The S7 ribosomal protein gene is truncated and overlaps a cytochrome c biogenesis gene in pea mitochondria

The pea mitochondrial genome contains a truncated rps7 gene lacking ca. 40 codons at its 5 terminus. This single-copy sequence is immediately downstream of and slightly overlapping an actively transcribed and edited reading frame of 744 bp (designated ccb248) homologous to the bacterial helC gene which encodes a subunit of the ABC-type heme transporter involved in cytochrome c biogenesis. This region of mitochondrial DNA appears recombinogenic, and the carboxy-termini of helC-type proteins are predicted to vary in sequence and length among plants. Sequences corresponding to the 5 coding region of rps7 were not detected elsewhere in the pea mitochondrial genome using wheat rps7 probes, and only a very short internal rps7 segment was observed in soybean mitochondrial DNA. The presence of rps7-homologous sequences in the nuclear genomes of pea and soybean is consistent with the recent transfer of a functional mitochondrial rps7 gene to the nucleus in certain plant lineages.     

Conserved sequence blocks 5′ to start codons of plant mitochondrial genes

Three sequence blocks of 10–12 bp are conserved in sequence and order 5 to putative start codons of several higher-plant mitochondrial genes. At least 25 examples were found, primarily associated with coxII, atp6, and orf25, in monocotyledons and dicotyledons. The proximal block can be 9 bp from start codons, and the three blocks generally occur within 100 bp 5 of start codons. In three examples 5 termini of the blocks represent recombination breakpoints, resulting in conservation of the blocks in resultant configurations. The two proximal blocks can form a secondary structure motif. The occurrence of the blocks near start codons, and conserved sequence and order, is consistent with a possible role in translation initiation or regulation.     

The mitochondrial coxII intron has been lost in two different lineages of dicots and altered in others

The central part of the mitochondrial coxII gene was amplified from 38 different dicots and two monocots using polymerase chain reaction. In 30 of the 40 plants studied, the amplified coxll gene-fragment contains an intron, ranging from 930 bp in Capsicum (pepper) in Solanaceae to 1,635 bp in Ampelamus albidans (climbing milkweed) in Asclepiadaceae. The composition of this intron varies as revealed by Southern hybridizations using oligonucleotide probes specific to the coxII intron-regions in maize, wheat, and rice. In the Apocynaceae, Calharanthus roseus (Madagascar periwinkle) and Vinca minor (common periwinkle) lack the coxII intron, while other members of the same family (various Mandevilla species, Nerium oleander and Apocynum cannabinum) and members of the closely related Asclepiadaceae (Asclepias incarnata, Ampelamus albidans and Asclepias tuberosa) retain the intron. Analysis of these data suggest a selective loss of the coxII intron from a plant, ancestral to both Catharanthus and Vinca, after the divergence of the Asclepiadaceae and Apocynaceae. The remaining eight plants from the Brassicaceae, Cucurbitaceae, Fabaceae, and Onagraceae lacking the intron fall into a single group or clade using the phylogenetic tree proposed by Chase et al. (Annals of the Missouri Botanical Garden: 80: 528–580, 1993) based on sequence of the chloroplast rbcL gene.     

Characterization of the S7 ribosomal protein gene in wheat mitochondria

Summary By screening a wheat mitoplast cDNA bank, we have identified an open reading frame of 444 by that has a derived amino acid sequence homologous to bacterial-type S7 ribosomal proteins. This gene, designated rps7, is located upstream of one of two 26S rRNA gene copies in the wheat mitochondrial genome and is expressed as an abundant mRNA of approximately 0.7 kb. Its 5 terminus maps to the end of an 80 by element that is closely related to sequences preceding the wheat coxII, orf25 and atp6 genes. Southern hybridization analysis indicates that rps7-homologous sequences are present in the mitochondria of rice and pea, but not soybean.     

DNA sequence and secondary structures of the large subunit rRNA coding regions and its two class I introns of mitochondrial DNA fromPodospora anserina

Summary DNA sequence analysis has shown that the gene coding for the mitochondrial (mt) large subunit ribosomal RNA (rRNA) fromPodospora anserina is interrupted by two class I introns. The coding region for the large subunit rRNA itself is 3715 bp and the two introns are 1544 (r1) and 2404 (r2) bp in length. Secondary structure models for the large subunit rRNA were constructed and compared with the equivalent structure fromEscherichia coli 23S rRNA. The two structures were remarkably similar despite an 800-base difference in length. The additional bases in theP. anserina rRNA appear to be mostly in unstructured regions in the 3 part of the RNA. Secondary structure models for the two introns show striking similarities with each other as well as with the intron models from the equivalent introns inSaccharomyces cerevisiae, Neurospora crassa, andAspergillus nidulans. The long open reading frames in each intron are different from each other, however, and the nucleotide sequence similarity diverges as it proceeds away from the core structure. Each intron is located within regions of the large subunit rRNA gene that are highly conserved in both sequence and structure. Computer analysis showed that the open reading frame for intron r1 contained a common maturase-like polypeptide. The open reading frames of intron r2 apeared to be chimeric, displaying high sequence similarity with the open reading frames in the r1 and ATPase 6 introns ofN. crassa.     

Multiple sites of crossing over within the Eb recombinational hotspot in the mouse

The Eb gene of the mouse major histocompatibility complex (MHC) contains a well-documented hotspot of recombination. Twelve cases of intra-Eb recombination derived from the b, d, k and s alleles of the Eb gene were sequenced to more precisely position the sites of meiotic recombination. This analysis was based on positioning recombination breakpoints between nucleotide polymorphisms found in the sequences of parental haplotypes. All twelve cases of recombination mapped within the second intron of the Eb gene. Six of these recombinants, involving the k and s haplotypes, mapped to two adjoining DNA segments of 394 and 955 base pairs (bp) in the 3 half of the intron. In an additional two cases derived by crossing over between the d and s alleles, breakpoints were positioned to adjoining segments of 28 and 433 bp, also in the 3 half of the intron. Finally, four b versus k recombinants were mapped to non-contiguous segments of DNA covering 2.9 kb and 1005 bp of the intron. An analysis of the map positions of crossover breakpoints defined in this study suggests that the second intron of the Eb gene contains a recombinational hotspot of approximately 800–1000 bp which contains at least two closely linked recombinationally active sites or segments. Further examination of the sequence data also suggests that the postulated location for the recombinational hotspot corresponds almost precisely to an 812 bp sequence that shows nucleotide sequence similarity to the MT family of middle repetitive DNA.     

Chloroplast DNA sequences integrated into an intron of a tomato nuclear gene

Summary DNA sequences capable of hybridizing with chloroplast DNA have previously been reported to exist in the nuclear genome of higher plants. Here we show that the third intron of the cultivated tomato (Lycopersicon esculentum) nuclear gene Cab-7, which resides on chromosome 10 and which we recently cloned and sequenced, contains two DNA fragments derived from the coding region of the chloroplast gene psbG. The first fragment, 133 bp long, is located at a site 63 bp from the 3 end of the 833 bp intron. The exact sequence of the 11 nucleotides at the 3 end of the inserting chloroplast sequence is also found at the 5 border of the insertion. A small (107 bp) chloroplast DNA fragment is inserted near the middle of the intron, again with the 3 end of the inserting element (6 bp) duplicated at the 5 border of the insertion. The second insert is a subfragment of the first insert, and is most likely directly derived from it. The psbG insertion sequence was found to be present in the Cab-7 gene of all tomato species examined but not in species from related genera (e.g. Solanum, Petunia, Nicotiana), suggesting that the original transposition event (chloroplast to nucleus) occurred relatively recently-since the divergence of the genus Lycopersicon from other genera in the family Solanaceae, but before radiation of species in that genus.     

Cloning of the fibroin gene from the oak silkworm, Antheraea yamamai and its complete sequence

The nucleotide sequences containing an entire genomic region and 5 upstream region of Antheraea yamamai fibroin gene have been determined. The gene consists of an initial exon encoding 14 amino acids, an intron (150 bp), and a long second exon coding for 2641 amino acids. The fibroin coding sequence shows a specialized organization with a highly repetitive region flanked by non repetitive 5 and 3 ends. Northern blot analyses confirmed that fibroin gene is actively expressed in the posterior silk gland of the final instar larvae of Antheraea yamamai.     

A pistil-specific thaumatin/PR5-like protein gene of Japanese pear (Pyrus serotina): sequence and promoter activity of the 5′ region in transgenic tobacco

The genomic clone encoding the pistil-specific thaumatin/PR5-like protein (PsTL1) was isolated from Japanese pear (Pyrus serotina). Sequence analysis showed that the genomic clone contained the 5-flanking sequence of 2.4 kb, the 3-flanking sequence of 648 bp and the coding region interrupted by a intron of 351 bp. A sequence motif conserved in some pistil self-incompatibility gene promoters of solanaceous and brassicaceous species was located in the 5-flanking region of the PsTL1 gene. The 2.4 kb 5-flanking region was fused to the GUS coding sequence and transferred to tobacco. Transgenic tobacco showed GUS activity in pistil and, at low level, in anther, but not in other floral organs and leaf. Histochemical analysis localized GUS activity to stigma, transmitting tissue, anther and pollen of transgenic tobacco.     

Expression of<Emphasis Type="Italic"> ENOD40</Emphasis> during tomato plant development

In legumes, ENOD40 expression is increased upon interaction of plants with rhizobia. Little is known of the expression pattern of ENOD40 during other stages of the plant life cycle. Studies of ENOD40 expression in non-legume development may give an indication of the function of the gene. To investigate the ENOD40 expression pattern during plant development, a fusion between the -glucuronidase (GUS) reporter gene and 150 bp of the 5 untranslated region plus 3,000 bp of 5 untranscribed tomato ENOD40 sequence was constructed and introduced into Lycopersicon esculentum Miller. Based on the observed GUS expression patterns in transgenic tomato we speculate that ENOD40 in tomato has a role in counteracting ethylene-provoked responses.Abbreviations GUS -glucuronidase - FISH fluorescence in situ hybridisation - RACE rapid amplification of cDNA ends - RFLC restriction fragment length polymorphism     

The plastid aldolase gene fromChlamydomonas reinhardtii: Intron/exon organization, evolution, and promoter structure

Genomic clones encoding the plastidic fructose- 1,6-bisphosphate aldolase ofChlamydomonas reinhardtii were isolated and sequenced. The gene contains three introns which are located within the coding sequence for the mature protein. No introns are located within or near the sequence encoding the transit-peptide, in contrast to the genes for plastidic aldolases of higher plants. Neither the number nor the positions of the three introns of theC. reinhardtii aldolase gene are conserved in the plastidic or cytosolic aldolase genes of higher plants and animals. The 5 border sequences of introns in the aldolase gene ofC. reinhardtii exhibit the conserved plant consensus sequence. The 3 acceptor splice sites for introns 1 and 3 show much less similarity to the eukaryotic consensus sequences than do those of intron 2. The plastidic aldolase gene has two tandemly repeated CAAT box motifs in the promoter region. Genomic Southern blots indicate that the gene is encoded by a single locus in theC. reinhardtii genome.     

The 3′ ends of two genes in the Balbiani ring c locus ofChironomus thummi

Summary The 3-end sequences of two nonallelic genes derived from the Balbiani ring c (BRc) locus ofChironomus thummi are described. Only one of the genes appears to be transcribed abundantly in normal late larval salivary glands. The two sequences are highly similar, even in the 3 untranslated regions, but sharply diverge beyond the polyadenylation site. Together with evidence from the 3 ends of BR1 and BR2 genes ofC. pallidivittatus andC. tentans, independently characterized by others, this result suggests the existence of a sequence-homogenization mechanism that operates across the 3 ends of all BR genes characterized to date. The 3-terminal coding region of each BRc gene is divided into two portions by a short intron. The upstream portion is homologous to and continuous with the tandem repeats that make up the internal core of each BR gene; however, that portion is variant in sequence relative to the core, and apparently is not subject to the homogenization process that operates on the core repeats. The portion downstream of the intron encodes a unique, 111-residue polypeptide highly different from the rest of the BRc product. The evolution of the various segments of the BRc genes is discussed.     

A Comparison of Intraspecific Patterns of DNA Sequence Variation in Mitochondrial DNA, α-Enolase,and MHC Class II B Loci in Auklets (Charadriiformes: Alcidae)

Patterns of DNA sequence variation can be used to learn about mechanisms of organismal evolution, but only if mechanisms of sequence evolution are well understood. Although theories of molecular evolution are well developed, few empirical studies have addressed patterns and mechanisms of sequence evolution in nuclear genes within species. In the present study, we compared DNA sequences among three loci with different evolutionary constraints to determine the influences of effective population size, balancing selection, and linkage on intraspecific patterns of sequence variation. Specifically, we assessed the degree and nature of polymorphism in a 307-base pair (bp) fragment of the mitochondrial cytochrome b gene, intron VIII of the gene for -enolase (a presumably neutral nuclear gene), and an ~600-bp fragment of an MHC class II B gene, including 155 bp of the hypervariable peptide binding region (a nuclear locus thought to be under balancing selection) for least and crested auklets (Aethia pusilla and A. cristatella; Charadriiformes: Alcidae). Transspecies polymorphism was found in both -enolase and the MHC but not cytochrome b and, given estimates of effective population size, probably represents retained ancestral variation. Biases in nucleotide composition suggested that mutational bias, tRNA availability, and the secondary structure of mRNA and/or DNA may influence base usage. Several lines of evidence indicated that balancing selection may be acting on the MHC II B exon 2. However, no evidence of balancing selection was observed in the intron and exon sequences immediately downstream of MHC II B exon 2. Current address (Hollie E. Walsh): Department of Zoology, University of Washington, Box 351800, Seattle, WA 98195-1800, USA     

Sequence and evolutionary history of the length polymorphism in intron 1 of the human red photopigment gene

The length of intron 1 of the red pigment gene is domorphic among Africans but not among Caucasians or Asians. This dimorphism was found to result from the presnce or basence of a block of 1,284 bp comprised of threeAlu elements and 328 bp of intervening unique-sequence DNA. This additional sequence in the long intron 1 of the red pigment gene was shown to have inserted into a fourth and olderAlu element present in the short form of intron 1. Furthermore, the size and sequence of the short intron 1 of the red pigment gene is equivalent to that of the adjacent green pigment gene. The block containing the threeAlu elements was not found in intron 1 of the red or green pigment genes of Old World monkeys and orangutans but was present in intron 1 ofboth the green and red pigment genes of gorillas and chimpanzees. The nucleotide sequence of this block in Old World primates and the estimated ages of the three elements suggest that their insertion occurred sequentially in the Old World monkey lineage prior to duplication of the ancestral X-chromosome-linked pigment gene. After gene duplication, deletion of the entire block containing the threeAlu elements from one of the genes created the short intron variant. Unequal recombination between the adjacent and highly homologous red and green pigment genes may have resulted in the formation of the short intron variant of the other gene.     

Molecular Basis of the Size Polymorphism of the First Intron of the<Emphasis Type="Italic">Adh-1</Emphasis> Gene of the Mediterranean Fruit Fly, <Emphasis Type="Italic">Ceratitis capitata</Emphasis>

The first intron of the gene encoding one of the alcohol dehydrogenase isoenzymes (ADH-1) in Ceratitis capitata is highly polymorphic in size. Five size variants of this intron were isolated from different strains and populations and characterized. Restriction map and sequence analysis showed that the intron size polymorphism is due to the presence or absence of (a) a copy of a defective mariner-like element, postdoc; (b) an 550-bp 3 indel which exhibits no similarity to any known sequence; and (c) a central duplication of 704 bp consisting of part of the 3 end of the postdoc element, the region between postdoc and the 3 indel, and the first 20 bp of the 3 indel. The homologous Adh-1 intron was amplified from the congeneric species, Ceratitis rosa, in order to obtain an outgroup for comparative and phylogenetic analyses. The C. rosa introns were polymorphic in size, ranging from about 1100 to 2000 bp, the major difference between them being the presence or absence of a mariner-like element Crmar2, unrelated to the postdoc element. Phylogenetic analysis suggests that the shorter intron variants in C. capitata may represent the ancestral form of the intron, the longest variants apparently being the most recent.     

Partial conservation of the 5′ ndhE-psaC-ndhD 3′ gene arrangement of chloroplasts in the cyanobacterium Synechocystis sp. PCC 6803: implications for NDH-D function in cyanobacteria and chloroplasts

The psaC gene, which encodes the 8.9 kDa iron-sulfur containing subunit of Photosystem I, has been sequenced from Synechocystis sp. PCC 6803 and shows greater similarity to reported plant sequences than other cyanobacterial psaC sequences. The deduced amino acid sequence of the protein encoded by the Synechocystis psaC gene is identical to the tobacco PSA-C sequence. In plants psaC is located in the small single-copy region of the chloroplast genome between two genes (designated ndhE and ndhD) with similarity to genes encoding subunits of the mitochondrial NADH Dehydrogenase Complex I. The 5 ndhE-psaC-ndhD3 gene arrangement of higher plants is only partially conserved in Synechocystis. An open reading frame (ORF) upstream of the Synechocystis psaC gene has 85% identity to the tobacco ndhE gene. Downstream of psaC there is a 273 bp ORF with 48% identity to the 5 portion of the tobacco ndhD gene (1527 bp). psaC, ndhE and the region of similarity to ndhD are present in a single copy in the Synechocystis genome. Part of the wheat ndhD gene was sequenced and used as a probe for the presence of the 3 portion of the ndhD gene. The wheat ndhD probe did not hybridize to Synechocystis or Anabaena sp. PCC 7120 genomic DNA, but did hybridize to Oenothera chloroplast DNA. These results indicate the complete ndhD gene is absent in two cyanobacteria, and raises the question of what role, if any, the ndhD gene product plays in the facultative heterotroph Synechocystis sp. PCC 6803.     

Tissue-specific and ABA-regulated Maize GIN gene expression in transgenic tobacco

Summary To study the regulatory functions of the ON promoter region, a ppG1b1GUS construct, consisting of 1402 bp 5 flanking sequence ofGlbl, 1919 by GUS coding sequence, and 283 by 3 NOS terminator, was cloned into a binary vector and introduced into tobacco plants byAgrobacterium-mediated transformation. Histochemical GUS assays of T_o tobacco mature seeds indicate that theGlbl promoter drives GUS expression in ABA treated seeds. Further GUS assays of the T, seeds at different developmental stages revealed that without ABA treatment, theGibl promoter drives GUS expression in immature seeds. The results from both T_o and T₁ tobacco plants indicated thatGlbl-driven GUS expression in tobacco is embryo specific.