Search PubMed⌕ Search

Biomedical subjects

M W Gray

Publications and source records attributed to M W Gray.

At least 127 records · Page 7Linked to original sources
PubMed1986

Structure and evolution of the small subunit ribosomal RNA gene of Crithidia fasciculata.

We present the cloning and sequence analysis of the nuclear-encoded Crithidia fasciculata small subunit (SSU) rRNA gene, the longest (2,206 bp) such gene yet characterized by direct sequence analysis. Much of the sequence can be folded to fit a phylogenetically conserved secondary structure model, with the additional length of this gene being accommodated within discrete variable domains that are present in eukaryotic SSU rRNAs. On the basis of sequence comparisons, we conclude that Crithidia contains the most highly diverged SSU rRNA described to date among the eukaryotes, and therefore represents one of the earliest branchings within the eukaryotic primary kingdom.

Animals↗
PubMed1985

The wheat mitochondrial gene for apocytochrome b: absence of a prokaryotic ribosome binding site.

The wheat mitochondrial gene for apocytochrome b (CYB) has been identified by its hybridization to a yeast CYB probe and its nucleotide sequence has been determined. The wheat CYB sequence predicts a cytochrome b apoprotein of 398 amino acids; it is almost identical to that of maize but has ten additional amino acids at the carboxy terminus. No introns are present in the wheat CYB gene, but an internal segment of the gene is repeated at another genomic location. Transcript analysis reveals a single wheat CYB mRNA of approximately 2.4 kb with a long untranslated leader. Sequences upstream of the CYB coding region are very similar in wheat and maize but the stretch proposed to be a ribosome binding site in maize is not conserved in wheat. The corresponding leader regions of the wheat mitochondrial mRNAs for cytochrome oxidase subunits I and II also lack complementarity to the 3'-end of the small subunit rRNA. We conclude that alternative signals are involved in the initiation of translation in plant mitochondria.

Amino Acid Sequence↗
PubMed1985

Phenylalanine and tyrosine transfer RNAs encoded by Tetrahymena pyriformis mitochondrial DNA: primary sequence, post-transcriptional modifications, and gene localization.

We have isolated Phe and Tyr tRNAs from Tetrahymena pyriformis mitochondria and have determined that these are "native" species, encoded by the mtDNA. A single gene for the tRNA(Phe) has been positioned 12-14 kbp from the left end of the linear Tetrahymena mtDNA, while duplicate tRNA(Tyr) genes have been localized within the inverted terminal repeats of this genome. Primary sequence analysis demonstrates that the tRNA(Tyr) has all of the characteristic primary and secondary structural features of a normal tRNA; however, the tRNA(Phe) displays several atypical features, including (i) replacement of the usual T psi sequence by UC, (ii) a U.U pair in the T psi C stem, and (iii) an extra 5'-nucleotide (U).

Animals↗
PubMed1985

Genes for respiratory chain proteins and ribosomal RNAs are present on a 16-kilobase-pair DNA species from Chlamydomonas reinhardtii mitochondria.

We have used heterologous hybridization and DNA sequence analysis to determine whether the 16-kilobase-pair (kbp) DNA from Chlamydomonas reinhardtii mitochondria is the functional equivalent of mtDNA in other eukaryotes. Restriction fragments corresponding to a continuous internal stretch spanning 75% of the 16-kbp DNA have been cloned and mapped, and regions hybridizing with probes specific for the cytochrome oxidase subunit I [CytOx I (acronym COI)] and apocytochrome b (Cyt b) genes of yeast and the mitochondrial 26S and 18S rRNA genes of wheat have been identified. Sequence analysis has verified the presence of CytOx I and the large and small subunit rRNA genes in the C. reinhardtii 16-kbp DNA. In the region of the 16-kbp DNA corresponding to exon 4 in the yeast CytOx I gene, the derived amino acid sequence is 61% and 63% identical with the CytOx I amino acid sequences of yeast and human mitochondria, respectively. Notably, tryptophan is specified by TGG rather than by TGA in this section of the C. reinhardtii CytOx I gene. A probe from the CytOx I region of the 16-kbp DNA hybridizes only with this 16-kbp DNA in Southern blots of total cellular DNA from C. reinhardtii but with a larger DNA species in the total cellular DNA of C. moewusii and C. eugametos--two species that lack a 16-kbp DNA. These observations provide evidence that C. reinhardtii 16-kbp DNA comprises at least part of the mitochondrial genome of this organism and that a homologous DNA exists in other species of Chlamydomonas.

Base Sequence↗
PubMed1984

On the evolutionary descent of organisms and organelles: a global phylogeny based on a highly conserved structural core in small subunit ribosomal RNA.

To probe the earliest evolutionary events attending the origin of the five known genome types (archaebacterial, eubacterial, nuclear, mitochondrial and plastid), we have analyzed sequences corresponding to a ubiquitous, highly conserved core of secondary structure in small subunit rRNA. Our results support (i) the existence of three primary lineages (archaebacterial, eubacterial, and nuclear), (ii) a specific eubacterial ancestry for plastids and mitochondria (plant, animal, fungal), and (iii) an endosymbiotic, evolutionary origin of the two types of organelle from within distinct groups of eubacteria (blue-green algae (cyanobacteria) in the case of plastids, nonphotosynthetic aerobic bacteria in the case of mitochondria). In addition, our analysis suggests (iv) a biphyletic origin of mitochondria, with animal and fungal mitochondria branching together but separately from plant mitochondria, and (v) a monophyletic origin of plastids. The method described here provides a powerful and generally applicable molecular taxonomic approach towards a global phylogeny encompassing all organisms and organelles.

Animals↗
PubMed1984

Evidence for homologous recombination between repeated sequences containing 18S and 5S ribosomal RNA genes in wheat mitochondrial DNA.

Closely linked genes for 18S and 5S rRNAs have been located on four different cloned SalI restriction fragments of wheat (Triticum aestivum L.) mitochondrial DNA. Restriction analysis has revealed that in each of the cloned fragments, the 18S and 5S rRNA genes are contained within the same basic structural unit, R, which is at least 4 kbp long. This unit is flanked by sequences designated u (0.8 kbp), v (13.7 kbp), w (0.7 kbp), and y (1.4 kbp), in the orientations v-R-w, v-R-y, u-R-w, and u-R-y in the four different SalI fragments. We conclude that 18S + 5S rRNA genes are located at several distinct sites in the wheat mitochondrial genome, and suggest that reciprocal intra- and/or intermolecular recombination between such repeated sequences could promote extensive genomic rearrangement and thus contribute to the physical heterogeneity that is a hallmark of most plant mitochondrial DNAs.

Journal Article↗
PubMed1984

The wheat cytochrome oxidase subunit II gene has an intron insert and three radical amino acid changes relative to maize.

We have determined the sequence of the wheat mitochondrial gene for cytochrome oxidase subunit II (COII) and find that its derived protein sequence differs from that of maize at only three amino acid positions. Unexpectedly, all three replacements are non-conservative ones. The wheat COII gene has a highly-conserved intron at the same position as in maize, but the wheat intron is 1.5 times longer because of an insert relative to its maize counterpart. Hybridization analysis of mitochondrial DNA from rye, pea, broad bean and cucumber indicates strong sequence conservation of COII coding sequences among all these higher plants. However, only rye and maize mitochondrial DNA show homology with wheat COII intron sequences and rye alone with intron-insert sequences. We find that a sequence identical to the region of the 5' exon corresponding to the transmembrane domain of the COII protein is present at a second genomic location in wheat mitochondria. These variations in COII gene structure and size, as well as the presence of repeated COII sequences, illustrate at the DNA sequence level, factors which contribute to higher plant mitochondrial DNA diversity and complexity.

Journal Article↗
PubMed1984

Pronounced structural similarities between the small subunit ribosomal RNA genes of wheat mitochondria and Escherichia coli.

We present here the nucleotide sequence of the small subunit (18S) rRNA gene from wheat mitochondria. Aside from five discrete variable domains, this gene and the analogous (16S) rRNA gene in Escherichia coli show essentially a one-to-one correspondence in their potential secondary structures, with regions accounting for 86% of the bacterial 16S rRNA having a strict secondary structure counterpart in the mitochondrial 18S rRNA. Primary sequence identity between the two rRNAs ranges from 73% to 85% (76% overall) within regions of conserved secondary structure. Within a smaller secondary structure core common to all small subunit rRNAs, the wheat mitochondrial sequence shares substantially more primary sequence identity with the E. coli (eubacterial) sequence (88%) than with the small subunit rRNA sequences of Halobacterium volcanii (an archaebacterium) (71%) or Xenopus laevis cytoplasm (61%). Moreover, the wheat mitochondrial sequence contains a very high proportion of certain lineage-specific residues that distinguish eubacterial/plastid 16S rRNAs from archaebacterial 16S and eukaryotic cytoplasmic 18S rRNAs. These data establish that the ancestry of the wheat mitochondrial 18S rRNA gene can be traced directly and specifically to the eubacterial primary kingdom, and the data provide compelling support for a relatively recent xenogenous (endosymbiotic) origin of plant mitochondria from eubacteria-like organisms.

Base Sequence↗
PubMed1983

Primary structures of four novel small ribosomal RNAs from Crithidia fasciculata.

We report here the complete primary structures of four novel small RNA species (designated e, f, g, and j) found in the large ribosomal subunit of Crithidia fasciculata, a trypanosomatid protozoan. These RNAs, which are distinct from Crithidia 5S (species h) and 5.8S (species i) rRNAs, do not have counterparts in the more conventional eukaryotic ribosomes characterized to date. The small RNAs are 212 (e), 183 (f), 135-136 (g), and 72-73 (j) nucleotides long, with g and j displaying 5'-terminal heterogeneity. All have unique sequences and all contain 5'-monophosphorylated and 3'-unphosphorylated termini. In their basic structural features, therefore, species e, f, g, and j are indistinguishable from other RNAs (including 5S and 5.8S) that are recognized components of eukaryotic ribosomes, although they are unrelated to 5S or 5.8S rRNA in sequence. Since previous work from this laboratory has ruled out the possibility that these small RNAs are generated by quantitative and highly specific (albeit artifactual) RNase cleavage of large rRNAs during isolation, we conclude that species e, f, g, and j are native components of the Crithidia ribosome. With the exception of e, which appears to contain a single pseudouridine residue, all of these novel RNA species are devoid of modified nucleosides. In connection with primary sequence analysis, we present a simple modification of the standard G-specific chemical sequencing reaction which in our hands yields reproducible and unambiguous results using commercially available dimethyl sulfate.

Animals↗
PubMed1982

3'-Terminal sequence of wheat mitochondrial 18S ribosomal RNA: further evidence of a eubacterial evolutionary origin.

We have determined the sequences of the 3'-terminal approximately 100 nucleotides of [5' -32P]pCp-labeled wheat mitochondrial, wheat cytosol, and E. coli small sub-unit rRNAs. Sequence comparison demonstrates that within this region, there is a substantially greater degree of homology between wheat mitochondrial 18S and E. coli 16S rRNAs than between either of these and wheat cytosol 18S rRNA. Moreover, at a position occupied by 3-methyluridine in E. coli 16S rRNA, the same (or a very similar) modified nucleoside is present in wheat mitochondrial 18S rRNA but not in wheat cytosol 18S rRNA. Further, E. coli 16S and 23S rRNAs hybridize extensively to wheat mitochondrial 18S and 26S rRNA genes, respectively, but wheat cytosol 18S and 26S rRNAs do not. No other mitochondrial system studies to date has provided comparable evidence that a mitochondrial rRNA is more closely related to its eubacterial homolog than is its counterpart in the cytoplasmic compartment of the same cell. The results reported here provide additional support for the view that plant mitochondria are of endosymbiotic, specifically eubacterial, origin.

Animals↗
PubMed1982

Nucleotide sequence of an exceptionally long 5.8S ribosomal RNA from Crithidia fasciculata.

In Crithidia fasciculata, a trypanosomatid protozoan, the large ribosomal subunit contains five small RNA species (e, f, g, i, j) in addition to 5S rRNA [Gray, M.W. (1981) Mol. Cell. Biol. 1, 347-357]. The complete primary sequence of species i is shown here to be pAACGUGUmCGCGAUGGAUGACUUGGCUUCCUAUCUCGUUGA ... AGAmACGCAGUAAAGUGCGAUAAGUGGUApsiCAAUUGmCAGAAUCAUUCAAUUACCGAAUCUUUGAACGAAACGG ... CGCAUGGGAGAAGCUCUUUUGAGUCAUCCCCGUGCAUGCCAUAUUCUCCAmGUGUCGAA(C)OH. This sequence establishes that species i is a 5.8S rRNA, despite its exceptional length (171-172 nucleotides). The extra nucleotides in C. fasciculata 5.8S rRNA are located in a region whose primary sequence and length are highly variable among 5.8S rRNAs, but which is capable of forming a stable hairpin loop structure (the "G+C-rich hairpin"). The sequence of C. fasciculata 5.8S rRNA is no more closely related to that of another protozoan, Acanthamoeba castellanii, than it is to representative 5.8S rRNA sequences from the other eukaryotic kingdoms, emphasizing the deep phylogenetic divisions that seem to exist within the Kingdom Protista.

Animals↗
PubMed1982

Comparative sequence analysis as an approach to evaluating structure, function, and evolution of 5S and 5.8S ribosomal RNAs.

Nucleotide sequences of nine eukaryotic and nine eubacterial 5S rRNAs have been selected for their diversity and subjected to analysis of primary and potential secondary structure. This analysis has allowed the quantitative confirmation of several previously made observations concerning 5S rRNA structure: (i) these two 5S rRNAs are derived from a common ancestor and probably perform essentially the same function in protein synthesis; (ii) one domain of 5S rRNA has undergone considerable divergence of structure (and presumably function) since the separation of the eukaryotic and eubacterial lineages; and (iii) single-stranded regions are more highly conserved than double-stranded regions. In addition, this analysis leads us to propose that (i) some of the highly conserved nucleotide residues in single-stranded regions interact in a specific manner with protein components of the translational apparatus, and (ii) repetitive folding and unfolding of helical regions occurs in two regions of eukaryotic 5S rRNA and one region of eubacterial 5S rRNA. In the context of these observations and propositions we also consider the potential secondary structure of plant mitochondrial 5S rRNA. Nucleotide sequences of 5.85 rRNAs have yielded less information about secondary structure and possible functional interactions. However, we have identified highly conserved and variable regions within this molecule and we show (in contrast to the situation with 5S rRNA) that these do not correlate well with proposed single-stranded and helical regions in a current model of 5.8S secondary structure.

Animals↗
PubMed1981

Is wheat mitochondrial 5S ribosomal RNA prokaryotic in nature?

Küntzel et al. (1981) (Nucleic Acids Res. 9, 1451-1461) recently concluded that the sequence of wheat mitochondrial 5S rRNA is significantly more related to prokaryotic than to eukaryotic 5S rRNA sequences, and displays an especially high affinity to that of the thermophilic Gram-negative bacterium, Thermus aquaticus. However, the sequence on which this conclusion was based, although attributed to us, differs in several places from the one determined by us. We show here that the correct sequence (Spencer, D.F., Bonen, L. and Gray, M.W. (1981) Biochemistry, in press) does not support the conclusions of Küntzel et al. about potential secondary structure in wheat mitochondrial 5S rRNA and its phylogenetic significance. We further show that when the wheat mitochondrial 5S rRNA sequence is matched against published alignments for E. coli, T. aquaticus, and wheat cytosol 5S rRNAs, the mitochondrial sequence shows no greater homology to the T. aquaticus sequence than to the E. coli sequence, and only slightly more homology to these two sequences than to wheat cytosol 5S rRNA. This analysis confirms our original view (Biochemistry, in press) that wheat mitochondrial 5S rRNA is neither obviously prokaryotic nor eukaryotic in nature, but shows characteristics of both classes of 5S rRNA, as well as some unique features.

Base Sequence↗
PubMed1981

Primary sequence of wheat mitochondrial 5S ribosomal ribonucleic acid: functional and evolutionary implications.

Using the procedures of Donis-Keller et al. [Donis-Keller, H., Maxam, A. M., & Gilbert, W. (1977) Nucleic Acids Res. 4, 2527--2538 (1977)] and Peattie [Peattie, D. A. (1979) Proc. Natl. Acad. Sci. U.S.A. 76, 1760--1764], we have determined the nucleotide sequence of wheat mitochondrial 5S ribosomal ribonucleic acid (rRNA). This sequence [Formula: see text] is the first to be reported for a plant mitochondrial RNA. A highly conserved region (underlined) readily identifies the molecule as a structural homologue of other 5S rRNAs, as do potential base-paired regions which are characteristic of all known (prokaryotic, chloroplast, eukaryotic cytosol) 5S rRNA sequences. However, when assessed in terms of those structural features which distinguish prokaryotic from eukaryotic 5S rRNAs, wheat mitochondrial 5S rRNA cannot be classified readily as one or the other but instead displays characteristics of both types. In addition, the mitochondrial 5S rRNA has several unusual features, including (i) a variable number (two to three) of A residues at both the 5' and 3' ends, (ii) a unique sequence (CGACC, italic) in place of the prokaryotic sequence (CGAAC) which has been postulated to interact with aminoacyl-tRNA during translation, and (iii) a novel sequence, AUAUAUAU, immediately following the highly conserved sequence. In terms of overall primary sequence, wheat mitochondrial and cytosol 5S rRNAs seem to be slightly more divergent from each other than either is from Escherichia coli 5S rRNA, with which they are about equally homologous. From these observations, we propose that wheat mitochondrial 5S rRNA represents a distinct class of 5S rRNA. Our observations raise a number of questions about the evolutionary origin and functional role(s) of plant mitochondrial 5S rRNA.

Base Composition↗
PubMed1981

Unusual pattern of ribonucleic acid components in the ribosome of Crithidia fasciculata, a trypanosomatid protozoan.

In a previous study from this laboratory, presumptive ribosomal ribonucleic acid (RNA) species were identified in the total cellular RNA directly extracted from intact cells of the trypanosomatid protozoan Crithidia fasciculata (M. W. Gray, Can. J. Biochem. 57:914-926, 1979). The results suggested that the C. fasciculata ribosome might be unusual in containing three novel, low-molecular-weight ribosomal RNA components, designated e, f, and g (apparent chain lengths 240, 195, and 135 nucleotides, respectively), in addition to analogs of eucaryotic 5S (species h) and 5.8S (species i) ribosomal RNAs. In the present study, all of the presumptive ribosomal RNAs were indeed found to be associated with purified C. fasciculata ribosomes, and their localization was investigated in subunits produced under different conditions of ribosome dissociation. When ribosomes were dissociated in a high-potassium (880 mM K+, 12.5 mM Mg2+) medium, species e to i were all found in the large ribosomal subunit, which also contained an additional, transfer RNA-sized component (species j). However, when subunits were prepared in a low-magnesium (60 mM K+, 0.1 mM Mg2+) medium, two of the novel species (e and g) did not remain with the large subunit, but were released, apparently as free RNAs. Control experiments have eliminated the possibility that the small RNAs are generated by quantitative and highly specific (albeit artifactual) ribonuclease cleavage of large ribosomal RNAs during isolation. In terms of RNA composition and dissociation properties, therefore, the ribosome of C. fasciculata is the most "atypical" eucaryotic ribosome yet described. These observations raise interesting questions about the function and evolutionary origin of C. fasciculata ribosomes and about the organization and expression of ribosomal RNA genes in this organism.

Animals↗