for maintaining telomere integrity. Although telomerase function is widely conserved, the integral telomerase RNA (TR) that provides a template for telomeric DNA synthesis has diverged dramatically. Nevertheless, TR molecules retain 2 highly conserved structural domains critical for catalysis: a template-proximal pseudoknot (PK) structure and a downstream stem-loop structure. Here we introduce the authentic TR from the plant Arabidopsis thaliana, called AtTR, identified through next-generation sequencing of RNAs copurifying with Arabidopsis TERT. This RNA is distinct from the RNA previously described as the templating telomerase RNA, AtTER1. AtTR is a 268-nt Pol III transcript necessary for telomere maintenance in vivo and sufficient with TERT to reconstitute telomerase activity in vitro. Bioinformatics analysis identified 85 AtTR orthologs from 3 major clades of plants: angiosperms, gymnosperms, and lycophytes. Through phylogenetic comparisons, a secondary structure model conserved among plant TRs was inferred and verified using in vitro and in vivo chemical probing. The conserved plant TR structure contains a template-PK core domain enclosed by a P1 stem and a 3′ long-stem P4/5/6, both of which resemble a corresponding structural element in ciliate and vertebrate TRs. However, the plant TR contains additional stems and linkers within the template-PK core, allowing for expansion of PK structure from the simple PK in the smaller ciliate TR during evolution. Thus, the plant TR provides an evolutionary bridge that unites the disparate structures of previously characterized TRs from ciliates and vertebrates.

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Potential Role of the Chemokine Macrophage Inflammatory Protein 1α in Human and Experimental Schistosomiasis

Infection and Immunity ◽

10.1128/iai.73.4.2515-2523.2005 ◽

2005 ◽

Vol 73 (4) ◽

pp. 2515-2523 ◽

Cited By ~ 23

Author(s):

Adriano L. S. Souza ◽

Ester Roffê ◽

Vanessa Pinho ◽

Danielle G. Souza ◽

Adriana F. Silva ◽

...

Keyword(s):

Potential Role ◽

Severe Disease ◽

Experimental Studies ◽

Interleukin 4 ◽

Enzyme Linked Immunosorbent Assay ◽

Inflammatory Protein ◽

Macrophage Inflammatory Protein ◽

Deficient Mice

ABSTRACT In human schistosomiasis, the concentrations of the chemokine macrophage inflammatory protein 1α (MIP-1α/CCL3) is greater in the plasma of patients with clinical hepatosplenic disease. The objective of the present study was to confirm the ability of CCL3 to detect severe disease in patients classified by ultrasonography (US) and to evaluate the potential role of CCL3 in Schistosoma mansoni-infected mice. CCL3 was measured by enzyme-linked immunosorbent assay in the plasma of S. mansoni-infected patients. CCL3-deficient mice were infected with 25 cercariae, and various inflammatory and infectious indices were evaluated. The concentration of CCL3 was higher in the plasma of S. mansoni-infected than noninfected patients. Moreover, CCL3 was greater in those with US-defined hepatosplenic than with the intestinal form of the disease. In CCL3-deficient mice, the size of the granuloma and the liver eosinophil peroxidase activity and collagen content were diminished compared to wild-type mice. In CCL3-deficient mice, the worm burden after 14 weeks of infection, but not after 9 weeks, was consistently smaller. The in vitro response of mesenteric lymph node cells to antigen stimulation was characterized by lower levels of interleukin-4 (IL-4) and IL-10. CCL3 is a marker of disease severity in infected humans, and experimental studies in mice suggest that CCL3 may be a causative factor in the development of severe schistosomiasis.

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Malaria and red cell genetic defects

Blood ◽

10.1182/blood.v74.4.1213.1213 ◽

1989 ◽

Vol 74 (4) ◽

pp. 1213-1221 ◽

Cited By ~ 75

Author(s):

RL Nagel ◽

EF Jr Roth

Keyword(s):

Culture System ◽

Potential Role ◽

Dna Analysis ◽

Red Cell ◽

Metabolic Products ◽

Recombinant Technology ◽

Host Parasite ◽

Made In

Abstract The study of inherited RBC resistance to malaria has increased our knowledge of the biochemistry and physiology of the host-parasite interaction and suggested potential sites for therapeutic intervention. Discovery by Jensen and Trager of the in vitro culture system for P falciparum has facilitated research in this area. Known RBC defects may affect invasion, growth, or merozoite liberation (Fig 1). Significant advances made in understanding mechanisms underlying protection against malaria should not obscure the fact that the data are far from complete. More knowledge is needed about the influence of the erythrocyte cytoskeleton on invasion and growth of parasites as well as the potential role of phospholipids, erythrocyte enzymes other than G6PD, or other metabolic products. Application of DNA analysis and recombinant technology may have an increasing impact on study of the interaction of RBC defects with malarial parasites.

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Intestinal miRNAs regulated in response to dietary lipids

Scientific Reports ◽

10.1038/s41598-020-75751-w ◽

2020 ◽

Vol 10 (1) ◽

Author(s):

Judit Gil-Zamorano ◽

João Tomé-Carneiro ◽

María-Carmen Lopez de las Hazas ◽

Lorena del Pozo-Acebo ◽

M. Carmen Crespo ◽

...

Keyword(s):

Small Intestine ◽

Lipid Metabolism ◽

Chronic Exposure ◽

Potential Role ◽

In Vitro Models ◽

Dietary Lipids ◽

Metabolism Regulation

Abstract The role of miRNAs in intestinal lipid metabolism is poorly described. The small intestine is constantly exposed to high amounts of dietary lipids, and it is under conditions of stress that the functions of miRNAs become especially pronounced. Approaches consisting in either a chronic exposure to cholesterol and triglyceride rich diets (for several days or weeks) or an acute lipid challenge were employed in the search for intestinal miRNAs with a potential role in lipid metabolism regulation. According to our results, changes in miRNA expression in response to fat ingestion are dependent on factors such as time upon exposure, gender and small intestine section. Classic and recent intestinal in vitro models (i.e. differentiated Caco-2 cells and murine organoids) partially mirror miRNA modulation in response to lipid challenges in vivo. Moreover, intestinal miRNAs might play a role in triglyceride absorption and produce changes in lipid accumulation in intestinal tissues as seen in a generated intestinal Dicer1-deletion murine model. Overall, despite some variability between the different experimental cohorts and in vitro models, results show that some miRNAs analysed here are modulated in response to dietary lipids, hence likely to participate in the regulation of lipid metabolism, and call for further research.

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