Telomere Length in Chromosomally Normal and Abnormal Miscarriages and Ongoing Pregnancies and Its Association with 5-hydroxymethylcytosine Patterns

The present study investigates telomere length (TL) in dividing chorionic cytotrophoblast cells from karyotypically normal and abnormal first trimester miscarriages and ongoing pregnancies. Using Q-FISH, we measured relative TLs in the metaphase chromosomes of 61 chorionic villous samples. Relative TLs did not differ between karyotypically normal samples from miscarriages and those from ongoing pregnancies (p = 0.3739). However, among the karyotypically abnormal samples, relative TLs were significantly higher in ongoing pregnancies than in miscarriages (p < 0.0001). Relative TLs were also significantly higher in chorion samples from karyotypically abnormal ongoing pregnancies than in those from karyotypically normal ones (p = 0.0018) in contrast to miscarriages, where relative TL values were higher in the karyotypically normal samples (p = 0.002). In the karyotypically abnormal chorionic cytotrophoblast, the TL variance was significantly lower than in any other group (p < 0.05). Assessed by TL ratios between sister chromatids, interchromatid TL asymmetry demonstrated similar patterns across all of the chorion samples (p = 0.22) but significantly exceeded that in PHA-stimulated lymphocytes (p < 0.0001, p = 0.0003). The longer telomere was predominantly present in the hydroxymethylated sister chromatid in chromosomes featuring hemihydroxymethylation (containing 5-hydroxymethylcytosine in only one sister chromatid)—a typical sign of chorionic cytotrophoblast cells. Our results suggest that the phenomena of interchromatid TL asymmetry and its association to 5hmC patterns in chorionic cytotrophoblast, which are potentially linked to telomere lengthening through recombination, are inherent to the development programme. The TL differences in chorionic cytotrophoblast that are associated with karyotype and embryo viability seem to be determined by heredity rather than telomere elongation mechanisms. The inheritance of long telomeres by a karyotypically abnormal embryo promotes his development, whereas TL in karyotypically normal first-trimester embryos does not seem to have a considerable impact on developmental capacity.

Download Full-text

The Role of Rif1 in telomere length regulation is separable from its role in origin firing

10.1101/2020.04.24.060491 ◽

2020 ◽

Author(s):

Calla B. Shubin ◽

Carol W. Greider

Keyword(s):

Telomere Length ◽

Protein Phosphatase ◽

Protein Phosphatase 1 ◽

Origin Firing ◽

Replication Complex ◽

Telomere Elongation ◽

Length Regulation ◽

Telomere Length Regulation ◽

Telomere Lengthening

AbstractTo examine the established link between DNA replication and telomere length, we tested whether firing of telomeric origins would cause telomere lengthening. We found that RIF1 mutants that block Protein Phosphatase 1 (PP1) binding activated telomeric origins but did not elongate telomeres. In a second approach, we found overexpression of ΔN-Dbf4 and Cdc7 increased DDK activity and activated telomeric origins, yet telomere length was unchanged. We tested a third mechanism to activate origins using the sld3-A mcm5-bob1 mutant that deregulates the pre-replication complex, and again saw no change in telomere length. Finally, we tested whether mutations in RIF1 that cause telomere elongation would affect origin firing. We found that neither rif1-Δ1322 nor rif1HOOK affected firing of telomeric origins. We conclude that telomeric origin firing does not cause telomere elongation, and the role of Rif1 in regulating origin firing is separable from its role in regulating telomere length.

Download Full-text

The role of Rif1 in telomere length regulation is separable from its role in origin firing

eLife ◽

10.7554/elife.58066 ◽

2020 ◽

Vol 9 ◽

Cited By ~ 1

Author(s):

Calla B Shubin ◽

Carol W Greider

Keyword(s):

Telomere Length ◽

Protein Phosphatase ◽

Protein Phosphatase 1 ◽

Origin Firing ◽

Replication Complex ◽

Telomere Elongation ◽

Length Regulation ◽

Telomere Length Regulation ◽

Telomere Lengthening

To examine the established link between DNA replication and telomere length, we tested whether firing of telomeric origins would cause telomere lengthening. We found that RIF1 mutants that block Protein Phosphatase 1 (PP1) binding activated telomeric origins but did not elongate telomeres. In a second approach, we found overexpression of ∆N-Dbf4 and Cdc7 increased DDK activity and activated telomeric origins, yet telomere length was unchanged. We tested a third mechanism to activate origins using the sld3-A mcm5-bob1 mutant that de-regulates the pre-replication complex, and again saw no change in telomere length. Finally, we tested whether mutations in RIF1 that cause telomere elongation would affect origin firing. We found that neither rif1-∆1322 nor rif1HOOK affected firing of telomeric origins. We conclude that telomeric origin firing does not cause telomere elongation, and the role of Rif1 in regulating origin firing is separable from its role in regulating telomere length.

Download Full-text

Sudden Telomere Lengthening Triggers a Rad53-dependent Checkpoint inSaccharomyces cerevisiae

Molecular Biology of the Cell ◽

10.1091/mbc.e02-11-0719 ◽

2003 ◽

Vol 14 (8) ◽

pp. 3126-3143 ◽

Cited By ~ 14

Author(s):

Valeria Viscardi ◽

Enrico Baroni ◽

Michele Romano ◽

Giovanna Lucchini ◽

Maria Pia Longhese

Keyword(s):

Telomere Length ◽

Double Strand Breaks ◽

Protein Fusion ◽

Dna Breaks ◽

Checkpoint Response ◽

Strand Breaks ◽

Telomere Elongation ◽

Cycle Arrest ◽

Telomeric Binding Protein ◽

Telomere Lengthening

Telomeres are specialized functional complexes that ensure chromosome stability by protecting chromosome ends from fusions and degradation and avoiding chromosomal termini from being sensed as DNA breaks. Budding yeast Tel1 is required both for telomere metabolism and for a Rad53-dependent checkpoint responding to unprocessed double-strand breaks. We show that overexpression of a GAL1-TEL1 fusion causes transient telomere lengthening and activation of a Rad53-dependent G2/M checkpoint in cells whose telomeres are short due to the lack of either Tel1 or Yku70. Sudden telomere elongation and checkpoint-mediated cell cycle arrest are also triggered in wild-type cells by overproducing a protein fusion between the telomeric binding protein Cdc13 and the telomerase-associated protein Est1. Checkpoint activation by GAL1-TEL1 requires ongoing telomere elongation. In fact, it is turned off concomitantly with telomeres reaching a new stable length and is partially suppressed by deletion of the telomerase EST2 gene. Moreover, both telomere length rebalancing and checkpoint inactivation under galactose-induced conditions are accelerated by high levels of either the Sae2 protein, involved in double-strand breaks processing, or the negative telomere length regulator Rif2. These data suggest that sudden telomere lengthening elicits a checkpoint response that inhibits the G2/M transition.

Download Full-text

91 SPERMATOZOA TELOMERE LENGTH DETERMINES EMBRYONIC TELOMERE LENGTH BEFORE EMBRYONIC GENOME ACTIVATION

Reproduction Fertility and Development ◽

10.1071/rdv25n1ab91 ◽

2013 ◽

Vol 25 (1) ◽

pp. 193

Author(s):

C. de Frutos ◽

R. Laguna-Barraza ◽

P. Bermejo-Alvarez ◽

D. Rizos ◽

A. Gutierrez-Adan

Keyword(s):

Telomere Length ◽

Mus Musculus ◽

Mus Spretus ◽

Telomere Elongation ◽

Embryonic Genome Activation ◽

Male Gametes ◽

Embryonic Genome ◽

Cell Embryo ◽

Telomere Lengthening ◽

Genome Activation

A critical issue for species integrity is the existence of a telomere elongation program during embryogenesis that ensures sufficient telomere reserves in mammalian newborns. Two different mechanisms have been reported to act on telomere elongation during early embryogenesis: first, the telomerase, the ribonucleoprotein that adds telomeric repeats onto the chromosome ends, known to be responsible for the telomere lengthening at the morula-blastocyst transition in mice and bovine; second, in laboratory mice strains, mature oocytes increase the length of their relatively short telomeres between the 1-cell and 2-cell stages by a recombination or ALT-like pathway. In contrast, spermatozoa, the terminally differentiated male gametes, exhibit a very long telomere length (TL). The aim of this study was to clarify the potential role of the spermatozoa TL in the telomere lengthening occurring between oocyte and the 2-cell stage. For this purpose, we used 2 mouse species known to differ greatly in their TL [Mus musculus (hybrid C57CBAF1), long TL, and Mus spretus, short TL]. First, we compared relative TL in sperm samples from 5 age-matched males of each species by quantitative real-time PCR, with the numbers of telomere repeats being normalized, to the amount of DNA present in the sample (based on quantification of the Rn18S gene) by the comparative Ct method. Then, 1- and 2-cell embryos were produced by fertilizing Mus musculus oocytes with either Mus musculus or Mus spretus spermatozoa. The TL analysis in oocytes, zygotes, or 2-cell embryos was carried out by absolute quantification of telomere repeats by qPCR and normalized to the highest Ct observed value. Twenty to thirty samples per stage were analyzed, with each sample consisting in 2 matured oocytes, 2 zygotes, or one 2-cell embryo, to allow comparisons between stages. One-way ANOVA was used for statistical analysis. Mus spretus spermatozoa had significantly shorter telomeres than did Mus musculus (1.0 ± 0.1 v. 9.0 ± 1.5, respectively; P ≤ 0.01). The TL increased after fertilization from oocyte to zygote and 2-cell embryo stages in Mus musculus (1.0 ± 0.1, 1.5 ± 0.1, and 2.4 ± 0.2, respectively; P ≤ 0.01). In contrast, no differences were found in the TLs between the 3 stages in Mus spretus hybrids (oocyte: 1.0 ± 0.1; zygote: 1.0 ± 0.1; and 2-cell embryo: 1.0 ± 0.1), indicating that no elongation occurred after fertilization with spermatozoa with short telomeres. Herein, we demonstrated that before embryonic genome activation occurs, spermatozoa TL determines TL of the early embryo, suggesting that spermatozoon telomeres may act as recombination templates for early telomere lengthening right after syngamia.

Download Full-text

Maternal Vitamin D and Newborn Telomere Length

Nutrients ◽

10.3390/nu13062012 ◽

2021 ◽

Vol 13 (6) ◽

pp. 2012

Author(s):

Lisa Daneels ◽

Dries S. Martens ◽

Soumia Arredouani ◽

Jaak Billen ◽

Gudrun Koppen ◽

...

Keyword(s):

Vitamin D ◽

Cord Blood ◽

Telomere Length ◽

First Trimester ◽

Maternal Serum ◽

Adverse Outcomes ◽

Dietary Vitamin ◽

Maternal Vitamin ◽

Vitamin D Intake ◽

Diet Supplements

Nutrition is important during pregnancy for offspring health. Gestational vitamin D intake may prevent several adverse outcomes and might have an influence on offspring telomere length (TL). In this study, we want to assess the association between maternal vitamin D intake during pregnancy and newborn TL, as reflected by cord blood TL. We studied mother–child pairs enrolled in the Maternal Nutrition and Offspring’s Epigenome (MANOE) cohort, Leuven, Belgium. To calculate the dietary vitamin D intake, 108 women were asked to keep track of their diet using the seven-day estimated diet record (EDR) method. TL was assessed in 108 cord blood using a quantitative real-time PCR method. In each trimester of pregnancy, maternal serum 25-hydroxyvitamin D (25-OHD) concentration was measured. We observed a positive association (β = 0.009, p-value = 0.036) between newborn average relative TL and maternal vitamin D intake (diet + supplement) during the first trimester. In contrast, we found no association between average relative TL of the newborn and mean maternal serum 25-OHD concentrations during pregnancy. To conclude, vitamin D intake (diet + supplements), specifically during the first trimester of pregnancy, is an important factor associated with TL at birth.

Download Full-text

RAD50 and RAD51 Define Two Pathways That Collaborate to Maintain Telomeres in the Absence of Telomerase

Genetics ◽

10.1093/genetics/152.1.143 ◽

1999 ◽

Vol 152 (1) ◽

pp. 143-152 ◽

Cited By ~ 12

Author(s):

Siyuan Le ◽

J Kent Moore ◽

James E Haber ◽

Carol W Greider

Keyword(s):

Cell Death ◽

Telomere Length ◽

Gene Conversion ◽

De Novo ◽

Dna Recombination ◽

Telomere Maintenance ◽

Triple Mutant ◽

Yeast Telomerase ◽

Telomere Lengthening

Abstract Telomere length is maintained by the de novo addition of telomere repeats by telomerase, yet recombination can elongate telomeres in the absence of telomerase. When the yeast telomerase RNA component, TLC1, is deleted, telomeres shorten and most cells die. However, gene conversion mediated by the RAD52 pathway allows telomere lengthening in rare survivor cells. To further investigate the role of recombination in telomere maintenance, we assayed telomere length and the ability to generate survivors in several isogenic DNA recombination mutants, including rad50, rad51, rad52, rad54, rad57, xrs2, and mre11. The rad51, rad52, rad54, and rad57 mutations increased the rate of cell death in the absence of TLC1. In contrast, although the rad50, xrs2, and mre11 strains initially had short telomeres, double mutants with tlc1 did not affect the rate of cell death, and survivors were generated at later times than tlc1 alone. While none of the double mutants of recombination genes and tlc1 (except rad52 tlc1) blocked the ability to generate survivors, a rad50 rad51 tlc1 triple mutant did not allow the generation of survivors. Thus RAD50 and RAD51 define two separate pathways that collaborate to allow cells to survive in the absence of telomerase.

Download Full-text

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.

Download Full-text

Development of software and modification of Q-FISH protocol for estimation of individual telomere length in immunopathology

Journal of Bioinformatics and Computational Biology ◽

10.1142/s0219720016500414 ◽

2017 ◽

Vol 15 (02) ◽

pp. 1650041 ◽

Cited By ~ 3

Author(s):

M. Sh. Barkovskaya ◽

A. G. Bogomolov ◽

N. Yu. Knauer ◽

N. B. Rubtsov ◽

V. A. Kozlov

Keyword(s):

Telomere Length ◽

Inflammatory Diseases ◽

Quantitative Information ◽

Computer Assisted ◽

Metaphase Chromosomes ◽

Important Indicator ◽

Background Fluorescence ◽

Immune Mediated ◽

Light Effects ◽

Assisted Analysis

Telomere length is an important indicator of proliferative cell history and potential. Decreasing telomere length in the cells of an immune system can indicate immune aging in immune-mediated and chronic inflammatory diseases. Quantitative fluorescent in situ hybridization (Q-FISH) of a labeled (C3TA[Formula: see text] peptide nucleic acid probe onto fixed metaphase cells followed by digital image microscopy allows the evaluation of telomere length in the arms of individual chromosomes. Computer-assisted analysis of microscopic images can provide quantitative information on the number of telomeric repeats in individual telomeres. We developed new software to estimate telomere length. The MeTeLen software contains new options that can be used to solve some Q-FISH and microscopy problems, including correction of irregular light effects and elimination of background fluorescence. The identification and description of chromosomes and chromosome regions are essential to the Q-FISH technique. To improve the quality of cytogenetic analysis after Q-FISH, we optimized the temperature and time of DNA-denaturation to get better DAPI-banding of metaphase chromosomes. MeTeLen was tested by comparing telomere length estimations for sister chromatids, background fluorescence estimations, and correction of nonuniform light effects. The application of the developed software for analysis of telomere length in patients with rheumatoid arthritis was demonstrated.

Download Full-text

Interleukin (IL)-1β Stimulates Migration and Survival of First-Trimester Villous Cytotrophoblast Cells through Endometrial Epithelial Cell-Derived IL-8

Endocrinology ◽

10.1210/en.2008-0264 ◽

2008 ◽

Vol 150 (1) ◽

pp. 350-356 ◽

Cited By ~ 30

Author(s):

Yasushi Hirota ◽

Yutaka Osuga ◽

Akiko Hasegawa ◽

Ako Kodama ◽

Toshiki Tajima ◽

...

Keyword(s):

Epithelial Cell ◽

First Trimester ◽

Endometrial Epithelial Cell ◽

Cytotrophoblast Cells

Download Full-text

Spermatozoa telomeres determine telomere length in early embryos and offspring

Reproduction ◽

10.1530/rep-15-0375 ◽

2016 ◽

Vol 151 (1) ◽

pp. 1-7 ◽

Cited By ~ 27

Author(s):

C de Frutos ◽

A P López-Cardona ◽

N Fonseca Balvís ◽

R Laguna-Barraza ◽

D Rizos ◽

...

Keyword(s):

Telomere Length ◽

Mus Musculus ◽

Mus Spretus ◽

Cell Stage ◽

Young Males ◽

Early Embryos ◽

Parent Of Origin ◽

Telomere Lengthening ◽

Zygote Stage

Offspring telomere length (TL) has been correlated with paternal TL, but the mechanism for this parent of origin-specific inheritance remains unclear. The objective of this study has been to determine the role of spermatozoa TL in embryonic telomere lengthening by using two mouse models showing dimorphism in their spermatozoa TL: Mus musculus vs Mus spretus and old vs young Mus musculus. Mus spretus spermatozoa displayed a shorter TL than Mus musculus. Hybrid offspring exhibited lower TL compared with Mus musculus starting at the two-cell stage, before the onset of telomerase expression. To analyze the role of spermatozoa telomeres in early telomere lengthening, we compared the TL in oocytes, zygotes, two-cell embryos and blastocysts produced by parthenogenesis or by fertilization with Mus musculus or Mus spretus spermatozoa. TL was significantly higher in spermatozoa compared with oocytes, and it increased significantly from the oocyte to the zygote stage in those embryos fertilized with Mus musculus spermatozoa, but not in those fertilized with Mus spretus spermatozoa or produced by parthenogenesis. A further increase was noted from the zygote to the two-cell stage in fertilized Mus musculus embryos, whereas hybrid embryos maintained the oocyte TL. Spermatozoa TL shortened with age in Mus musculus and the offspring from young males showed a significantly higher TL compared with that fathered by old males. These significant differences were already noticeable at the two-cell stage. These results suggest that spermatozoa telomeres act as a guide for telomerase-independent telomere lengthening resulting in differences in TL that persist after birth.Free Spanish abstract: A Spanish translation of this abstract is freely available at http://www.reproduction-online.org/content/151/1/1/suppl/DC1.

Download Full-text