scholarly journals Mapping the Germ-Line and Somatic Genomes of a Ciliated Protozoan,Tetrahymena thermophila

1998 ◽  
Vol 8 (2) ◽  
pp. 91-99 ◽  
Author(s):  
Eduardo Orias
Keyword(s):  
Tetrahymena Thermophila ◽  
Germ Line ◽  
Ciliated Protozoan

scholarly journals A method for mapping germ line sequences in Tetrahymena thermophila using the polymerase chain reaction.

Genetics ◽  
1994 ◽  
Vol 137 (1) ◽  
pp. 95-106 ◽  
Author(s):  
D Cassidy-Hanley ◽  
M C Yao ◽  
P J Bruns
Keyword(s):  
Polymerase Chain Reaction ◽  
Dna Sequences ◽  
Tetrahymena Thermophila ◽  
Germ Line ◽  
Repetitive Sequences ◽  
Mapping Technique ◽  
Ciliated Protozoan ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Template Dna

Abstract A method for mapping DNA sequences to specific germinal chromosomes in the ciliated protozoan Tetrahymena thermophila has been developed. This mapping technique (PCR mapping) utilizes the polymerase chain reaction and template DNA derived from nullisomic strains to directly assign micronuclear DNA sequences to specific micronuclear chromosomes. Using this technique, a number of unique sequences and short repetitive sequences flanked by unique sequences have been mapped to four of the five germinal chromosomes.

scholarly journals Germ line-specific DNA sequences are present on all five micronuclear chromosomes in Tetrahymena thermophila.

1983 ◽  
Vol 3 (11) ◽  
pp. 1909-1919 ◽  
Author(s):  
K M Karrer
Keyword(s):  
In Situ Hybridization ◽  
Dna Sequences ◽  
Tetrahymena Thermophila ◽  
Germ Line ◽  
Plasmid Vector ◽  
Ciliated Protozoan ◽  
Specific Sequence ◽  
Individual Probe ◽  
Vector Pbr322

The development of the macronucleus from the zygotic micronucleus in the ciliated protozoan Tetrahymena spp. involves the elimination of specific DNA sequences (M. C. Yao and M. Gorovsky, Chromosoma 48:1-18 1974). The present study demonstrates that micronucleus-specific DNA is present on all five of the micronuclear chromosomes. Fragments of micronuclear DNA from Tetrahymena thermophila were cloned in the plasmid vector pBR322. A procedure was developed to examine the organization of the cloned sequences in micro- and macronuclear DNA without nick translating each individual probe. Twenty-three percent of randomly selected DNA sequences examined by this method were micronucleus (germ line) specific. They were all members of families of repeated sequences. Hybridization of six micronucleus-specific DNA sequences to micronuclear DNA from nullisomic strains of T. thermophila, which are lacking one or more pairs of chromosomes in the micronucleus, suggested that these sequences are present on several chromosomes. One micronucleus-specific sequence was shown by in situ hybridization to be present on all five of the micronuclear chromosomes.

scholarly journals Germ line-specific DNA sequences are present on all five micronuclear chromosomes in Tetrahymena thermophila

1983 ◽  
Vol 3 (11) ◽  
pp. 1909-1919
Author(s):  
K M Karrer
Keyword(s):  
In Situ Hybridization ◽  
Dna Sequences ◽  
Tetrahymena Thermophila ◽  
Germ Line ◽  
Plasmid Vector ◽  
Ciliated Protozoan ◽  
Specific Sequence ◽  
Individual Probe ◽  
Vector Pbr322

The development of the macronucleus from the zygotic micronucleus in the ciliated protozoan Tetrahymena spp. involves the elimination of specific DNA sequences (M. C. Yao and M. Gorovsky, Chromosoma 48:1-18 1974). The present study demonstrates that micronucleus-specific DNA is present on all five of the micronuclear chromosomes. Fragments of micronuclear DNA from Tetrahymena thermophila were cloned in the plasmid vector pBR322. A procedure was developed to examine the organization of the cloned sequences in micro- and macronuclear DNA without nick translating each individual probe. Twenty-three percent of randomly selected DNA sequences examined by this method were micronucleus (germ line) specific. They were all members of families of repeated sequences. Hybridization of six micronucleus-specific DNA sequences to micronuclear DNA from nullisomic strains of T. thermophila, which are lacking one or more pairs of chromosomes in the micronucleus, suggested that these sequences are present on several chromosomes. One micronucleus-specific sequence was shown by in situ hybridization to be present on all five of the micronuclear chromosomes.

scholarly journals FURTHER EVIDENCE FOR LACK OF GENE EXPRESSION IN THE TETRAHYMENA MICRONUCLEUS

Genetics ◽  
1981 ◽  
Vol 98 (4) ◽  
pp. 747-762
Author(s):  
Kristen A Mayo ◽  
Eduardo Orias
Keyword(s):  
Gene Expression ◽  
Rna Synthesis ◽  
Tetrahymena Thermophila ◽  
Germ Line ◽  
Ciliated Protozoan ◽  
Quantitative Evidence ◽  
Soluble Cell ◽  
Cell Extracts ◽  
Radiometric Assay ◽  
Galactokinase Activity

ABSTRACT Certain galA mutations in the ciliated protozoan Tetrahymena thermophila confer an almost total loss of galactokinase activity in homozygotes. Heterokaryons have been constructed that are homogeneous for the galA1 mutation in the (45n) macronucleus, but which contain a galA  + (2n) micronucleus. Soluble cell extracts prepared from these heterokaryons have been assayed for galactokinase activity, using a radiometric assay for the conversion of galactose to galactose-1-phosphate (gal-1-P). No galactokinase activity attributable to the micronuclear genes is observed in such heterokaryons. These results, obtained with the galA1 marker, provide the first direct, quantitative evidence for the lack of micronuclear (germ line) gene expression in Tetrahymena during vegetative growth, and substantiate the predictions of previous phenotypic observations on heterokaryons and autoradiographic studies of micronuclear RNA synthesis. The generality of this conclusion will be established in the future when other enzymically assayable mutations become available for similar studies.

scholarly journals Role of Micronucleus-Limited DNA in Programmed Deletion of mse2.9 during Macronuclear Development of Tetrahymena thermophila

2004 ◽  
Vol 3 (2) ◽  
pp. 288-301 ◽  
Author(s):  
Jeffrey S. Fillingham ◽  
Ronald E. Pearlman
Keyword(s):  
Epigenetic Regulation ◽  
Tetrahymena Thermophila ◽  
Germ Line ◽  
Sexual Cycle ◽  
Ciliated Protozoan ◽  
Dna Rearrangements ◽  
Macronuclear Development ◽  
Dna Deletion

ABSTRACT Extensive programmed DNA rearrangements occur during the development of the somatic macronucleus from the germ line micronucleus in the sexual cycle of the ciliated protozoan Tetrahymena thermophila. Using an in vivo processing assay, we analyzed the role of micronucleus-limited DNA during the programmed deletion of mse2.9, an internal eliminated sequence (IES). We identified a 200-bp region within mse2.9 that contains an important cis-acting element which is required for the targeting of efficient programmed deletion. Our results, obtained with a series of mse2.9-based chimeric IESs, led us to suggest that the cis-acting elements in both micronucleus-limited and macronucleus-retained flanking DNAs stimulate programmed deletion to different degrees depending on the particular eliminated sequence. The mse2.9 IES is situated within the second intron of the micronuclear locus of the ARP1 gene. We show that the expression of ARP1 is not essential for the growth of Tetrahymena. Our results also suggest that mse2.9 is not subject to epigenetic regulation of DNA deletion, placing possible constraints on the scan RNA model of IES excision.

scholarly journals A Non-Long Terminal Repeat Retrotransposon Family Is Restricted to the Germ Line Micronucleus of the Ciliated Protozoan Tetrahymena thermophila

2004 ◽  
Vol 3 (1) ◽  
pp. 157-169 ◽  
Author(s):  
Jeffrey S. Fillingham ◽  
Trine A. Thing ◽  
Nama Vythilingum ◽  
Alex Keuroghlian ◽  
Deanna Bruno ◽  
...  
Keyword(s):  
Long Terminal Repeat ◽  
Tetrahymena Thermophila ◽  
Germ Line ◽  
Terminal Repeat ◽  
Sexual Cycle ◽  
Northern Analysis ◽  
Ciliated Protozoan ◽  
Dna Rearrangements ◽  
Retrotransposon Family ◽  
The Relationship

ABSTRACT The ciliated protozoan Tetrahymena thermophila undergoes extensive programmed DNA rearrangements during the development of a somatic macronucleus from the germ line micronucleus in its sexual cycle. To investigate the relationship between programmed DNA rearrangements and transposable elements, we identified several members of a family of non-long terminal repeat (LTR) retrotransposons (retroposons) in T. thermophila, the first characterized in the ciliated protozoa. This multiple-copy retrotransposon family is restricted to the micronucleus of T. thermophila. The REP (Tetrahymena non-LTR retroposon) elements encode an ORF2 typical of non-LTR elements that contains apurinic/apyrimidinic endonuclease (APE) and reverse transcriptase (RT) domains. Phylogenetic analysis of the RT and APE domains indicates that the element forms a deep-branching clade within the non-LTR retrotransposon family. Northern analysis with a probe to the conserved RT domain indicates that transcripts from the element are small and heterogeneous in length during early macronuclear development. The presence of a repeated transposable element in the genome is consistent with the model that programmed DNA deletion in T. thermophila evolved as a method of eliminating deleterious transposons from the somatic macronucleus.

scholarly journals A Developmentally Regulated Gene, ASI2, Is Required for Endocycling in the Macronuclear Anlagen of Tetrahymena

2010 ◽  
Vol 9 (9) ◽  
pp. 1343-1353 ◽  
Author(s):  
Lihui Yin ◽  
Susan T. Gater ◽  
Kathleen M. Karrer
Keyword(s):  
Signal Transduction ◽  
Mitotic Cycle ◽  
Tetrahymena Thermophila ◽  
Germ Line ◽  
Ciliated Protozoa ◽  
Dna Rearrangement ◽  
Developmentally Regulated ◽  
Content Type ◽  
Molecular Events ◽  
Multicellular Organisms

ABSTRACT Ciliated protozoa contain two types of nuclei, germ line micronuclei (Mic) and transcriptionally active macronuclei (Mac). During sexual reproduction, the parental Mac degenerates and a new Mac develops from a mitotic product of the zygotic Mic. Macronuclear development involves extensive endoreplication of the genome. The present study shows that endoreplication of macronuclear DNA in Tetrahymena is an example of endocyling, a variant of the mitotic cycle with alternating S and G phases in the absence of cell division. Thus, endocycling is conserved from ciliates to multicellular organisms. The gene ASI2 in Tetrahymena thermophila encodes a putative signal transduction receptor. ASI2 is nonessential for vegetative growth, but it is upregulated during development of the new Mac. Cells that lack ASI2 in the developing Mac anlagen are arrested in endoreplication of the DNA and die. This study shows that ASI2 is also transcribed in the parental Mac early in conjugation and that transcription of ASI2 in the parental Mac supports endoreplication of the DNA during early stages of development of the Mac anlagen. Other molecular events in Mac anlage development, including developmentally regulated DNA rearrangement, occur normally in matings between ASI2 knockouts, suggesting that ASI2 specifically regulates endocycling in Tetrahymena.

scholarly journals Constitutive expression, not a particular primary sequence, is the important feature of the H3 replacement variant hv2 in Tetrahymena thermophila.

1997 ◽  
Vol 17 (11) ◽  
pp. 6303-6310 ◽  
Author(s):  
L Yu ◽  
M A Gorovsky
Keyword(s):  
Protein Sequence ◽  
Tetrahymena Thermophila ◽  
Histone H3 ◽  
Constitutive Expression ◽  
Histone Variants ◽  
Wild Type ◽  
Ciliated Protozoan ◽  
Knockout Mutant ◽  
Double Knockout ◽  
Constitutive Synthesis

Although quantitatively minor replication-independent (replacement) histone variants have been found in a wide variety of organisms, their functions remain unknown. Like the H3.3 replacement variants of vertebrates, hv2, an H3 variant in the ciliated protozoan Tetrahymena thermophila, is synthesized and deposited in nuclei of nongrowing cells. Although hv2 is clearly an H3.3-like replacement variant by its expression, sequence analysis indicates that it evolved independently of the H3.3 variants of multicellular eukaryotes. This suggested that it is the constitutive synthesis, not the particular protein sequence, of these variants that is important in the function of H3 replacement variants. Here, we demonstrate that the gene (HHT3) encoding hv2 or either gene (HHT1 or HHT2) encoding the abundant major H3 can be completely knocked out in Tetrahymena. Surprisingly, when cells lacking hv2 are starved, a major histone H3 mRNA transcribed by the HHT2 gene, which is synthesized little, if at all, in wild-type nongrowing cells, is easily detectable. Both HHT2 and HHT3 knockout strains show no obvious defect during vegetative growth. In addition, a mutant with the double knockout of HHT1 and HHT3 is viable while the HHT2 HHT3 double-knockout mutant is not. These results argue strongly that cells require a constitutively expressed H3 gene but that the particular sequence being expressed is not critical.

Structure and expression of two temperature-specific surface proteins in the ciliated protozoan Tetrahymena thermophila

1986 ◽  
Vol 6 (9) ◽  
pp. 3240-3245
Author(s):  
G A Bannon ◽  
R Perkins-Dameron ◽  
A Allen-Nash
Keyword(s):  
Cell Surface ◽  
Specific Surface ◽  
Incubation Temperature ◽  
Tetrahymena Thermophila ◽  
Sodium Dodecyl ◽  
Surface Protein ◽  
Surface Proteins ◽  
Ciliated Protozoan ◽  
Temperature Dependent ◽  
Molecular Sizes

The presence of specific proteins (known as immobilization antigens) on the surface of the ciliated protozoan Tetrahymena thermophila is under environmental regulation. There are five different classes (serotypes) of surface proteins which appear on the cell surface when T. thermophila is cultured under different conditions of temperature or incubation medium; three of these are temperature dependent. The appearance of these proteins on the cell surface is mutually exclusive. We used polyclonal antibodies raised against 30 degrees C (designated SerH3)- and 40 degrees C (designated SerT)-specific surface antigens to study their structure and expression. We showed that these surface proteins contain at least one disulfide bridge. On sodium dodecyl sulfate-denaturing polyacrylamide gels, the nonreduced 30 degrees C- and 40 degrees C-specific surface proteins migrated with molecular sizes of 69 and 36 kilodaltons, respectively. The reduced forms of the proteins migrated with molecular sizes of 58 and 30 kilodaltons, respectively. The synthesis of the surface proteins responded rapidly and with a time course similar to that of the incubation temperature. The synthesis of each surface protein was greatly reduced within 1 h and undetectable by 2 h after a shift to the temperature at which the protein is not expressed. Surface protein synthesis resumed by the end of 1 h after a shift to the temperature at which the protein is expressed. The temperature-dependent induction of these surface proteins appears to be dependent on the synthesis of new mRNA, as indicated by a sensitivity to actinomycin D. Surface protein syntheses were mutually exclusive except at a transition temperature. At 35 degrees C both surface proteins were synthesized by a cell population. These data support the potential of this system as a model for the study of the effects of environmental factors on the genetic regulation of cell surface proteins.

scholarly journals Site-specific methylation of adenine in the nuclear genome of a eucaryote, Tetrahymena thermophila.

1986 ◽  
Vol 6 (7) ◽  
pp. 2364-2370 ◽  
Author(s):  
G S Harrison ◽  
R C Findly ◽  
K M Karrer
Keyword(s):  
Heat Shock ◽  
Protein Gene ◽  
De Novo ◽  
Tetrahymena Thermophila ◽  
Nuclear Genome ◽  
Histone H4 ◽  
Ciliated Protozoan ◽  
Site Specific ◽  
Logarithmic Growth ◽  
I Gene

DNA in the polyploid macronucleus of the ciliated protozoan Tetrahymena thermophila contains the modified base N6-methyladenine. We identified two GATC sites which are methylated in most or all of the 45 copies of the macronuclear genome. One site is 2 kilobases 5' to the histone H4-I gene, and the other is 5 kilobases 3' to the 73-kilodalton heat shock protein gene. These sites are de novo methylated between 10 and 16 h after initiation of conjugation, during macronuclear anlage development. The methylation states of these two GATC sites and four other unmethylated GATC sites do not change in the DNA of cells cultured under conditions which change the activity of the genes, including logarithmic growth, starvation, and heat shock.

Sign in / Sign up

Export Citation Format

Share Document