scholarly journals DAF-16/Foxo suppresses the transgenerational sterility of prg-1 piRNA mutants via a systemic small RNA pathway

2018 ◽  
Author(s):  
Matt Simon ◽  
Maya Spichal ◽  
Bree Heestand ◽  
Stephen Frenk ◽  
Ashley Hedges ◽  
...  
Keyword(s):  
Germ Cell ◽  
Small Rna ◽  
Immune Surveillance ◽  
Somatic Cells ◽  
Rnai Pathway ◽  
Transmembrane Channel ◽  
Dsrna Binding ◽  
Dsrna Binding Protein ◽  
Systemic Rnai ◽  
Cell Immortality

AbstractMutation of the daf-2 insulin/IGF-1 receptor activates the DAF-16/Foxo transcription factor to suppress the transgenerational sterility phenotype of prg-1/piRNA mutants that are deficient for piRNA-mediated genome silencing. As with PRG-1/piRNAs, mutations in the nuclear RNA interference gene nrde-1 compromised germ cell immortality, but deficiency for daf-2 did not suppress the transgenerational sterility of nrde-1 or nrde-4 single mutants or of prg-1; nrde-4 or prg-1; hrde-1 double mutants. NRDE-1 and NRDE-4 promote transcriptional silencing in somatic cells via the nuclear Argonaute protein NRDE-3, which was dispensable for germ cell immortality. However, daf-2 deficiency failed to promote germ cell immortality in prg-1; nrde-3 mutants. Consistently, we found that DAF-16 activity in somatic cells suppressed the transgenerational sterility of prg-1 mutants via the SID-1 dsRNA transmembrane channel that promotes systemic RNAi as well as Dicer, the dsRNA binding protein RDE-4 and the RDRP RRF-3. We conclude that DAF-16 activates a cell-non-autonomous systemic RNAi pathway that promotes small RNA-mediated genome silencing in germ cells to suppress loss of the genomic immune surveillance factor Piwi/PRG-1.

scholarly journals Deficiency for Piwi results in transmission of a heritable stress that promotes longevity via DAF-16/Foxo

2018 ◽  
Author(s):  
Bree Heestand ◽  
Matt Simon ◽  
Stephen Frenk ◽  
Shawn Ahmed
Keyword(s):  
Germ Cell ◽  
Germ Cells ◽  
Stress Resistance ◽  
Rnai Pathway ◽  
Early Generation ◽  
C Elegans ◽  
Systemic Rnai ◽  
Late Generation ◽  
Cell Immortality ◽  
F1 Cross

AbstractThe C. elegans Piwi Argonaute protein PRG-1 and associated piRNAs protect the genomes of germ cells by suppressing the expression of transposons and potentially deleterious foreign nucleic acids. Deficiency for prg-1 compromises germ cell immortality, resulting in normal fertility for many generations followed by progressively reduced fertility and ultimately sterility. The sterility phenotype of prg-1 mutants was recently shown to be a form of reproductive arrest, which implies that prg-1 mutants may become sterile in response to a form of heritable stress. The DAF-16 stress resistance and longevity factor can promote germ cell immortality of prg-1 mutants by activating a systemic RNAi pathway. We found that this RNAi pathway was not required for the somatic longevity function of DAF-16. Given that prg-1 mutant germ cells may transmit a form of heritable stress, we studied the somatic longevity of prg-1 mutant adults. We found that early generation prg-1 mutants had normal lifespans, but that late-generation adults that displayed reduced fertility or sterility were long lived. Germ cells of long-lived late-generation prg-1 mutants gave rise to F1 cross progeny that were heterozygous for prg-1, fertile and also long lived. However, in the absence of DAF-16, the heritable stress transmitted by prg-1 mutant germ cells was deleterious and caused lifespan to shorten. We conclude that deficiency for the genomic surveillance factor PRG-1/Piwi results in germ cells that transmit a heritable stress that promotes somatic longevity via DAF-16/Foxo, which could be relevant transgenerational regulationof aging.

scholarly journals Each cell needs long double-stranded RNA for silencing by feeding RNAi in C. elegans

2016 ◽  
Author(s):  
Pravrutha Raman ◽  
Soriayah M Zaghab ◽  
Edward C Traver ◽  
Antony M Jose
Keyword(s):  
Rna Interference ◽  
Single Copy ◽  
Animal Cells ◽  
Tissue Specific ◽  
Double Stranded Rna ◽  
C Elegans ◽  
Dsrna Binding ◽  
Dsrna Binding Protein ◽  
Systemic Rnai ◽  
Derivatives Of

ABSTRACTLong double-stranded RNA (dsRNA) can silence genes of matching sequence upon ingestion in many invertebrates and is therefore being developed as a pesticide. Such feeding RNA interference (RNAi) is best understood in the worm C. elegans, where it is thought that derivatives of ingested dsRNA, including short dsRNAs, move between cells and cause systemic silencing. Movement of short dsRNAs has been inferred using tissue-specific rescue of the long dsRNA-binding protein RDE-4 by expressing it from repetitive transgenes. We found that the use of repetitive transgenes for the tissue-specific rescue of a gene could inhibit RNAi within that tissue and could result in misexpression of the gene in other tissues. Both inhibition and misexpression were not detectable when a single-copy transgene was used for tissue-specific rescue. In animals with single-copy rescue of RDE-4, RNAi was restricted to the tissue with RDE-4 expression. Thus, unlike previous observations using repetitive transgenes, these results suggest that binding of long dsRNA by RDE-4 in each silenced cell is required for systemic RNAi. Taken together with the requirement for long dsRNA to trigger RNAi in insects, these results suggest that the entry of long dsRNA is a necessary first step for feeding RNAi in animal cells.

Chemical shift assignments of DRB4 (1–153), a dsRNA binding protein in A. thaliana RNAi pathway

2014 ◽  
Vol 9 (2) ◽  
pp. 253-256 ◽  
Author(s):  
Sai Chaitanya Chiliveri ◽  
Mandar V. Deshmukh
Keyword(s):  
Chemical Shift ◽  
Binding Protein ◽  
Rnai Pathway ◽  
Chemical Shift Assignments ◽  
Dsrna Binding ◽  
Dsrna Binding Protein

scholarly journals The origin and evolution of loqs2: a gene encoding an antiviral dsRNA binding protein in Aedes mosquitoes

2021 ◽  
Author(s):  
Carlos F. Estevez-Castro ◽  
Murillo F. Rodrigues ◽  
Antinéa Babarit ◽  
Flávia Viana Ferreira ◽  
Eric Marois ◽  
...  
Keyword(s):  
Rna Binding ◽  
Binding Protein ◽  
Mosquito Species ◽  
Rnai Pathway ◽  
Gene Encoding ◽  
Origin And Evolution ◽  
Larval Stages ◽  
Dsrna Binding ◽  
Dsrna Binding Protein ◽  
Antiviral Gene

Mosquito borne viruses such as dengue, Zika, yellow fever and Chikungunya cause millions of infections every year. These viruses are mostly transmitted by two urban-adapted mosquito species, Aedes aegypti and Aedes albopictus, that appear to be more permissive to arbovirus infections compared to closely related species. Although mechanistic understanding remains, Aedes mosquitoes may have evolved specialized antiviral mechanisms that potentially contribute to the low impact of viral infection. Recently, we reported the identification of an Aedes specific double-stranded RNA binding protein (dsRBP), named Loqs2, that is involved in the control of infection by dengue and Zika viruses in Ae. aegypti. Loqs2 interacts with two important co-factors of the RNA interference (RNAi) pathway, Loquacious (Loqs) and R2D2, and seems to be a strong regulator of the antiviral defense. However, the origin and evolution of loqs2 remains unclear. Here, we describe that loqs2 likely originated from two independent duplications of the first dsRNA binding domain (dsRBD) of loquacious that occurred before the radiation of the Aedes Stegomya subgenus. After its origin, our analyses suggest that loqs2 evolved by relaxed positive selection towards neofunctionalization. In fact, loqs2 is evolving at a faster pace compared to other RNAi components such as loquacious, r2d2 and Dicer-2 in Aedes mosquitoes. Unlike loquacious, transcriptomic analysis showed that loqs2 expression is tightly regulated, almost restricted to reproductive tissues in Ae. aegypti and Ae. albopictus. Transgenic mosquitoes engineered to ubiquitously express loqs2 show massive dysregulation of stress response genes and undergo developmental arrest at larval stages. Overall, our results uncover the possible origin and neofunctionalization of a novel antiviral gene, loqs2, in Aedes mosquitoes that ultimately may contribute to their effectiveness as vectors for arboviruses.

scholarly journals The conserved phosphatase GSP-2/PP1 promotes germline immortality via small RNA-mediated genome silencing during meiosis

2018 ◽  
Author(s):  
Katherine Kretovich Billmyre ◽  
Anna-lisa Doebley ◽  
Bree Heestand ◽  
Tony Belicard ◽  
Aya Sato-Carlton ◽  
...  
Keyword(s):  
Germ Cell ◽  
Chromosome Segregation ◽  
Small Rna ◽  
Protein Phosphatase ◽  
Germ Line ◽  
Meiotic Chromosome ◽  
Loss Of Function ◽  
Homologous Chromosomes ◽  
C Elegans ◽  
Cell Immortality

AbstractGenomic silencing can promote germ cell immortality, or transgenerational maintenance of the germ line, via mechanisms that may occur during mitosis or meiosis. Here we report that the gsp-2 PP1/Glc7 phosphatase promotes germ cell immortality. We identified a separation-of-function allele of C. elegans GSP-2 that caused a meiosis-specific chromosome segregation defect and defects in transgenerational small RNA-induced genome silencing. GSP-2 is recruited to meiotic chromosomes by LAB-1, which also promoted germ cell immortality. Sterile gsp-2 and lab-1 mutant adults displayed germline degeneration, univalents and histone phosphorylation defects in oocytes, similar to small RNA genome silencing mutants. Epistasis and RNA analysis suggested that GSP-2 functions downstream of small RNAs. We conclude that a meiosis-specific function of GSP-2/LAB-1 ties small RNA-mediated silencing of the epigenome to germ cell immortality. Given that hemizygous genetic elements can drive transgenerational epigenomic silencing, and given that LAB-1 promotes pairing of homologous chromosomes and localizes to the interface between homologous chromosomes during pachytene, we suggest that discontinuities at this interface could promote nuclear silencing in a manner that depends on GSP-2.Author SummaryThe germ line of an organism is considered immortal in its capacity to give rise to an unlimited number of future generations. To protect the integrity of the germ line, mechanisms act to suppress the accumulation of transgenerational damage to the genome or epigenome. Loss of germ cell immortality can result from mutations that disrupt the small RNA-mediated silencing pathway that helps to protect the integrity of the epigenome. Here we report for the first time that the C. elegans protein phosphatase GSP-2 that promotes core chromosome biology functions during meiosis is also required for germ cell immortality. Specifically, we identified a partial loss of function allele of gsp-2 that exhibits defects in meiotic chromosome segregation and is also dysfunctional for transgenerational small RNA-mediated genome silencing. Our results are consistent with a known role of Drosophila Protein Phosphatase 1 in heterochromatin silencing, and point to a meiotic phosphatase function that is relevant to germ cell immortality, conceivably related to its roles in chromosome pairing or sister chromatid cohesion.

scholarly journals The Human dsRNA binding protein PACT is unable to functionally substitute for the Drosophila dsRNA binding protein R2D2

F1000Research ◽  
2014 ◽  
Vol 2 ◽  
pp. 220
Author(s):  
Benjamin K Dickerman ◽  
Jocelyn A McDonald ◽  
Ganes C Sen
Keyword(s):  
Small Rna ◽  
Binding Protein ◽  
Dependent Protein Kinase ◽  
Dsrna Binding ◽  
Dsrna Binding Protein ◽  
Rna Biogenesis ◽  
Dependent Protein ◽  
Response To Stress ◽  
Stress Signals ◽  
Transcriptional Gene Regulation

The dsRNA binding protein (dsRBP) PACT was first described as an activator of the dsRNA dependent protein kinase PKR in response to stress signals.  Additionally, it has been identified as a component of the small RNA processing pathway.  A role for PACT in this pathway represents an important interplay between two modes of post-transcriptional gene regulation.  The function of PACT in this context is poorly understood.  Thus, additional approaches are required to clarify the mechanism by which PACT functions.  In this study, the genetic utility of Drosophila melanogaster was employed to identify dsRNA-binding proteins that are functionally orthologous to PACT.  Transgenic Drosophila expressing human PACT were generated to determine whether PACT is capable of functionally substituting for the Drosophila dsRBP R2D2, which has a well-defined role in small RNA biogenesis.  Results presented here indicate that PACT is unable to substitute for R2D2 at the whole organism level.

scholarly journals The Human dsRNA binding protein PACT is unable to functionally substitute for the Drosophila dsRNA binding protein R2D2

F1000Research ◽  
2013 ◽  
Vol 2 ◽  
pp. 220
Author(s):  
Benjamin K Dickerman ◽  
Jocelyn A McDonald ◽  
Ganes C Sen
Keyword(s):  
Small Rna ◽  
Binding Protein ◽  
Dependent Protein Kinase ◽  
Dsrna Binding ◽  
Dsrna Binding Protein ◽  
Rna Biogenesis ◽  
Dependent Protein ◽  
Response To Stress ◽  
Stress Signals ◽  
Transcriptional Gene Regulation

The primary function of the dsRNA binding protein (dsRBP) PACT/RAX is to activate the dsRNA dependent protein kinase PKR in response to stress signals.  Additionally, it has been identified as a component of the small RNA processing pathway.  A role for PACT/RAX in this pathway represents an important interplay between two modes of post-transcriptional gene regulation.  The function of PACT/RAX in this context is poorly understood.  Thus, additional models are required to clarify the mechanism by which PACT/RAX functions.  In this study, Drosophila melanogaster was employed to identify functionally orthologous dsRNA-binding proteins.  Transgenic Drosophila expressing human PACT were generated to determine whether PACT is capable of functionally substituting for the Drosophila dsRBP R2D2, which has a well-defined role in small RNA biogenesis.  Results presented here indicate that PACT is unable to substitute for R2D2 at the whole organism level.

scholarly journals Environmental RNAi pathways in the two-spotted spider mite

BMC Genomics ◽  
2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Mosharrof Mondal ◽  
Jacob Peter ◽  
Obrie Scarbrough ◽  
Alex Flynt
Keyword(s):  
Gene Expression ◽  
Small Rna ◽  
Spider Mite ◽  
Genetic Manipulation ◽  
Model Organisms ◽  
Rnai Pathway ◽  
Plant Host ◽  
Rnai Technology ◽  
Two Spotted Spider Mite ◽  
Multicellular Organisms

Abstract Background RNA interference (RNAi) regulates gene expression in most multicellular organisms through binding of small RNA effectors to target transcripts. Exploiting this process is a popular strategy for genetic manipulation and has applications that includes arthropod pest control. RNAi technologies are dependent on delivery method with the most convenient likely being feeding, which is effective in some animals while others are insensitive. The two-spotted spider mite, Tetranychus urticae, is prime candidate for developing RNAi approaches due to frequent occurrence of conventional pesticide resistance. Using a sequencing-based approach, the fate of ingested RNAs was explored to identify features and conditions that affect small RNA biogenesis from external sources to better inform RNAi design. Results Biochemical and sequencing approaches in conjunction with extensive computational assessment were used to evaluate metabolism of ingested RNAs in T. urticae. This chelicerae arthropod shows only modest response to oral RNAi and has biogenesis pathways distinct from model organisms. Processing of synthetic and plant host RNAs ingested during feeding were evaluated to identify active substrates for spider mite RNAi pathways. Through cataloging characteristics of biochemically purified RNA from these sources, trans-acting small RNAs could be distinguished from degradation fragments and their origins documented. Conclusions Using a strategy that delineates small RNA processing, we found many transcripts have the potential to enter spider mite RNAi pathways, however, trans-acting RNAs appear very unstable and rare. This suggests potential RNAi pathway substrates from ingested materials are mostly degraded and infrequently converted into regulators of gene expression. Spider mites infest a variety of plants, and it would be maladaptive to generate diverse gene regulators from dietary RNAs. This study provides a framework for assessing RNAi technology in organisms where genetic and biochemical tools are absent and benefit rationale design of RNAi triggers for T.urticae.

scholarly journals Müllerian Inhibiting Substance Is Required for Germ Cell Proliferation during Early Gonadal Differentiation in Medaka (Oryzias latipes)

Endocrinology ◽  
2007 ◽  
Vol 149 (4) ◽  
pp. 1813-1819 ◽  
Author(s):  
Eri Shiraishi ◽  
Norifumi Yoshinaga ◽  
Takeshi Miura ◽  
Hayato Yokoi ◽  
Yuko Wakamatsu ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Germ Cell ◽  
Germ Cells ◽  
Sex Differentiation ◽  
Somatic Cells ◽  
Oryzias Latipes ◽  
Cell Number ◽  
Gonadal Differentiation ◽  
Loss Of Function ◽  
Germ Cell Proliferation

Müllerian inhibiting substance (MIS) is a glycoprotein belonging to the TGF-β superfamily. In mammals, MIS is responsible for the regression of Müllerian ducts in the male fetus. However, the role of MIS in gonadal sex differentiation of teleost fish, which have no Müllerian ducts, has yet to be clarified. In the present study, we examined the expression pattern of mis and mis type 2 receptor (misr2) mRNAs and the function of MIS signaling in early gonadal differentiation in medaka (teleost, Oryzias latipes). In situ hybridization showed that both mis and misr2 mRNAs were expressed in the somatic cells surrounding the germ cells of both sexes during early sex differentiation. Loss-of-function of either MIS or MIS type II receptor (MISRII) in medaka resulted in suppression of germ cell proliferation during sex differentiation. These results were supported by cell proliferation assay using 5-bromo-2′-deoxyuridine labeling analysis. Treatment of tissue fragments containing germ cells with recombinant eel MIS significantly induced germ cell proliferation in both sexes compared with the untreated control. On the other hand, culture of tissue fragments from the MIS- or MISRII-defective embryos inhibited proliferation of germ cells in both sexes. Moreover, treatment with recombinant eel MIS in the MIS-defective embryos dose-dependently increased germ cell number in both sexes, whereas in the MISRII-defective embryos, it did not permit proliferation of germ cells. These results suggest that in medaka, MIS indirectly stimulates germ cell proliferation through MISRII, expressed in the somatic cells immediately after they reach the gonadal primordium.

Transcription of the rat and mouse proenkephalin genes is initiated at distinct sites in spermatogenic and somatic cells

1990 ◽  
Vol 10 (7) ◽  
pp. 3717-3726
Author(s):  
D L Kilpatrick ◽  
S A Zinn ◽  
M Fitzgerald ◽  
H Higuchi ◽  
S L Sabol ◽  
...  
Keyword(s):  
Germ Cell ◽  
Somatic Cells ◽  
Tata Box ◽  
Spermatogenic Cell ◽  
Spermatogenic Cells ◽  
Consensus Binding Site ◽  
Base Pairs ◽  
Promoter Motif ◽  
Specific Manner ◽  
Rna Protection

During spermatogenesis, several genes are expressed in a germ cell-specific manner. Previous studies have demonstrated that rat and mouse spermatogenic cells produce a 1,700-nucleotide proenkephalin RNA, while somatic cells that express the proenkephalin gene contain a 1,450-nucleotide transcript. Using cDNA cloning, RNA protection, and primer extension analyses, we showed that transcription of the rat and mouse spermatogenic-cell RNAs is initiated downstream from the proenkephalin somatic promoter in the first somatic intron (intron As). In both species, the germ cell cap site region consists of multiple start sites distributed over a length of approximately 30 base pairs. Within rat and mouse intron As, the region upstream of the germ cell cap sites is GC rich and lacks TATA sequences. A consensus binding site for the transcription factor SP1 was identified in intron As downstream of the proenkephalin germ cell cap site region. These features are characteristic of several previously described promoters that lack TATA sequences. Homologies were also identified between the proenkephalin and rat cytochrome c spermatogenic-cell promoters, including the absence of a TATA box, a multiple start site region, and several common sequences. This promoter motif thus may be shared with other genes expressed in male germ cells.

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