The nucleotide sequence of the tobacco chloroplast gene for the large subunit of ribulose-l,5-bisphosphate carboxylase/oxygenase

The gene for the large subunit (LS) of ribulose-l,5-bisphosphate carboxylase/oxygenase (RuBPCase/ Oase) from tobacco has been cloned in pBR322 and sequenced. The coding region contains 1431 bp (477 codons). The deduced arnino acid sequence of tobacco LS protein shows 90% homology with those of maize and spinach LS. The positions in the gene corresponding to the 5' and the 3' ends of tobacco LS mRNA have been located on the DNA sequence by the S1 nuclease mapping procedure. The LS gene promoter sequence has homology with Escherichia coli promoter sequences; its terminator sequence is capable of forming a stem-and-loop structure. A sequence GGAGG, which is complementary to a sequence near the 3' end of tobacco chloroplast 16S rRNA and a putative ribosome binding site, occurs 6–10 bp upstream from the initiation codon.     

Nucleotide sequence of tobacco chloroplast gene for the alpha subunit of proton-translocating ATPase.

The tobacco chloroplast gene for the alpha subunit of proton-translocating ATPase has been cloned and sequenced. The coding region contains 1521 bp (507 codons). The nucleotide sequence and the deduced amino acid sequence show 55% and 54% homologies with those of the E. coli alpha subunit, respectively. The deduced amino acid composition is quite similar to that estimated for the spinach alpha subunit.     

The gene for the large subunit of ribulose-1,5-bisphosphate carboxylase in Euglena gracilis chloroplast DNA: location, polarity, cloning, and evidence for an intervening sequence

The gene for the large subunit (LS) of ribulose-1,5,-bisphosphate carboxylase of Euglena gracilis Z chloroplast DNA has been mapped by heterologous hybridization with DNA restriction fragments containing internal sequences from the Zea mays and Chlamydomonas reinhardii LS genes. The Euglena LS gene which has the same polarity as the Euglena rRNA genes has been located with respect to Pst I, Pvu I, and HindIII sites within the Eco RI fragment Eco A. The region of Euglena chloroplast DNA complementary to an 887 bp internal fragment from the Chlamydomonas chloroplast LS gene is interrupted by a 0.5-1.1 kbp non-complementary sequence. This is the first chloroplast protein gene located on the Euglena genome, and the first evidence for an intervening sequence within any chloroplast protein gene.     

Nucleotide sequence of a region of the mitochondrial genome of Aspergillus nidulans including the gene for ATPase subunit 6.

A 1500 bp fragment of the Aspergillus nidulans mitochondrial genome contains genes for arginine and asparagine tRNAs, an unassigned reading frame, and the structural gene for ATPase subunit 6. The tRNA genes possess 66% nucleotide homology and possibly originated by a relatively recent duplication event. The unassigned reading frame displays a low level of homology with the human URF A6L. The predicted amino acid sequence of the A-nidulans ATPase subunit 6 gene is 40% homologous to the yeast polypeptide and includes the short, highly conserved regions also present in the equivalent subunits from other mitochondrial systems and from Escherichia coli.     

Cloning, sequence, and expression of the Drosophila cAMP-dependent protein kinase catalytic subunit gene

Genomic DNA containing the protein coding region for Drosophila cAMP-dependent protein kinase catalytic subunit has been cloned and sequenced. The probe used to detect and isolate the gene fragment was constructed from two partially complementary synthetic oligonucleotides and contains 60 base pairs that encode (using Drosophila codon preferences) amino acids 195-214 of the beef heart catalytic subunit. In reduced stringency hybridization conditions, the probe recognizes two target sites in fly genomic DNA with 85% homology. One of these sites is in the cAMP-dependent protein kinase catalytic subunit gene, which was isolated as a 3959-base pair HindIII fragment. This fragment contains all of the protein coding portion, 900 base pairs upstream of the initiator ATG, and 2000 base pairs downstream of the termination codon (TAG). The coding portion of the gene contains no introns and yields a protein of 352 amino acids. There is a 2-amino acid insertion near the N terminus of the fly protein relative to the beef and mouse enzymes. Of the remaining 350 amino acids, 273 are invariant in the three species. A probe derived from the coding sequence of the HindIII clone hybridizes strongly to a 5100-base poly(A)+ RNA and weakly to 4100- and 3400-base poly(A)+ RNAs expressed in adult flies. A 2100-base pair EcoRI genomic fragment containing the second site recognized by the 60-base pair probe has also been cloned. DNA sequence analysis demonstrates that this fragment is part of the cGMP-dependent protein kinase gene or a close homolog. The catalytic subunit gene and the cGMP-dependent protein kinase gene have been located in regions 30C and 21D, respectively, of chromosome 2.     

Molecular cloning of the human thyrotropin-beta subunit gene

Genomic DNA fragments that carried a gene for human thyrotropin-beta (hTSH beta) subunit were isolated. Nucleotide sequence analysis of the gene showed that the hTSH beta subunit precursor consists of 138 amino acid residues. There is an N-terminal sequence of 20 amino acids as a signal peptide, followed by 112 amino acids, whose sequence is in agreement with that known for the secretory form of hTSH beta subunit. This is followed by an additional stretch of 6 hydrophobic amino acids, which may be eliminated post-translationally. The coding region is separated by an intron of about 460 bp. Genomic Southern blot hybridization analysis suggested that the hTSH beta gene is a unique single copy gene.     

Characterization and nucleotide sequence of the gene for the common alpha subunit of the bovine pituitary glycoprotein hormones.

The gene coding for the common alpha subunit of the bovine pituitary glycoprotein hormones was isolated from a bovine genomic library. The gene spans roughly 16.5 kbp, contains three intervening sequences, and codes for a message of approximately 730 nucleotides. The complete coding region of the gene was sequenced as well as 315 nucleotides of 5' flanking sequence and the entire intron C. Only a single base difference was found when the sequence of the gene was compared with that of the cDNA. Genomic blotting experiments suggest the presence of a single alpha subunit gene. Comparison of the bovine and human alpha subunit genes indicated that the high level of homology observed in the coding regions has been maintained throughout the 5' and 3' untranslated regions, and at least 90 nucleotides of the 5'flanking regions. Additionally, there is an 18 base pair sequence present in both the 5' flanking and 5' untranslated regions of the gene that is homologous to a region of the chick ovalbumin gene. This ovalbumin sequence has been suggested as a binding site for the progesterone receptor-complex.     

The mitochondrial genome of the fission yeast Schizosaccharomyces pombe: highly homologous introns are inserted at the same position of the otherwise less conserved cox1 genes in Schizosaccharomyces pombe and Aspergillus nidulans.

The DNA sequence of the second intron in the mitochondrial gene for subunit 1 of cytochrome oxidase (cox1), and the 3'' part of the structural gene have been determined in Schizosaccharomyces pombe. Comparing the presumptive amino acid sequence of the 3'' regions of the cox1 genes in fungi reveals similarly large evolutionary distances between Aspergillus nidulans, Saccharomyces cerevisiae and S. pombe. The comparison of exon sequences also reveals a stretch of only low homology and of general size variation among the fungal and mammalian genes, close to the 3'' ends of the cox1 genes. The second intron in the cox1 gene of S. pombe contains an open reading frame, which is contiguous with the upstream exon and displays all characteristics common to class I introns. Three findings suggest a recent horizontal gene transfer of this intron from an Aspergillus type fungus to S. pombe. (i) The intron is inserted at exactly the same position of the cox1 gene, where an intron is also found in A. nidulans. (ii) Both introns contain the highest amino acid homology between the intronic unassigned reading frames of all fungi identified so far (70% identity over a stretch of 253 amino acids). However, in the most homologous region, a GC-rich sequence is inserted in the A. nidulans intron, flanked by two direct repeats of 5 bp. The 37-bp insert plus 5 bp of direct repeat amounts to an extra 42 bp in the A. nidulans intron. (iii) TGA codons are the preferred tryptophan codons compared with TGG in all mitochondrial protein coding sequences of fungi and mammalia.(ABSTRACT TRUNCATED AT 250 WORDS)     

Protein phosphatase 2A: identification in Oryza sativa of the gene encoding the regulatory A subunit

A 2225 bp cDNA, designated RPA1, was isolated from an Oryza sativa cDNA library. Analysis revealed a 1761 bp coding sequence with 15 non-identical repeat units. The ORF encoded the A regulatory subunit of protein phosphatase 2A (PP2A-A) as ascertained by complementation of the yeast tpd3 mutant defective in this gene. The corresponding genomic DNA from a rice genome BAC library revealed that the gene contains eleven introns. The rice genome contains only a single copy of this gene as judged by Southern blot analysis. The PP2A protein is highly conserved in nature; the rice protein shows 88% amino acid identity with its counterparts in Arabidopsis or Nicotiana tabacum.     

Three variant introns of the same general class in the mitochondrial gene for cytochrome oxidase subunit 1 in Aspergillus nidulans

The oxiA gene of Aspergillus nidulans, coding for cytochrome oxidase subunit 1, is shown by DNA sequencing to contain three introns. An AUG start codon is not present at the beginning of the sequence, suggesting that either another codon, possibly the four base codon AUGA, is used for initiation or there is a further short intron between the true start codon and the beginning of the recognisable coding region. The second and third introns have long open reading frames, which could code for maturase proteins. The lack of conservation of amino acid sequence in the putative region of proteolytic cleavage for maturase formation suggests that the first conserved decapeptide may act as the recognition signal for protein processing. The third intron is remarkably (70%) homologous to the second intron of the cytochrome oxidase subunit 1 gene of Schizosaccharomyces pombe and both are located in exactly the same position. The third Aspergillus intron has an in-frame insertion of a 37-bp GC-rich DNA sequence which is now flanked by a 5-bp repeat, a well-known feature of transposable elements. All three introns in the oxiA gene have a 'core' RNA secondary structure found in a class of introns fitting the RNA splicing model of Davies et al. (1982). This core RNA structure may play a catalytic as well as a structural role in intron splicing. A sequence within the intron could act as a guide to align the splice sites of two of the introns in accordance with the model of Davies et al.     

Depression of the xylanase-encoding cgxA gene of Chaetomium gracile in Aspergillus nidulans

Regulation of the Chaetomium gracile xylanase A gene (cgxA) was investigated using Aspergillus nidulans as an intermediate host. Deletion of a 185 bp DNA fragment from its promoter region led to higher levels of the cgxA gene expression, indicating that the 185 bp DNA fragment contains an element involved in repression of the gene. A nuclear extract was assayed for proteins which bind to the 185 bp DNA fragment. A protein designated AnRP bound sequence specifically to the DNA fragment. The minimum sequence required for AnRP binding, 5'TTGACAAAT-3', was determined by means of gel mobility shift assays with various double-stranded oligonucleotides. Furthermore, this sequence repressed the expression of the cgxA gene when inserted at the 5' end of the cgxA gene on pXAH, which was deleted for the repressive element from the promoter region.     

RPC40, a unique gene for a subunit shared between yeast RNA polymerases A and C

Yeast RNA polymerases A and C share an approximately equal to 40 kd subunit. We have identified, sequenced, and mutagenized in vitro the AC40 subunit gene. The RPC40 gene is unique in the yeast genome and is required for cell viability. This gene contains an open reading frame encoding a 37.6 kd protein having no significant homology with bacterial RNA polymerase subunits. The promoter region contains a 19 bp sequence also present in the largest subunit of RNA polymerase C. It also contains a well-conserved RPG box, a sequence found in the promoter region of many genes encoding the translational apparatus. A novel, plasmid-shuffling method was developed to isolate a large number of RPC40 ts mutants. One of these, ts4, was shown to be defective in the synthesis of RNA polymerases A and C at the restrictive temperature. In contrast, RNA polymerase B was made normally.     

Isolation and nucleotide sequence of the ribosomal protein S16-encoding gene from Aspergillus nidulans.

A genomic clone has been isolated from Aspergillus nidulans which is homologous to the ribosomal (r) protein S16-encoding gene of Saccharomyces cerevisiae (S16A) and the r-protein S19-encoding gene of rat (S19). The amino acid (aa) sequences, deduced from nucleotide (nt) sequence analysis, show that in both cases more than 63% of the aa are conserved. The proposed A. nidulans r-protein S16 gene (rps16) differs from that of S. cerevisiae in that it occurs as a single copy in the haploid genome (rather than two copies as in yeast) and contains two putative introns (rather than one). The mRNA leader is long compared to many Aspergillus genes, commencing 293 nt upstream from the coding region, and contains an open reading frame of 13 codons.     

Molecular cloning and chromosomal mapping of the human gene for the testis-specific catalytic subunit of calmodulin-dependent protein phosphatase (calcineurin A).

A cDNA for an alternatively spliced variant of the testis-specific catalytic subunit of calmodulin dependent protein phosphatase (CaM-PrP) was cloned from a human testis library. The nucleotide sequence of 2134 base pairs (bp) encodes a protein of 502 amino acids (Mr approximately 57,132) and pI 7.0. The cDNA sequence differs from the murine form of this gene by a 30 bp deletion in the coding region, the position of which matches those in the two other genes for the catalytic subunit. These data indicate that this alternative splicing event arose prior to the divergence of the three genes. The deduced sequence of the human protein is only 88% identical to the homologous murine form, in striking contrast to the other two CaM-PrP catalytic subunits which are highly conserved between mouse and human (approximately 99%); this indicates a more rapid rate of evolution for the testis-specific gene. Analysis of Southern blots containing DNA from human-hamster somatic cell hybrids show that the gene is on human chromosome 8.     

Sequence of a genomic DNA clone for the small subunit of ribulose bis-phosphate carboxylase-oxygenase from tobacco.

We have cloned and sequenced a gene for the small subunit (SS) of ribulose bis-phosphate carboxylase-oxygenase from Nicotiana tabacum. The tobacco gene is most closely related to the SS genes from the dicots soybean and pea, and less so to the monocots wheat and Lemna; the deduced amino acid sequence of the mature protein is in all cases more closely conserved than is its chloroplast transit sequence. Unlike the genomic sequences of the two monocots, which have one intron, and the two other dicots, which have two introns, the tobacco gene has three introns. The third tobacco intron lies within a highly conserved region of the protein. Its position coincides with the boundary of a 12 amino acid insertion in the SS genes of higher plants, relative to those of blue green algae. The 5' flanking end of the gene carries 67 bp inverted repeats, which flank a series of eight direct repeats; the direct repeats themselves each carry inverted repeats. The 3' untranslated end of this gene differs by only 2 bp from that of an N. sylvestris SS gene.     

Cloning and DNA sequence of the 5'-exonuclease gene of bacteriophage T5

The nucleotide sequence of the BalI-PstI fragment of T5 DNA, 1347 bp in length, coding for 5'-exonuclease (D15 gene), has been determined. A coding region of the gene contains 873 bp and is preceded by a typical Shine-Dalgarno sequence. The D15 gene belongs to a cluster, consisting of at least 3 genes, in which a termination codon of a preceding gene overlaps an initiation codon of the following one. The sequence contains an open reading frame for 291 amino acid residues. The molecular mass of the 5'-exonuclease calculated from the predicted amino acid sequence is 33 400 Da.