Key role of piRNAs in telomeric chromatin maintenance and telomere nuclear positioning in Drosophila germline

AbstractTelomeric small RNAs related to PIWI-interacting RNAs (piRNAs) were discovered in different species, however, their role in germline-specific telomere function remains poorly understood. Using a Drosophila model, we show that the piRNA pathway provides a strong germline-specific mechanism of telomere homeostasis. We show that telomeric retrotransposon arrays belong to a unique class of dual-strand piRNA clusters whose transcripts, required for telomere elongation, serve simultaneously as piRNA precursors and their only targets. However, the ability to produce piRNAs and bind Rhino – a germline-specific homolog of heterochromatic protein 1 (HP1) – varies along telomeres. Most likely, this heterogeneity is determined by the peculiarities of telomeric retrotransposons themselves. piRNAs play a pivotal role in the establishment and maintenance of telomeric and subtelomeric chromatin in the germline facilitating loading of HP1 and histone 3 lysine 9 trimethylation mark – highly conservative telomere components – at different telomeric regions. piRNA pathway disruption results in telomere dysfunction characterized by a loss of heterochromatic components and translocation of telomeres from the periphery to the nuclear interior but does not affect the telomere end capping.

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Chromosome End Maintenance by Telomerase

Journal of Biological Chemistry ◽

10.1074/jbc.r900011200 ◽

2009 ◽

Vol 284 (24) ◽

pp. 16061-16065 ◽

Cited By ~ 66

Author(s):

Jennifer L. Osterhage ◽

Katherine L. Friedman

Keyword(s):

Genome Stability ◽

Chromosomal Abnormalities ◽

Human Aging ◽

Dna Complexes ◽

Telomeric Sequences ◽

Telomere Elongation ◽

Telomere Function ◽

Multiple Levels ◽

Linear Chromosomes

Telomeres, protein-DNA complexes at the ends of eukaryotic linear chromosomes, are essential for genome stability. The accumulation of chromosomal abnormalities in the absence of proper telomere function is implicated in human aging and cancer. Repetitive telomeric sequences are maintained by telomerase, a ribonucleoprotein complex containing a reverse transcriptase subunit, a template RNA, and accessory components. Telomere elongation is regulated at multiple levels, including assembly of the telomerase holoenzyme, recruitment of telomerase to the chromosome terminus, and telomere accessibility. This minireview provides an overview of telomerase structure, function, and regulation and the role of telomerase in human disease.

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Germline silencing of UASt depends on the piRNA pathway

10.1101/290726 ◽

2018 ◽

Author(s):

Yi-Chun Huang ◽

Henry Moreno ◽

Sarayu Row ◽

Dongyu Jia ◽

Wu-Min Deng

Keyword(s):

Binary System ◽

Small Rnas ◽

Transgene Expression ◽

Rna Seq ◽

Somatic Tissues ◽

Germline Expression ◽

Germline Cells ◽

Pirna Pathway ◽

Nonsense Mediated Rna Decay

AbstractOne of the most extensively used techniques in Drosophila is the Gal4/UAS binary system, which allows tissue-specific misexpression or knockdown of specific genes of interest. The original UAS vector, UASt, can only be activated for transgene expression in somatic tissues and not in the germline cells. Rørth (1998) generated UASp, a modified UAS vector that is responsive to Gal4 in both somatic and germline tissues, by replacing both the hsp70 promoter and the SV40 3’UTR with the P transposase promoter and the K10 3’UTR respectively. At present, the mechanisms by which UASt is silenced in germline cells are not fully understood. Here, we report that the piRNA pathway is involved in suppressing UASt expression in ovarian germline cells. Individually knocking down or mutating components of the piRNA biogenesis pathway (e.g., Piwi, AGO3, Aub, Spn-E, and Vasa) resulted in the expression of the UASt-reporter (GFP or RFP) in the germline. An RNA-seq analysis of small RNAs revealed that the hsp70 promoter of UASt is targeted by piRNAs, and in the aub mutant ovary, the amount of piRNAs targeting the hsp70 promoter is reduced by around 40 folds. In contrast, the SV40 3’UTR of the UASt, which happens to be targeted by the Nonsense-mediated RNA decay (NMD) pathway, is not responsible for germline UASt suppression, as UASt-reporters with NMD-insensitive 3’UTRs fail to show germline expression. Taken together, our studies reveal a crucial role of the piRNA pathway, potentially via the suppression of the hsp70 promoter, in germline UASt silencing in Drosophila ovaries.

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Regulation of telomere homeostasis and genomic stability in cancer by N6-adenosine methylation (m6A)

Science Advances ◽

10.1126/sciadv.abg7073 ◽

2021 ◽

Vol 7 (31) ◽

pp. eabg7073

Author(s):

Ji Hoon Lee ◽

Juyeong Hong ◽

Zhao Zhang ◽

Bárbara de la Peña Avalos ◽

Cecilia J. Proietti ◽

...

Keyword(s):

Genomic Stability ◽

Cancer Genome ◽

Mrna Levels ◽

Telomere Dysfunction ◽

Rna Methylation ◽

P53 Signaling ◽

Per Se ◽

Telomere Homeostasis ◽

Underlying Mechanisms

The role of RNA methylation on N6-adenosine (m6A) in cancer has been acknowledged, but the underlying mechanisms remain obscure. Here, we identified homeobox containing 1 (HMBOX1) as an authentic target mRNA of m6A machinery, which is highly methylated in malignant cells compared to the normal counterparts and subject to expedited degradation upon the modification. m6A-mediated down-regulation of HMBOX1 causes telomere dysfunction and inactivation of p53 signaling, which leads to chromosome abnormalities and aggressive phenotypes. CRISPR-based, m6A-editing tools further prove that the methyl groups on HMBOX1 per se contribute to the generation of altered cancer genome. In multiple types of human cancers, expression of the RNA methyltransferase METTL3 is negatively correlated with the telomere length but favorably with fractions of altered cancer genome, whereas HMBOX1 mRNA levels show the opposite patterns. Our work suggests that the cancer-driving genomic alterations may potentially be fixed by rectifying particular epitranscriptomic program.

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MicroRNAs: Biological Regulators in Pathogen–Host Interactions

Cells ◽

10.3390/cells9010113 ◽

2020 ◽

Vol 9 (1) ◽

pp. 113 ◽

Cited By ~ 3

Author(s):

Stephanie Maia Acuña ◽

Lucile Maria Floeter-Winter ◽

Sandra Marcia Muxel

Keyword(s):

Immune Response ◽

Infectious Diseases ◽

Small Rnas ◽

Immune Cell ◽

Fine Tuning ◽

Biological Processes ◽

Host Interactions ◽

The Past ◽

Biological Aspects

An inflammatory response is essential for combating invading pathogens. Several effector components, as well as immune cell populations, are involved in mounting an immune response, thereby destroying pathogenic organisms such as bacteria, fungi, viruses, and parasites. In the past decade, microRNAs (miRNAs), a group of noncoding small RNAs, have emerged as functionally significant regulatory molecules with the significant capability of fine-tuning biological processes. The important role of miRNAs in inflammation and immune responses is highlighted by studies in which the regulation of miRNAs in the host was shown to be related to infectious diseases and associated with the eradication or susceptibility of the infection. Here, we review the biological aspects of microRNAs, focusing on their roles as regulators of gene expression during pathogen–host interactions and their implications in the immune response against Leishmania, Trypanosoma, Toxoplasma, and Plasmodium infectious diseases.

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Connecting the αα-hubs: same fold, disordered ligands, new functions

Cell Communication and Signaling ◽

10.1186/s12964-020-00686-8 ◽

2021 ◽

Vol 19 (1) ◽

Author(s):

Lasse Staby ◽

Katrine Bugge ◽

Rasmus Greve Falbe-Hansen ◽

Edoardo Salladini ◽

Karen Skriver ◽

...

Keyword(s):

Intrinsically Disordered Proteins ◽

Disordered Proteins ◽

Telomere Elongation ◽

Protein Protein Interaction ◽

Intrinsically Disordered ◽

Functional Understanding ◽

Signal Specificity ◽

Telomere Regulation ◽

Secondary Structure Motif

Abstract Background Signal fidelity depends on protein–protein interaction–‘hubs’ integrating cues from large interactomes. Recently, and based on a common secondary structure motif, the αα-hubs were defined, which are small α-helical domains of large, modular proteins binding intrinsically disordered transcriptional regulators. Methods Comparative structural biology. Results We assign the harmonin-homology-domain (HHD, also named the harmonin N-terminal domain, NTD) present in large proteins such as harmonin, whirlin, cerebral cavernous malformation 2, and regulator of telomere elongation 1 to the αα-hubs. The new member of the αα-hubs expands functionality to include scaffolding of supra-modular complexes mediating sensory perception, neurovascular integrity and telomere regulation, and reveal novel features of the αα-hubs. As a common trait, the αα-hubs bind intrinsically disordered ligands of similar properties integrating similar cellular cues, but without cross-talk. Conclusion The inclusion of the HHD in the αα-hubs has uncovered new features, exemplifying the utility of identifying groups of hub domains, whereby discoveries in one member may cross-fertilize discoveries in others. These features make the αα-hubs unique models for decomposing signal specificity and fidelity. Using these as models, together with other suitable hub domain, we may advance the functional understanding of hub proteins and their role in cellular communication and signaling, as well as the role of intrinsically disordered proteins in signaling networks.

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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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Telomere Formation by Rap1p Binding Site Arrays Reveals End-Specific Length Regulation Requirements and Active Telomeric Recombination

Molecular and Cellular Biology ◽

10.1128/mcb.21.23.8117-8128.2001 ◽

2001 ◽

Vol 21 (23) ◽

pp. 8117-8128 ◽

Cited By ~ 28

Author(s):

Simona Grossi ◽

Alessandro Bianchi ◽

Pascal Damay ◽

David Shore

Keyword(s):

Binding Sites ◽

De Novo ◽

Repeat Sequence ◽

Independent Manner ◽

Telomere Repeat ◽

Length Regulation ◽

End Capping ◽

Original Array

ABSTRACT Rap1p, the major telomere repeat binding protein in yeast, has been implicated in both de novo telomere formation and telomere length regulation. To characterize the role of Rap1p in these processes in more detail, we studied the generation of telomeres in vivo from linear DNA substrates containing defined arrays of Rap1p binding sites. Consistent with previous work, our results indicate that synthetic Rap1p binding sites within the internal half of a telomeric array are recognized as an integral part of the telomere complex in an orientation-independent manner that is largely insensitive to the precise spacing between adjacent sites. By extending the lengths of these constructs, we found that several different Rap1p site arrays could never be found at the very distal end of a telomere, even when correctly oriented. Instead, these synthetic arrays were always followed by a short (≈100-bp) “cap” of genuine TG repeat sequence, indicating a remarkably strict sequence requirement for an end-specific function(s) of the telomere. Despite this fact, even misoriented Rap1p site arrays promote telomere formation when they are placed at the distal end of a telomere-healing substrate, provided that at least a single correctly oriented site is present within the array. Surprisingly, these heterogeneous arrays of Rap1p binding sites generate telomeres through a RAD52-dependent fusion resolution reaction that results in an inversion of the original array. Our results provide new insights into the nature of telomere end capping and reveal one way by which recombination can resolve a defect in this process.

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The role of exploratory mechanisms in evolution

Philosophical Transactions of the Royal Society B Biological Sciences ◽

10.1098/rstb.1995.0116 ◽

1995 ◽

Vol 349 (1329) ◽

pp. 297-297

Keyword(s):

Dynamic Instability ◽

Specific Mechanism ◽

Cellular Mechanisms ◽

Selection Processes ◽

Wide Range ◽

Rapid Changes ◽

Neuronal Processes ◽

History Of ◽

Variation And Selection

Many cellular mechanisms use a process of variation and selection to generate specific patterns. Among these, dynamic instability of microtubules has been shown to employ a specific mechanism to intentionally generate variation. In many systems the growth of neurons or neuronal processes is excessive, the final connections being established by stabilization of functional interactions. When changes in neuronal networks take place, such as in metamorphosis, use is made of the plasticity of neuronal connectivity. In the immune system, specific responses are generated by variation and selection. Processes that explore a wide range of conditions and a wide range of structures can be called exploratory processes. These are very robust and capable of responding to damage, variability in the environment and ontogenic changes in the organisms. Such robustness would be useful for adapting to changes that occur during phylogenetic changes as well. Given the extensive history of extinction and radiation in evolution, it may be supposed that these mechanisms have themselves been selected for their capacity to survive rapid changes in the organism and for their ability to generate cellular variation.

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