A self-encoded capsid derivative restricts Ty1 retrotransposition in Saccharomyces

ABSTRACT Ty1 reverse transcriptase/RNase H (RT/RH) is exquisitely sensitive to manganese concentrations. Elevated intracellular free Mn2+ inhibits Ty1 retrotransposition and in vitro Ty1 RT-polymerizing activity. Furthermore, Mn2+ inhibition is not limited to the Ty1 RT, as this ion similarly inhibits the activities of both avian myeloblastosis virus and human immunodeficiency virus type 1 RTs. To further characterize Mn2+ inhibition, we generated RT/RH suppressor mutants capable of increased Ty1 transposition in pmr1Δ cells. PMR1 codes for a P-type ATPase that regulates intracellular calcium and manganese ion homeostasis, and pmr1 mutants accumulate elevated intracellular manganese levels and display 100-fold less transposition than PMR1 + cells. Mapping of these suppressor mutations revealed, surprisingly, that suppressor point mutations localize not to the RT itself but to the RH domain of the protein. Furthermore, Mn2+ inhibition of in vitro RT activity is greatly reduced in all the suppressor mutants, whereas RH activity and cleavage specificity remain largely unchanged. These intriguing results reveal that the effect of these suppressor mutations is transmitted to the polymerase domain and suggest biochemical communication between these two domains during reverse transcription.

Download Full-text

Post-transcriptional Cosuppression of Ty1 Retrotransposition

Genetics ◽

10.1093/genetics/165.1.83 ◽

2003 ◽

Vol 165 (1) ◽

pp. 83-99

Author(s):

David J Garfinkel ◽

Katherine Nyswaner ◽

Jun Wang ◽

Jae-Yong Cho

Keyword(s):

Gene Silencing ◽

Reverse Transcription ◽

Copy Number ◽

Transcript Level ◽

Gag Protein ◽

Protein Levels ◽

Copy Number Control ◽

Gal1 Promoter ◽

Pgk1 Promoter ◽

Ty1 Retrotransposition

Abstract To determine whether homology-dependent gene silencing or cosuppression mechanisms underlie copy number control (CNC) of Ty1 retrotransposition, we introduced an active Ty1 element into a naïve strain. Single Ty1 element retrotransposition was elevated in a Ty1-less background, but decreased dramatically when additional elements were present. Transcription from the suppressing Ty1 elements enhanced CNC but translation or reverse transcription was not required. Ty1 CNC occurred with a transcriptionally active Ty2 element, but not with Ty3 or Ty5 elements. CNC also occurred when the suppressing Ty1 elements were transcriptionally silenced, fused to the constitutive PGK1 promoter, or contained a minimal segment of mostly TYA1-gag sequence. Ty1 transcription of a multicopy element expressed from the GAL1 promoter abolished CNC, even when the suppressing element was defective for transposition. Although Ty1 RNA and TyA1-gag protein levels increased with the copy number of expressible elements, a given element's transcript level varied less than twofold regardless of whether the suppressing elements were transcriptionally active or repressed. Furthermore, a decrease in the synthesis of Ty1 cDNA is strongly associated with Ty1 CNC. Together our results suggest that Ty1 cosuppression can occur post-transcriptionally, either prior to or during reverse transcription.

Download Full-text

Posttranslational Regulation of Ty1 Retrotransposition by Mitogen-Activated Protein Kinase Fus3

Molecular and Cellular Biology ◽

10.1128/mcb.18.9.5620-a ◽

1998 ◽

Vol 18 (9) ◽

pp. 5620-5620

Author(s):

Darryl Conte ◽

Ellen Barber ◽

Mukti Banerjee ◽

David J. Garfinkel ◽

M. Joan Curcio

Keyword(s):

Protein Kinase ◽

Mitogen Activated Protein Kinase ◽

Posttranslational Regulation ◽

Mitogen Activated Protein ◽

Ty1 Retrotransposition

Download Full-text

Erratum for Saha et al., Atrans-Dominant Form of Gag Restricts Ty1 Retrotransposition and Mediates Copy Number Control

Journal of Virology ◽

10.1128/jvi.00418-16 ◽

2016 ◽

Vol 90 (10) ◽

pp. 5210-5210

Author(s):

Agniva Saha ◽

Jessica A. Mitchell ◽

Yuri Nishida ◽

Jonathan E. Hildreth ◽

Joshua A. Arribere ◽

...

Keyword(s):

Copy Number ◽

Dominant Form ◽

Copy Number Control ◽

Ty1 Retrotransposition

Download Full-text

Posttranslational control of Ty1 retrotransposition occurs at the level of protein processing.

Molecular and Cellular Biology ◽

10.1128/mcb.12.6.2813 ◽

1992 ◽

Vol 12 (6) ◽

pp. 2813-2825 ◽

Cited By ~ 43

Author(s):

M J Curcio ◽

D J Garfinkel

Keyword(s):

Protein Processing ◽

Yeast Cells ◽

Translational Efficiency ◽

Gene Products ◽

Protease Domain ◽

High Level Expression ◽

Gal1 Promoter ◽

Low Levels ◽

Ty1 Retrotransposition ◽

High Level

High-level expression of a transpositionally competent Ty1 element fused to the inducible GAL1 promoter on a 2 microns plasmid (pGTy1) overcomes transpositional dormancy in Saccharomyces cerevisiae. To investigate the mechanisms controlling the rate of Ty1 retrotransposition, we quantitated transposition and Ty1 gene products in cells induced and uninduced for expression of pGTy1. The increase in Ty1 transposition was 45- to 125-fold greater than the increase in Ty1 RNA effected by pGTy1 induction. Translational efficiency of Ty1 RNA was not altered in transposition-induced cells, since p190TYA1-TYB1 protein synthesis increased in proportion to steady-state Ty1 RNA levels. Therefore, expression of a pGTy1 element increases the efficiency of Ty1 transposition at a posttranslational level. Galactose induction of pGTy1 enhanced TYA1 protein processing and allowed detection of processed TYB1 proteins, which are normally present at very low levels in uninduced cells. When the ability of genomic Ty1 elements to complement defined mutations in HIS3-marked pGTy1 elements was examined, mutations in the protease domain or certain mutations in the integrase domain failed to be complemented, but mutations in the reverse transcriptase domain were partially complemented by genomic Ty1 elements. Therefore, the activity of Ty1 elements in yeast cells may be limited by the availability of Ty1 protease and possibly integrase. These results suggest that Ty1 transposition is regulated at the level of protein processing and that this regulation is overcome by expression of a pGTy1 element.

Download Full-text

A nuclear pore sub-complex restricts the propagation of Ty retrotransposons by limiting their transcription

PLoS Genetics ◽

10.1371/journal.pgen.1009889 ◽

2021 ◽

Vol 17 (11) ◽

pp. e1009889

Author(s):

Amandine Bonnet ◽

Carole Chaput ◽

Noé Palmic ◽

Benoit Palancade ◽

Pascale Lesage

Keyword(s):

Transcriptional Control ◽

Nuclear Pore ◽

Nuclear Pore Complexes ◽

Ltr Retrotransposons ◽

Loss Of Function ◽

Protein Coding ◽

Multiple Copies ◽

Ty1 Retrotransposition ◽

Pore Complexes ◽

The Impact

Beyond their canonical function in nucleocytoplasmic exchanges, nuclear pore complexes (NPCs) regulate the expression of protein-coding genes. Here, we have implemented transcriptomic and molecular methods to specifically address the impact of the NPC on retroelements, which are present in multiple copies in genomes. We report a novel function for the Nup84 complex, a core NPC building block, in specifically restricting the transcription of LTR-retrotransposons in yeast. Nup84 complex-dependent repression impacts both Copia and Gypsy Ty LTR-retrotransposons, all over the S. cerevisiae genome. Mechanistically, the Nup84 complex restricts the transcription of Ty1, the most active yeast retrotransposon, through the tethering of the SUMO-deconjugating enzyme Ulp1 to NPCs. Strikingly, the modest accumulation of Ty1 RNAs caused by Nup84 complex loss-of-function is sufficient to trigger an important increase of Ty1 cDNA levels, resulting in massive Ty1 retrotransposition. Altogether, our study expands our understanding of the complex interactions between retrotransposons and the NPC, and highlights the importance for the cells to keep retrotransposon under tight transcriptional control.

Download Full-text

Nucleotide Excision Repair/TFIIH Helicases Rad3 and Ssl2 Inhibit Short-Sequence Recombination and Ty1 Retrotransposition by Similar Mechanisms

Molecular and Cellular Biology ◽

10.1128/mcb.20.7.2436-2445.2000 ◽

2000 ◽

Vol 20 (7) ◽

pp. 2436-2445 ◽

Cited By ~ 29

Author(s):

Bum-Soo Lee ◽

Liu Bi ◽

David J. Garfinkel ◽

Adam M. Bailis

Keyword(s):

Nucleotide Excision Repair ◽

Genome Stability ◽

Synthetic Lethality ◽

Excision Repair ◽

Genome Structure ◽

Physical Stability ◽

Short Sequence ◽

Protein Levels ◽

Nucleotide Excision ◽

Ty1 Retrotransposition

ABSTRACT Eukaryotic genomes contain potentially unstable sequences whose rearrangement threatens genome structure and function. Here we show that certain mutant alleles of the nucleotide excision repair (NER)/TFIIH helicase genes RAD3 and SSL2(RAD25) confer synthetic lethality and destabilize theSaccharomyces cerevisiae genome by increasing both short-sequence recombination and Ty1 retrotransposition. Therad3-G595R and ssl2-rtt mutations do not markedly alter Ty1 RNA or protein levels or target site specificity. However, these mutations cause an increase in the physical stability of broken DNA molecules and unincorporated Ty1 cDNA, which leads to higher levels of short-sequence recombination and Ty1 retrotransposition. Our results link components of the core NER/TFIIH complex with genome stability, homologous recombination, and host defense against Ty1 retrotransposition via a mechanism that involves DNA degradation.

Download Full-text

Posttranslational Regulation of Ty1 Retrotransposition by Mitogen-Activated Protein Kinase Fus3

Molecular and Cellular Biology ◽

10.1128/mcb.18.5.2502 ◽

1998 ◽

Vol 18 (5) ◽

pp. 2502-2513 ◽

Cited By ~ 37

Author(s):

Darryl Conte ◽

Ellen Barber ◽

Mukti Banerjee ◽

David J. Garfinkel ◽

M. Joan Curcio

Keyword(s):

Protein Kinase ◽

Proteolytic Processing ◽

Mitogen Activated Protein Kinase ◽

Pheromone Response ◽

Environmental Signals ◽

Pheromone Response Pathway ◽

Mitogen Activated Protein ◽

Associated Proteins ◽

Steady State Levels ◽

Ty1 Retrotransposition

ABSTRACT Ty1 retrotransposons in Saccharomyces cerevisiae are maintained in a state of transpositional dormancy. We isolated a mutation, rtt100-1, that increases the transposition of genomic Ty1 elements 18- to 56-fold but has little effect on the transposition of related Ty2 elements. rtt100-1 was shown to be a null allele of the FUS3 gene, which encodes a haploid-specific mitogen-activated protein kinase. In fus3mutants, the levels of Ty1 RNA, protein synthesis, and proteolytic processing were not altered relative to those in FUS3strains but steady-state levels of TyA, integrase, and reverse transcriptase proteins and Ty1 cDNA were all increased. These findings suggest that Fus3 suppresses Ty1 transposition by destabilizing viruslike particle-associated proteins. The Fus3 kinase is activated through the mating-pheromone response pathway by phosphorylation at basal levels in naive cells and at enhanced levels in pheromone-treated cells. We demonstrate that suppression of Ty1 transposition in naive cells requires basal levels of Fus3 activation. Substitution of conserved amino acids required for activation of Fus3 derepressed Ty1 transposition. Moreover, epistasis analyses revealed that components of the pheromone response pathway that act upstream of Fus3, including Ste4, Ste5, Ste7, and Ste11, are required for the posttranslational suppression of Ty1 transposition by Fus3. The regulation of Ty1 transposition by Fus3 provides a haploid-specific mechanism through which environmental signals can modulate the levels of retrotransposition.

Download Full-text

A rare tRNA-Arg(CCU) that regulates Ty1 element ribosomal frameshifting is essential for Ty1 retrotransposition in Saccharomyces cerevisiae.

Genetics ◽

10.1093/genetics/135.2.309 ◽

1993 ◽

Vol 135 (2) ◽

pp. 309-320 ◽

Cited By ~ 7

Author(s):

K Kawakami ◽

S Pande ◽

B Faiola ◽

D P Moore ◽

J D Boeke ◽

...

Keyword(s):

Saccharomyces Cerevisiae ◽

Protein Processing ◽

Ribonuclease H ◽

Wild Type ◽

Ribosomal Frameshifting ◽

Translational Frameshifting ◽

Rate Limiting Step ◽

Limiting Step ◽

Ty1 Retrotransposition ◽

Rate Limiting

Abstract Translation of the yeast retrotransposon Ty1 TYA1(gag)-TYB1(pol) gene occurs by a +1 ribosomal frameshifting event at the sequence CUU AGG C. Because overexpression of a low abundance tRNA-Arg(CCU) encoded by the HSX1 gene resulted in a reduction in Ty1 frameshifting, it was suggested that a translational pause at the AGG-Arg codon is required for optimum frameshifting. The present work shows that the absence of tRNA-Arg(CCU) affects Ty1 transposition, translational frameshifting, and accumulation of mature TYB1 proteins. Transposition of genetically tagged Ty1 elements decreases at least 50-fold and translational frameshifting increases 3-17-fold in cells lacking tRNA-Arg(CCU). Accumulation of Ty1-integrase and Ty1-reverse transcriptase/ribonuclease H is defective in an hsx1 mutant. The defect in Ty1 transposition is complemented by the wild-type HSX1 gene or a mutant tRNA-Arg(UCU) gene containing a C for T substitution in the first position of the anticodon. Overexpression of TYA1 stimulates Ty1 transposition 50-fold above wild-type levels when the level of transposition is compared in isogenic hsx1 and HSX1 strains. Thus, the HSX1 gene determines the ratio of the TYA1 to TYA1-TYB1 precursors required for protein processing or stability, and keeps expression of TYB1 a rate-limiting step in the retrotransposition cycle.

Download Full-text

Ty1 Mobilizes Subtelomeric Y′ Elements in Telomerase-Negative Saccharomyces cerevisiae Survivors

Molecular and Cellular Biology ◽

10.1128/mcb.24.22.9887-9898.2004 ◽

2004 ◽

Vol 24 (22) ◽

pp. 9887-9898 ◽

Cited By ~ 30

Author(s):

Patrick H. Maxwell ◽

Candice Coombes ◽

Alison E. Kenny ◽

Joseph F. Lawler ◽

Jef D. Boeke ◽

...

Keyword(s):

Saccharomyces Cerevisiae ◽

Transcription Factor ◽

Cell Division ◽

Long Terminal Repeat ◽

Reverse Transcription ◽

Telomere Maintenance ◽

Nonpermissive Temperature ◽

Telomeric Dna ◽

Copy Numbers ◽

Ty1 Retrotransposition

ABSTRACT When telomerase is inactivated in Saccharomyces cerevisiae, telomeric DNA shortens with every cell division, and cells stop dividing after ∼100 generations. Survivors that form in these senescent populations and resume growing have variably amplified arrays of subtelomeric Y′ elements. We marked a chromosomal Y′ element with the his3AI retrotransposition indicator gene and found that Y′HIS3 cDNA was incorporated into the genome at ∼10- to 1,000-fold-higher frequencies in survivors compared to telomerase-positive strains. Y′HIS3 cDNA mobility was significantly reduced if assayed at 30°C, a nonpermissive temperature for Ty1 retrotransposition, or in the absence of Tec1p, a transcription factor for Ty1. Microarray analysis revealed that Y′ RNA is preferentially associated with Ty1 virus-like particles (VLPs). Genomic copies of Y′HIS3 cDNA typically have downstream oligo(A) tracts, followed by a complete Ty1 long terminal repeat and TYA1 or TYB1 sequences. These data are consistent with the use of Ty1 cDNA to prime reverse transcription of polyadenylated Y′ RNA within Ty1 VLPs. Unmarked Y′-oligo(A)-Ty1 cDNA was also detected in survivors, reaching copy numbers of ∼10−2 per genome. We propose that Y′-oligo(A)-Ty1 cDNA recombines with Y′ elements at eroding telomeres in survivors and may play a role in telomere maintenance in the absence of telomerase.

Download Full-text