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At least 343 records · Page 19Linked to original sources
PubMed2004

The scrambled actin I gene in Uroleptus pisces.

The micronuclear gene encoding actin I in Uroleptus pisces occurs in two segments. Segment I contains 638 bp divided into six macronuclear destined subsegments, or MDSs, by five internal eliminated segments, or IESs. The MDSs in segment 1 are in the scrambled disorder, 1-2-4-8-6-15, with MDSs 8 and 6 inverted. Segment II contains 2452 bp divided into ten MDSs by nine IESs in the scrambled disorder, 3-5-7-10-13-12-9-14-16-11, with MDSs 12, 9, and 11 inverted. Extensive attempts by polymerase chain reaction to connect the two segments failed. We conclude that the two segments are separated by a very long IES or are in different loci. The pattern of the 16 scrambled MDSs is entirely different from the scrambled pattern observed for the actin I gene in six other stichotrichs. We conclude that the actin I gene became scrambled on two separate occasions during stichotrich evolution: once in the lineage leading to the group of six stichotrichs, which includes, among others, Sterkiella species and Stylonychia lemnae, and once in the lineage leading to Uroleptus pisces. Repeated sequence pairs (pointers) of three to 14 bases at the ends of MDSs appear to be essential for correct splicing of MDSs during macronuclear development. However, the micronuclear actin gene also contains 40 matches of eight or more bases between IESs and MDSs that do not function as pointers. To prevent these ectopic repeats from causing improper processing of the micronuclear gene appears to demand a template of DNA or RNA from the old macronucleus to guide splicing of MDSs in the orthodox order.

Actins↗
PubMed1997

The intranuclear organization of normal, hemizygous and excision-deficient rRNA genes during developmental amplification in Tetrahymena thermophila.

In the ciliated protozoan, Tetrahymena thermophila, the diploid germinal micronucleus contains two allelic copies of the gene for ribosomal RNA (rDNA). During genesis of new somatic macronuclei the germline rDNA gene is excised by developmentally programmed chromosome breakage and preferentially amplified to approximately 9, 000 copies. We have studied this process by fluorescence in situ hybridization. We find that initially rDNA amplification is restricted to two separate and highly confined regions of the nucleus. Analysis of nuclei that are hemizygous for the rDNA locus reveals that each focus of hybridization is derived from a single allele of the rDNA. As rDNA amplification progresses these two foci of hybridization disperse and spread throughout the macronucleus, eventually forming approximately 100-500 new nucleoli. These events are correlated with morphologically distinct developmental stages. We investigated the amplification of the C3 allele of the rDNA that confers a replication advantage over the B allele during vegetative propagation, and find no evidence for preferential amplification of the C3 early in rDNA maturation. We also show that the rmm 11 rDNA mutant allele, which is defective for developmentally programmed rDNA excision, can be amplified during the two-foci stage in mutant homozygotes and heterozygotes, but fails to amplify further and disperse into multiple nucleoli. These data indicate that amplification of the rmm 11 allele is not delayed during the initial rounds of amplification, and suggest that efficient excision is not required for this amplification to occur. We propose that rDNA amplification is a two-step process. First, the two rDNA alleles are independently amplified, while allelic copies remain closely associated. Later, copies of the rDNA disperse and are further amplified, presumably because rDNA excision has occurred, generating fully mature rDNA minichromosomes that are able to replicate to high copy number.

Alleles↗
PubMed2000

First record of Cycloposthium edentatum Strelkow, 1928 from the black-striped wallaby, Macropus dorsalis.

The cycloposthiid ciliate Cycloposthium edentatum Strelkow 1928 is redescribed from the stomach content of the black-striped wallaby, Macropus dorsalis (Gray) following protargol staining and light and electron microscopy. This is the first record of a cycloposthiid ciliate occurring in a marsupial, as all previous records have been restricted to eutherian herbivores, especially equids. C. edentatum was found in 7 (39%) of 18 black-striped wallabies examined, but not in any of 263 animals belonging to another 20 macropodid species examined. C. edentatum is distinguished by the form of the macronucleus, the presence of a longitudinal pellicular groove, a broad posterior tail and pellicular patterning. C. edentatum specimens recovered from M. dorsalis were smaller than those reported from horses, being up to 50% shorter and 42% thinner.

Animals↗
PubMed1975

DNA-dependent DNA polymerase activities from Paramecia macronuclei.

Three DNA-dependent DNA polymerase activities, A, B, and C, have been detected from the macronucleus of Paramecium. The enzymes were separated and partially purified by DEAE-cellulose and phosphocellulose chromatography. These three enzymes can be further distinguished from each other by the effect of KCl, araCTP and different DNA templates on their activities. All three enzymes have the same molecular weight (900 000-110 000), are dependent on Mg2+ and added DNA template and are inhibited by sodium pyrophosphate. The relative amounts of the three enzyme activities vary with the growth stage of the Paramecia from which they are isolated. In balanced growth, three polymerase activities (A, B AND C) are observed, whereas in late log or stationary phase polymerase C is absent, while the level of polymerase B decreases by a larger factor than polymerase A. These results have been related to the possible functional roles of the three activities and their relationship to activities in other organisms studies is discussed.

Animals↗
PubMed1984

Histone rearrangements accompany nuclear differentiation and dedifferentiation in Tetrahymena.

Histone synthesis and deposition into specific classes of nuclei has been investigated in starved and conjugating Tetrahymena. During starvation and early stages of conjugation (between 0 and 5 hr after opposite mating types are mixed), micronuclei selectively lose preexisting micronuclear-specific histones alpha, beta, gamma, and H3F. Of these histones, only alpha appears to accumulate in micronuclear chromatin through active synthesis and deposition during the mating process. Curiously, alpha is not observed (by stain or label) in young macronuclear anlagen (4C, 10 hr of conjugation). Thus, young macronuclear anlagen are missing all of the histones which are known to be specific to micronuclei of vegetative cells. By 14-16 hr of conjugation, we observe active synthesis and deposition of macronuclear-specific histones, hv1, hv2, and H1, into new macronuclear anlagen (8C). Thus macronuclear differentiation seems well underway by this time of conjugation. It is also in this time period (14-16 hr) that we first detect significant amounts of micronuclear-specific H1-like polypeptides beta and gamma in micronuclear extracts. These polypeptides do not seem to be synthesized during this period, which suggests that beta and gamma are derived from a precursor molecule(s). Since these micronuclear-specific histones do not appear in micronuclear chromatin until after other micronuclei have been selected to differentiate as macronuclei, we suspect that micronuclear differentiation is also an important process which occurs in 10-16 hr mating cells. Our results also suggest that proteolytic processing of micronuclear H3S into H3F (which occurs in a cell cycle dependent fashion during vegetative growth) is not operative during most if not all of conjugation. Thus micronuclei of mating cells contain only H3S which also seems consistent with the fact that some micronuclei differentiate into new macronuclei (micronuclear H3S is indistinguishable from macronuclear H3). Interestingly, the only H3 synthesized and deposited into the former macronucleus of mating cells is the relatively minor macronuclear-specific H3-like variant, hv2. These results demonstrate that significant histone rearrangements occur during conjugation in Tetrahymena in a manner consistent with the fact that during conjugation some micronuclei eventually differentiate into new macronuclei. Our results suggest that selective synthesis and deposition of specific histones (and histone variants) plays an important role in the nuclear differentiation process in Tetrahymena.

Animals↗
PubMed1986

Developmental regulation of gene expression in Tetrahymena.

The onset of gene expression in Tetrahymena thermophila during macronuclear differentiation was investigated by assay of galactokinase in conjugating deoxygalactose-resistant heterokaryons. Our results distinguish three successive states of galactokinase gene expression for cells developing a new macronucleus: stage 0, refractory to induction; stage 1, inducible by refeeding; and stage 2, induced. The refractory period ends at 12 to 13 hr after the onset of conjugation; this corresponds to the time of pair separation, and occurs several hours after the new macronuclei have become morphologically distinguishable. Stage 1 cells behave indistinguishably from mature starved cells. Inhibitor studies suggest that galactokinase synthesis is induced coincidentally with the induction of bulk protein synthesis during conjugation: thus it behaves developmentally like a typical protein; and that galactokinase mRNA is probably transcribed within 1 hr prior to its translation. Thus, when conjugating cells are refed during the refractory period, some developmental condition prevents the swift induction of protein (and galactokinase) synthesis observed upon refeeding starved (nonmating) cells. The possible nature of this developmental phenomenon is discussed.

Animals↗
PubMed1985

A transmissible developmental block in Tetrahymena thermophila.

When the amicronucleate mutant BI3840 of Tetrahymena thermophila is mated with normal micronucleate cells, it receives a pronucleus from its partner but there is no further nuclear development and the conjugants separate, retaining their original macronuclei. Both of these sexually mature exconjugants and any cells with which they are mated show an unconditional block in macronuclear development. Although prezygotic nuclear divisions, nuclear transfer and post-zygotic nuclear divisions appeared normal upon cytological analysis of Giemsa-stained conjugants, macronuclear development was invariably aborted. Since the original macronucleus was resorbed, the cells were rendered amacronucleate and they died. When no macronuclear development was initiated, as in crosses with the aneuploid strain A* (III), the exconjugants were viable and retained their original macronuclei. This pattern was invariant with three different strains serving as the original micronuclear source, and in the case of one non-BI3840 exconjugant, persisted for over 200 cell generations. Exconjugants from a cross of one of the micronuclear donors with strain A* (III) did not show arrested development when crossed. It thus appears likely that there is conjugal transfer of non-nuclear information originating in BI3840 which is self-replicating and which causes an arrest in macronuclear development.

Animals↗
PubMed1987

Nucleotide sequence and expression of two beta-tubulin genes in Stylonychia lemnae.

The gene-sized macronuclear DNA of the hypotrichous ciliate Stylonychia lemnae contains one size class of DNA molecules of 1.85 kb (1 kb = 10(3) base-pairs) coding for beta-tubulin. These DNA molecules consist of two different beta-tubulin genes, beta 1 and beta 2, which are amplified to about 150,000 (beta 1) and 30,000 (beta 2) copies per macronucleus. Both genes were cloned and sequenced entirely. The coding sequences of the two molecules (1329 base-pairs including TGA) predict identical amino acid sequences for the proteins and show a nucleotide homology of 97.2%. The nucleotide as well as the encoded amino acid sequences are highly conserved, when compared to beta-tubulin genes from vertebrates. The ciliate-specific codon TAA specifying glutamine is present only in the beta 2-tubulin gene, whereas glutamine is encoded soley by CAA in the beta 1-tubulin gene. The 5' and 3'-non-coding regions of both beta-tubulin genes are similar in length, but differ extremely in nucleotide sequence. Both beta-tubulin genes are transcriptionally active in S. lemnae, although not all putative transcription-regulatory sequences known from higher eukaryotes can be detected within the non-coding regions. The two transcription products localized by S1-mapping experiments show a similar length of about 1.40 kb and transcription seems to be regulated differently for beta 1 and beta 2.

Amino Acid Sequence↗
PubMed1979

Germinal aging in Tetrahymena thermophila.

The manifestations of germinal aging in the ciliate Tetrahymena thermophila include death of the cells at conjugation and macronuclear retention in which the normal replacement of the old macronucleus by a new one fails to occur. Available data suggest that methods of routine maintenance that reduce the number of fissions may delay aging. Differences in breeding performance following maintenance for 1-5 years in axenic peptone broth vs. bacterized Cerophyl were not significant; those following maintenance at different temperatures were significant. The analysis of several hundred crosses is consistent with a random mutational basis for aging in the micronucleus and does not support the hypothesis of an age-correlated program in which the rate of deterioration increases with time. Following routine cultural maintenance for as long as 9-11 years, some lines show no deficiencies in their ability to produce viable progeny, and sublines of the same clone frequently differ significantly in their breeding performance. Moreover, breeding degeneration occurs at constant, but different, rates in different inbred strains.

Animals↗
PubMed1981

Nucleolar changes in aging and autogamous Paramecium tetraurelia.

A significant loss in nucleolar volume density (proportion of nuclear volume occupied by nucleoli) occurs in the macronucleus as clonal age increases, which suggests an age-correlated loss of ribosomal RNA synthesizing capacity. In the macronuclear fragments of cells undergoing autogamy, however, a significant gain in nucleolar volume density takes place.

Aging↗
PubMed1984

Clonal aging in Paramecium tetraurelia. Absence of evidence for a cytoplasmic factor.

Cells of Paramecium tetraurelia show clonal aging with characteristics comparable to the aging seen in tissue culture cell lines. An investigation is underway to determine the relative contributions, if any, of the cytoplasm and the macronucleus to the onset of senescence. Using a microinjection protocol, the importance of the cytoplasm to aging was tested. Cytoplasm was transferred from young cells into old cells to see if the mean age of death of the injected cell lines could be increased, compared with uninjected controls and sham injected controls. Cytoplasm from old cells was transferred into young cells to see if the mean age of death of the injected cell lines could be decreased, compared with sham injected and uninjected controls. In neither case was there any statistically significant change in mean ages of death. Furthermore, no changes can be demonstrated even with a protocol which transplanted 3 times the normal cytoplasmic volume. Thus, a cytoplasmic effect upon vegetative aging in P. tetraurelia cannot be demonstrated. Tests of a macronuclear influence are now underway.

Aging↗
PubMed1984

Resistance of germinal nucleus to aging in Paramecium: evidence obtained by micronuclear transplantation.

A diversity of time when increase in mortality after conjugation occurs during the lifespan was found in subclones of three stocks of Paramecium caudatum. A possible micronuclear contribution to the increase in sterility has been investigated by micronuclear transplantation. We found two classes of micronuclei in aged clones: those that can function normally if the cytoplasmic environment is young, and those that cannot, even in a young cytoplasmic environment. The results indicate that in the former, the age-dependent increase in sterility is due to a deleterious macronucleus and/or cytoplasm, and in the latter it is due to micronuclear damage. The micronuclear damage in aged clones is probably induced by a deleterious cytoplasmic environment because aged clones with transplanted young micronuclei showed an abrupt decrease in progeny survival between 14 and 42 cell divisions after transplantation. Overall, the micronucleus seems not to be a source of age-related damage but rather is subjected to damage from macronuclear and/or cytoplasmic sources.

Animals↗
PubMed1979

An intervening sequence in the gene coding for 25S ribosomal RNA of Tetrahymena pigmentosa.

An intervening sequence of approximately 400 bp has been discovered in the 25S ribosomal RNA gene (rDNA) from macronuclei of Tetrahymena pigmentosa strain 6UM. Another strain, 8ALP, lacks the intervening sequence in its macronuclear rDNA. The presence of the intervening sequence was first suggested by a comparison of the restriction endonuclease maps of the rDNA from each strain. A region which differed in size between the two strains was shown to lie within the region coding for 25S ribosomal RNA by hybridization of the restriction fragments to labeled 25S rRNA. Direct visual evidence for the existence of the intervening sequence in 6UM and its absence from 8ALP was obtained by electron microscopy of R loop hybrids formed between the rDNA and 25S rRNA. The size and position of the intervening sequence within the gene were further defined by detailed restriction analyses of cloned fragments of the rDNA from each strain. The rDNA within the macronucleus of each of these strains is homogeneous, and thus the intervening sequence in the rDNA of strain 6UM must not interfere with transcription of the gene. Furthermore, the presence or absence of the intervening sequence has no obvious functional importance, since both strains can survive and grow equally well.

Base Sequence↗
PubMed1979

The intervening sequence in the 26S rRNA coding region of T. thermophila is transcribed within the largest stable precursor for rRNA.

We studied the transcription of the intervening sequence in the 26S rRNA coding region of the extrachromosomal rDNA molecules in the macronucleus of T. thermophila by hybridization of purified nuclear rRNA precursors or cytoplasmic 26S rRNA to purified native rDNA or specific rDNA restriction fragments. Examination of R loop hybrids in the electron microscope and analyses of S1-protected rDNA fragments in alkaline agarose gels showed that mature 26S rRNA, nuclear pre-26S rRNA and a fraction of the pre-rRNA molecules containing both the sequences for 17S and 26S rRNA all lack the region corresponding to the intervening sequence. The rest of the pre-rRNA molecules, however, hybridize in a colinear fashion to the whole coding region, and thus must contain the intervening sequence. We can conclude from these results that the intervening sequence is transcribed within the primary transcription product of the rDNA, and that the post-transcriptional removal of the intervening RNA sequence is a very early processing event in the organism.

Animals↗
PubMed1982

A conserved histone variant enriched in nucleoli of mammalian cells.

An antibody has been raised to Tetrahymena histone variant hv1 that specifically stains the macronucleus, but not the micronucleus, of Tetrahymena. This antiserum also stains small punctate regions in nucleoli of several mammalian cell lines. These observations suggest that this histone variant has been highly conserved in evolution and may be associated with transcribed sequences.

Animals↗
PubMed1982

Tandemly repeated hexanucleotide at Tetrahymena rDNA free end is generated from a single copy during development.

The linear extrachromosomal ribosomal DNA of Tetrahymena is generated from a single integrated copy during macronuclear development. The free ends of this extrachromosomal gene contain 20-70 tandem repeats of the hexanucleotide CCCCAA. We have determined the nucleotide sequence at the same (3') end of the single, integrated micronuclear gene. In contrast to the extrachromosomal gene, only a single CCCCAA sequence was found at this position. The same result was obtained from two independently isolated DNA clones, and was therefore not likely an artifact of cloning. Comparisons of the genomic DNA with the cloned fragment by Southern hybridization also supported this argument. Thus the tandemly repetitive hexanucleotide at the free ends of the extrachromosomal rDNA is not an inherited feature, and must be generated during the development of the macronucleus.

Animals↗
PubMed1984

Internal sequences are eliminated from genes during macronuclear development in the ciliated protozoan Oxytricha nova.

During the life cycle of the hypotrichous ciliate Oxytricha nova, a macronucleus containing short, gene-sized DNA molecules is produced from a copy of the chromosomal micronuclear genome. In order to characterize the process of macronuclear development, we have isolated and determined the DNA sequence of a particular macronuclear gene and its micronuclear precursor. The results of this analysis indicate that macronuclear telomeric sequences (5'C4A4(3') repeats) are not present at the ends of the gene in its micronuclear chromosomal location and must be added during development. In addition, the micronuclear copy of the gene contains three short blocks of sequence that must be removed during development, implying the involvement of a nucleic acid-splicing process in generating mature macronuclear genes.

Amino Acid Sequence↗
PubMed1984

Telomeric properties of C4A4-homologous sequences in micronuclear DNA of Oxytricha fallax.

The hypotrichous ciliated protozoan Oxytricha fallax has both macronuclei and micronuclei. The short DNA molecules of the macronucleus are terminated with repeats of the octanucleotide C4A4. Micronuclear DNA carries a family of long sequence blocks, which share a consensus restriction map and have homology with the C4A4 repeats. Each block contains a 3 to 6 kb subregion, which is refractory to all restriction enzymes tested and includes the C4A4 homology. We conclude that these refractory sequences are at DNA termini because Sau 3A fragments containing them fail to ligate to form circles, and they are rapidly degraded by Bal 31 nuclease. These terminal C4A4-homologous sequence blocks likely represent micronuclear chromosomal telomeres. Possible relationships between these sequences, internal micronuclear C4A4 repeats, and macronuclear DNA termini are discussed.

Animals↗