scholarly journals Essential and nonessential histone H2A variants in Tetrahymena thermophila.

1996 ◽  
Vol 16 (8) ◽  
pp. 4305-4311 ◽  
Author(s):  
X Liu ◽  
B Li ◽  
GorovskyMA
Keyword(s):  
Tetrahymena Thermophila ◽  
Essential Gene ◽  
Histone Variants ◽  
Gene Replacement ◽  
Histone H2a ◽  
Gene Encoding ◽  
Genes Encoding ◽  
Simple Mass ◽  
Essential Function ◽  
High Degree

Although variants have been identified for every class of histone, their functions remain unknown. We have been studying the histone H2A variant hv1 in the ciliated protozoan Tetrahymena thermophila. Sequence analysis indicates that hv1 belongs to the H2A.F/Z type of histone variants. On the basis of the high degree of evolutionary conservation of this class of histones, they are proposed to have one or more distinct and essential functions that cannot be performed by their major H2A counterparts. Considerable evidence supports the hypothesis that the hv1 protein in T. thermophila and hv1-like proteins in other eukaryotes are associated with active chromatin. In T. thermophila, simple mass transformation and gene replacement techniques have recently become available. In this report, we demonstrate that either the HTA1 gene or the HTA2 gene, encoding the major H2As, can be completely replaced by disrupted genes in the polyploid, transcriptionally active macronucleus, indicating that neither of the two genes is essential. However, only some of the HTA3 genes encoding hv1 can be replaced by disrupted genes, indicating that the H2A.F/Z type variants have an essential function that cannot be performed by the major H2A genes. Thus, an essential gene in T. thermophila can be defined by the fact that it can be partially, but not completely, eliminated from the polyploid macronucleus. To our knowledge, this study represents the first use of gene disruption technology to study core histone gene function in any organism other than yeast and the first demonstration of an essential gene in T. thermophila using these methods. When a rescuing plasmid carrying a wild-type HTA3 gene was introduced into the T. thermophila cells, the endogenous chromosomal HTA3 could be completely replaced, defining a gene replacement strategy that can be used to analyze the function of essential genes.

scholarly journals Structure, expression and phylogenetic analysis of the gene encoding actin I in Pneumocystis carinii.

Genetics ◽  
1994 ◽  
Vol 137 (3) ◽  
pp. 743-750 ◽  
Author(s):  
L D Fletcher ◽  
J M McDowell ◽  
R R Tidwell ◽  
R B Meagher ◽  
C C Dykstra
Keyword(s):  
Phylogenetic Analysis ◽  
Essential Gene ◽  
Pneumocystis Carinii ◽  
Comparative Sequence Analysis ◽  
Cdna Clones ◽  
Actin Gene ◽  
Gene Encoding ◽  
Actin Protein ◽  
Relationship Of ◽  
High Degree

Abstract Actin is a major component of the cytoskeleton and one of the most abundant proteins found in eukaryotic cells. Comparative sequence analysis shows that this essential gene has been highly conserved throughout eukaryotic evolution making it useful for phylogenetic analysis. Complete cDNA clones for the actin-encoding gene were isolated and characterized from Pneumocystis carinii purified from immunosuppressed rat lungs. The nucleotide sequence encodes a protein of 376 amino acids. The predicted actin protein of P. carinii shares a high degree of conservation to other known actins. Only one major actin gene was found in P. carinii. The P. carinii actin sequence was compared with 30 other actin sequences. Gene phylogenies constructed using both neighbor-joining and protein parsimony methods places the P. carinii actin sequence closest to the majority of the fungi. Since the phylogenetic relationship of P. carinii to fungi and protists has been questioned, these data on the actin gene phylogeny support the grouping of P. carinii with the fungi.

scholarly journals A Locus on Mouse Chromosome 2 Is Involved in Susceptibility to Congenital Hypothyroidism and Contains an Essential Gene Expressed in Thyroid

Endocrinology ◽  
2010 ◽  
Vol 151 (4) ◽  
pp. 1948-1958 ◽  
Author(s):  
Elena Amendola ◽  
Remo Sanges ◽  
Antonella Galvan ◽  
Nina Dathan ◽  
Giacomo Manenti ◽  
...  
Keyword(s):  
Congenital Hypothyroidism ◽  
Chromosomal Region ◽  
Essential Gene ◽  
Chromosome 2 ◽  
Nucleotide Polymorphisms ◽  
Single Nucleotide ◽  
Genes Encoding ◽  
Mouse Chromosome 2 ◽  
Essential Function

We report here the mapping of a chromosomal region responsible for strain-specific development of congenital hypothyroidism in mice heterozygous for null mutations in genes encoding Nkx2-1/Titf1 and Pax8. The two strains showing a differential predisposition to congenital hypothyroidism contain several single-nucleotide polymorphisms in this locus, one of which leads to a nonsynonymous amino acid change in a highly conserved region of Dnajc17, a member of the type III heat-shock protein-40 (Hsp40) family. We demonstrate that Dnajc17 is highly expressed in the thyroid bud and had an essential function in development, suggesting an important role of this protein in organogenesis and/or function of the thyroid gland.

scholarly journals The Nonessential H2A N-Terminal Tail Can Function as an Essential Charge Patch on the H2A.Z Variant N-Terminal Tail

2003 ◽  
Vol 23 (8) ◽  
pp. 2778-2789 ◽  
Author(s):  
Qinghu Ren ◽  
Martin A. Gorovsky
Keyword(s):  
Tetrahymena Thermophila ◽  
Gene Replacement ◽  
In Vitro Mutagenesis ◽  
Wild Type ◽  
Wild Type Protein ◽  
Lysine Residues ◽  
In The Wild ◽  
Essential Function

ABSTRACT Tetrahymena thermophila cells contain three forms of H2A: major H2A.1 and H2A.2, which make up ∼80% of total H2A, and a conserved variant, H2A.Z. We showed previously that acetylation of H2A.Z was essential (Q. Ren and M. A. Gorovsky, Mol. Cell 7:1329-1335, 2001). Here we used in vitro mutagenesis of lysine residues, coupled with gene replacement, to identify the sites of acetylation of the N-terminal tail of the major H2A and to analyze its function in vivo. Tetrahymena cells survived with all five acetylatable lysines replaced by arginines plus a mutation that abolished acetylation of the N-terminal serine normally found in the wild-type protein. Thus, neither posttranslational nor cotranslational acetylation of major H2A is essential. Surprisingly, the nonacetylatable N-terminal tail of the major H2A was able to replace the essential function of the acetylation of the H2A.Z N-terminal tail. Tail-swapping experiments between H2A.1 and H2A.Z revealed that the nonessential acetylation of the major H2A N-terminal tail can be made to function as an essential charge patch in place of the H2A.Z N-terminal tail and that while the pattern of acetylation of an H2A N-terminal tail is determined by the tail sequence, the effects of acetylation on viability are determined by properties of the H2A core and not those of the N-terminal tail itself.

scholarly journals The carB Gene Encoding the Large Subunit of Carbamoylphosphate Synthetase from Lactococcus lactis Is Transcribed Monocistronically

1998 ◽  
Vol 180 (17) ◽  
pp. 4380-4386 ◽  
Author(s):  
Jan Martinussen ◽  
Karin Hammer
Keyword(s):  
Lactococcus Lactis ◽  
Large Subunit ◽  
Arginine Deiminase ◽  
Gene Encoding ◽  
Pyrimidine Nucleotides ◽  
Reading Frame ◽  
Glutathione Peroxidases ◽  
Carbamoylphosphate Synthetase ◽  
Genes Encoding ◽  
High Degree

ABSTRACT The biosynthesis of carbamoylphosphate is catalyzed by the heterodimeric enzyme carbamoylphosphate synthetase. The genes encoding the two subunits of this enzyme in procaryotes are normally transcribed as an operon, but the gene encoding the large subunit (carB) in Lactococcus lactis is shown to be transcribed as an isolated unit. Carbamoylphosphate is a precursor in the biosynthesis of both pyrimidine nucleotides and arginine. By mutant analysis,L. lactis is shown to possess only onecarB gene; the same gene product is thus required for both biosynthetic pathways. Furthermore, arginine may satisfy the requirement for carbamoylphosphate in pyrimidine biosynthesis through degradation by means of the arginine deiminase pathway. The expression of the carB gene is subject to regulation at the level of transcription by pyrimidines, most probably by an attenuator mechanism. Upstream of the carB gene, an open reading frame showing a high degree of similarity to those of glutathione peroxidases from other organisms was identified.

scholarly journals Hidden resources in the Escherichia coli genome restore PLP synthesis and robust growth after deletion of the essential gene pdxB

2019 ◽  
Vol 116 (48) ◽  
pp. 24164-24173 ◽  
Author(s):  
Juhan Kim ◽  
Jake J. Flood ◽  
Michael R. Kristofich ◽  
Cyrus Gidfar ◽  
Andrew B. Morgenthaler ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Carbon Source ◽  
Sole Carbon Source ◽  
Essential Gene ◽  
Alternative Pathway ◽  
Gene Encoding ◽  
Genes Encoding ◽  
Phosphoglycerate Dehydrogenase ◽  
Functional Pathway

PdxB (erythronate 4-phosphate dehydrogenase) is expected to be required for synthesis of the essential cofactor pyridoxal 5′-phosphate (PLP) in Escherichia coli. Surprisingly, incubation of the ∆pdxB strain in medium containing glucose as a sole carbon source for 10 d resulted in visible turbidity, suggesting that PLP is being produced by some alternative pathway. Continued evolution of parallel lineages for 110 to 150 generations produced several strains that grow robustly in glucose. We identified a 4-step bypass pathway patched together from promiscuous enzymes that restores PLP synthesis in strain JK1. None of the mutations in JK1 occurs in a gene encoding an enzyme in the new pathway. Two mutations indirectly enhance the ability of SerA (3-phosphoglycerate dehydrogenase) to perform a new function in the bypass pathway. Another disrupts a gene encoding a PLP phosphatase, thus preserving PLP levels. These results demonstrate that a functional pathway can be patched together from promiscuous enzymes in the proteome, even without mutations in the genes encoding those enzymes.

scholarly journals RNA Polymerase II Localizes in Tetrahymena thermophila Meiotic Micronuclei When Micronuclear Transcription Associated with Genome Rearrangement Occurs

2004 ◽  
Vol 3 (5) ◽  
pp. 1233-1240 ◽  
Author(s):  
Kazufumi Mochizuki ◽  
Martin A. Gorovsky
Keyword(s):  
Life Cycle ◽  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Genome Rearrangement ◽  
Tetrahymena Thermophila ◽  
Germ Line ◽  
Essential Gene ◽  
Gene Encoding ◽  
Sexual Process ◽  
Rnap Ii

ABSTRACT The germ line micronucleus in Tetrahymena thermophila is transcriptionally silent in vegetatively growing cells. However, micronuclear transcription has been observed in the early (“crescent”) stages of the sexual process, conjugation. This transcription is proposed to play a central role in identifying sites for subsequent genome rearrangements that accompany development of the somatic macronucleus from the micronucleus. RPB3 (cnjC), a gene encoding a protein homologous to the third largest subunit of RNA polymerase II (RNAP II), was previously reported to be expressed specifically during conjugation, suggesting a role in micronucleus-specific transcription. Rpb3p localized in the micronucleus only during the meiotic prophase, when micronuclear transcription occurs, and its intranuclear distribution is strikingly similar to that for previously described sites of micronuclear RNA synthesis. By contrast, Rpc5p, the homologous subunit shared by RNAPs I and III, was not detectable in the micronucleus at any stage of the life cycle. However, Rpb3p is not specific to the transcribing micronucleus. Like Rpc5p, it also localizes to macronuclei in all stages of the life cycle. Rpb3p is encoded by a unique, essential gene in Tetrahymena. Thus, RNAP II is associated with both somatic transcription and crescent transcription and probably has an important role in genome rearrangement.

scholarly journals The essential function of Swc4p - a protein shared by two chromatin-modifying complexes of the yeast Saccharomyces cerevisiae - resides within its N-terminal part.

2008 ◽  
Vol 55 (3) ◽  
pp. 603-612 ◽  
Author(s):  
Arkadiusz Miciałkiewicz ◽  
Anna Chełstowska
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Essential Gene ◽  
Random Mutagenesis ◽  
Growth Conditions ◽  
Histone H2a ◽  
Terminal Part ◽  
Yeast Saccharomyces Cerevisiae ◽  
Chromatin Remodeling Complexes ◽  
Essential Function

The Swc4p protein, encoded by an essential gene, is shared by two chromatin-remodeling complexes in Saccharomyces cerevisiae cells: NuA4 (nucleosome acetyltransferase of H4) and SWR1. The SWR1 complex catalyzes ATP-dependent exchange of the nucleosomal histone H2A for H2AZ (Htz1p). The activity of NuA4 is responsible mainly for the acetylation of the H4 histone but also for the acetylation of H2A and H2AZ. In this work we investigated the role of the Swc4p protein. Using random mutagenesis we isolated a collection of swc4 mutants and showed that the essential function of Swc4p resides in its N-terminal part, within the first 269 amino acids of the 476-amino acid-long protein. We also demonstrated that Swc4p is able to accommodate numerous mutations without losing its functionality under standard growth conditions. However, when swc4 mutants were exposed to methyl methanesulfonate (MMS), hydroxyurea or benomyl, severe growth deficiencies appeared, pointing to an involvement of Swc4p in many chromatin-based processes. The mutants' phenotypes did not result from an impairment of histone acetylation, as in the mutant which bears the shortest isolated variant of truncated Swc4p, the level of overall H4 acetylation was unchanged.

scholarly journals Functional constraints on replacing an essential gene with its ancient and modern homologs

2016 ◽  
Author(s):  
Betül Kacar ◽  
Eva Garmendia ◽  
Nurcan Tunçbağ ◽  
Dan I. Andersson ◽  
Diarmaid Hughes
Keyword(s):  
Protein Function ◽  
Essential Gene ◽  
Elongation Factor ◽  
Network Connectivity ◽  
Protein Translation ◽  
Protein Activity ◽  
Gene Encoding ◽  
E Coli ◽  
Functional Conservation ◽  
Genes Encoding

AbstractThe complexity hypothesis posits that network connectivity and protein function are two important determinants of how a gene adapts to and functions in a foreign genome. Genes encoding proteins that carry out essential informational tasks in the cell, in particular where multiple interaction partners are involved, are less likely to be transferable to a foreign organism. Here we investigated the constraints on transfer of a gene encoding a highly conserved informational protein, translation elongation factor Tu (EF-Tu), by systematically replacing the endogenoustufAgene in theEscherichia coligenome with its extant and ancestral homologs. The extant homologs representedtufvariants from both near and distant homologous organisms. The ancestral homologs represented phylogenetically resurrectedtufsequences dating from 0.7 to 3.6 bya. Our results demonstrate that all of the foreigntufgenes are transferable to theE. coligenome, provided that an additional copy of the EF-Tu gene,tufB, remains present in theE. coligenome. However, when thetufBgene was removed, only the variants obtained from the γ-proteobacterial family (extant and ancestral), supported growth. This demonstrates the limited functional interchangability ofE. coli tufwith its homologs. Our data show a linear correlation between relative bacterial fitness and the evolutionary distance of the extanttufhomologs inserted into theE. coligenome. Our data and analysis also suggest that the functional conservation of protein activity, and its network interactivity, act to constrain the successful transfer of this essential gene into foreign bacteria.

scholarly journals Mutations that improve the efficiency of a weak-link enzyme are rare compared to adaptive mutations elsewhere in the genome

2019 ◽  
Author(s):  
Andrew B. Morgenthaler ◽  
Wallis R. Kinney ◽  
Christopher C. Ebmeier ◽  
Corinne M. Walsh ◽  
Daniel J. Snyder ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Weak Link ◽  
Experimental Studies ◽  
Gene Encoding ◽  
Evolutionary Trajectory ◽  
Adaptive Mutations ◽  
Parental Allele ◽  
Genes Encoding ◽  
Essential Function ◽  
Beneficial Allele

AbstractNew enzymes often evolve by amplification and divergence of genes encoding enzymes with a weak ability to provide a new function. Experimental studies to date have followed the evolutionary trajectory of an amplified gene, but have not addressed other mutations in the genome when fitness is limited by an evolving gene. We have adapted Escherichia coli in which an enzyme’s weak secondary activity has been recruited to serve an essential function. While the gene encoding the “weak-link” enzyme amplified in all eight populations, mutations improving the new activity occurred in only one. This beneficial allele quickly swept the amplified array, displacing the parental allele. Most adaptive mutations, however, occurred elsewhere in the genome. We have identified the mechanisms by which three of the classes of mutations increase fitness. These mutations may be detrimental once a new enzyme has evolved, and require reversion or compensation, leading to permanent changes in the genome.

scholarly journals Transcript encoded on the opposite strand of the human steroid 21-hydroxylase/complement component C4 gene locus

1989 ◽  
Vol 86 (17) ◽  
pp. 6582-6586 ◽  
Author(s):  
Y Morel ◽  
J Bristow ◽  
S E Gitelman ◽  
W L Miller
Keyword(s):  
Genetic Disorders ◽  
Gene Encoding ◽  
Gene Deletions ◽  
Adrenal Steroid ◽  
Genes Encoding ◽  
Opposite Strand ◽  
Human Chromosome 6 ◽  
Essential Function ◽  
Fourth Component ◽  
Considerable Detail

The gene encoding human adrenal steroid 21-hydroxylase (P450c21) and its highly similar pseudogene are duplicated in tandem with the two genes encoding the fourth component of human serum hemolytic complement (C4). This 60-kilobase gene complex, which lies within the major histocompatibility complex on the short arm of human chromosome 6, has been studied in considerable detail because genetic disorders in steroid 21-hydroxylation and in C4 are common. We have cloned a cDNA encoded by a previously unidentified gene in this region. This gene lies on the strand of DNA opposite from the strand containing the P450c21 and C4 genes, and it overlaps the last exon of P450c21. The newly identified gene encodes mRNAs of 3.5 and 1.8 kilobases that are expressed in the adrenal and in a Leydig cell tumor but are not expressed in nonsteroidogenic tissues. The sequence of the longest cDNA (2.7 kilobases) shows no similarity to known sequences available in two computerized data bases. The 5' end of this sequence is characterized by three repeats, each encoding about 100 amino acids flanked by potential sites for proteolytic cleavage. Although numerous studies have shown that gene deletions causing congenital adrenal hyperplasia occur in this region, none of these gene deletions extends into this newly identified gene, suggesting that it encodes an essential function.

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