Sequence and comparative analysis of the rabbit alpha-like globin gene cluster reveals a rapid mode of evolution in a G + C-rich region of mammalian genomes

A sequence of 10,621 base-pairs from the alpha-like globin gene cluster of rabbit has been determined. It includes the sequence of gene zeta 1 (a pseudogene for the rabbit embryonic zeta-globin), the functional rabbit alpha-globin gene, and the theta 1 pseudogene, along with the sequences of eight C repeats (short interspersed repeats in rabbit) and a J sequence implicated in recombination. The region is quite G + C-rich (62%) and contains two CpG islands. As expected for a very G + C-rich region, it has an abundance of open reading frames, but few of the long open reading frames are associated with the coding regions of genes. Alignments between the sequences of the rabbit and human alpha-like globin gene clusters reveal matches primarily in the immediate vicinity of genes and CpG islands, while the intergenic regions of these gene clusters have many fewer matches than are seen between the beta-like globin gene clusters of these two species. Furthermore, the non-coding sequences in this portion of the rabbit alpha-like globin gene cluster are shorter than in human, indicating a strong tendency either for sequence contraction in the rabbit gene cluster or for expansion in the human gene cluster. Thus, the intergenic regions of the alpha-like globin gene clusters have evolved in a relatively fast mode since the mammalian radiation, but not exclusively by nucleotide substitution. Despite this rapid mode of evolution, some strong matches are found 5' to the start sites of the human and rabbit alpha genes, perhaps indicating conservation of a regulatory element. The rabbit J sequence is over 1000 base-pairs long; it contains a C repeat at its 5' end and an internal region of homology to the 3'-untranslated region of the alpha-globin gene. Part of the rabbit J sequence matches with sequences within the X homology block in human. Both of these regions have been implicated as hot-spots for recombination, hence the matching sequences are good candidates for such a function. All the interspersed repeats within both gene clusters are retroposon SINEs that appear to have inserted independently in the rabbit and human lineages.     

Staphylococcus aureus has clustered tRNA genes.

The polymerase chain reaction (PCR) was used to detect large tRNA gene clusters in Bacillus subtilis, Bacillus badius, Bacillus megaterium, Lactobacillus brevis, Lactobacillus casei, and Staphylococcus aureus. The primers were based on conserved sequences of known gram-positive bacterial tRNA(Arg) and tRNA(Phe) genes. This PCR procedure detected an unusually large tRNA gene cluster in S. aureus. PCR-generated probes were used to identify a 4.5-kb EcoRI fragment that contained 27 tRNA genes immediately 3' to an rRNA operon. Some of these 27 tRNA genes are very similar, but only 1 is exactly repeated in the cluster. The 5' end of this cluster has a gene order similar to that found in the 9- and 21-tRNA gene clusters of B. subtilis. The 3' end of this S. aureus cluster exhibits more similarity to the 16-tRNA gene cluster of B. subtilis. The 24th, 25th, and 26th tRNA genes of this S. aureus tRNA gene cluster code for three similar, unusual Gly-tRNAs that may be used in the synthesis of the peptidoglycan in the cell wall but not in protein synthesis. Southern analysis of restriction digests of S. aureus DNA indicate that there are five to six rRNA operons in this bacterium's genome and that most or all may have large tRNA gene clusters at the 3' end.     

Comparative sequence analysis of the sorghum Rph region and the maize Rp1 resistance gene complex

A 268-kb chromosomal segment containing sorghum (Sorghum bicolor) genes that are orthologous to the maize (Zea mays) Rp1 disease resistance (R) gene complex was sequenced. A region of approximately 27 kb in sorghum was found to contain five Rp1 homologs, but most have structures indicating that they are not functional. In contrast, maize inbred B73 has 15 Rp1 homologs in two nearby clusters of 250 and 300 kb. As at maize Rp1, the cluster of R gene homologs is interrupted by the presence of several genes that appear to have no resistance role, but these genes were different from the ones found within the maize Rp1 complex. More than 200 kb of DNA downstream from the sorghum Rp1-orthologous R gene cluster was sequenced and found to contain many duplicated and/or truncated genes. None of the duplications currently exist as simple tandem events, suggesting that numerous rearrangements were required to generate the current genomic structure. Four truncated genes were observed, including one gene that appears to have both 5' and 3' deletions. The maize Rp1 region is also unusually enriched in truncated genes. Hence, the orthologous maize and sorghum regions share numerous structural features, but all involve events that occurred independently in each species. The data suggest that complex R gene clusters are unusually prone to frequent internal and adjacent chromosomal rearrangements of several types.     

Hox-3.6: isolation and characterization of a new murine homeobox gene located in the 5' region of the Hox-3 cluster.

Most members of the murine Hox gene system can be grouped into two subclasses based on their structural similarity to either one of the Drosophila homeotic genes Antennapedia (Antp) or Abdominal B (AbdB). All the AbdB-like genes reported thus far are located in the 5' region of their respective cluster. We describe here the isolation, structural characterization and spatio-temporal expression pattern of a new AbdB-like homeobox gene designated Hox-3.6 that is located in the 5' region of the Hox-3 cluster. Hox-3.6 has an extreme posterior expression domain in embryos of 12.5 days of gestation, a feature that has thus far only been observed for the 5' most genes of the Hox-4 cluster. Like the other members of the AbdB subfamily, Hox-3.6 exhibits spatially restricted expression in the hindlimb bud, but the expression domain is antero-proximal in contrast to the postero-distal domain reported for its cognate gene Hox-4.5. Structural analysis of the 5' region revealed the presence of a 35 bp sequence which shares homology and relative 5' position with an upstream sequence present in its two nearest downstream neighbors, Hox-3.2 and -3.1.     

Identification of the cellulose-binding domain of the cellulosome subunit S1 from Clostridium thermocellum YS

The 3' region of a gene designated cipB, which shows strong homology with cipA that encodes the cellulosome SL subunit of Clostridium thermocellum ATCC 27405, was isolated from a gene library of C. thermocellum strain YS. The truncated S1 protein encoded by the cipB derivative bound tightly to cellulose. The cellulose-binding domain in this polypeptide consisted of a C-terminal proximal 167 residue sequence which showed complete identity with residues 337-503 of mature SL from C. thermocellum strain ATCC 27405. The cellulose-binding domain interacted with both crystalline and amorphous cellulose, but not with xylan.     

Aflatoxin biosynthesis gene clusters and flanking regions

AIMS: To compare the biosynthetic gene cluster sequences of the main aflatoxin (AF)-producing Aspergillus species. METHODS AND RESULTS: Sequencing was on fosmid clones selected by homology to Aspergillus parasiticus sequence. Alignments revealed that gene order is conserved among AF gene clusters of Aspergillus nomius, A. parasiticus, two sclerotial morphotypes of Aspergillus flavus, and an unnamed Aspergillus sp. Phylogenetic relationships were established using the maximum likelihood method implemented in PAUP. Based on the Eurotiomycete/Sordariomycete divergence time, the A. flavus-type cluster has been maintained for at least 25 million years. Such conservation of the genes and gene order reflects strong selective constraints on rearrangement. Phylogenetic comparison of individual genes in the cluster indicated that ver-1, which has homology to a melanin biosynthesis gene, experienced selective forces distinct from the other pathway genes. Sequences upstream of the polyketide synthase-encoding gene vary among the species, but a four-gene sugar utilization cluster at the distal end is conserved, indicating a functional relationship between the two adjacent clusters. CONCLUSIONS: The high conservation of cluster components needed for AF production suggests there is an adaptive value for AFs in character-shaping niches important to those taxa. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first comparison of the complete nucleotide sequences of gene clusters harbouring the AF biosynthesis genes of the main AF-producing species. Such a comparison will aid in understanding how AF biosynthesis is regulated in experimental and natural environments.     

The nucleotide sequence of the large ribosomal RNA gene and the adjacent tRNA genes from rat mitochondria.

We have sequenced the Eco R(1) fragment D from rat mitochondrial DNA. It contains one third of the tRNA (Val) gene (the remaining part has been sequenced from the 3' end of the Eco R(1) fragment A) the complete gene for the large mt 16S rRNA, the tRNA (Leu) gene and the 5' end of an unidentified reading frame. The mt gene for the large rRNA from rat has been aligned with the homologous genes from mouse and human using graphic computer programs. Hypervariable regions at the center of the molecule and highly conserved regions toward the 3' end have been detected. The mt gene for tRNA Leu is of the conventional type and its primary structure is highly conserved among mammals. The mt gene for tRNA(Val) shows characteristics similar to those of other mt tRNA genes but the degree of homology is lower. Comparative studies confirm that AGA and AGG are read as stop codons in mammalian mitochondria.     

Characterization of the cellulolytic complex (cellulosome) of Clostridium acetobutylicum

A large cellulosomal gene cluster was identified in the recently sequenced genome of Clostridium acetobutylicum ATCC 824. Sequence analysis revealed that this cluster contains the genes for the scaffolding protein CipA, the processive endocellulase Cel48A, several endoglucanases of families 5 and 9, the mannanase Man5G, and a hydrophobic protein, OrfXp. Surprisingly, genetic organization of this large cluster is very similar to that of Clostridium cellulolyticum, the model of mesophilic clostridial cellulosomes. As C. acetobutylicum is unable to grow on cellulosic substrates, the existence of a cellulosomal gene cluster in the genome raises questions about its expression, function and evolution. Biochemical evidence for the expression of a cellulosomal protein complex was investigated. The results of sodium dodecyl sulfate-polyacrylamide gel electrophoresis, N-terminal sequencing and Western blotting with antibodies against specific components of the C. cellulolyticum cellulosome suggest that at least four major cellulosomal proteins are present. In addition, despite the fact that no cellulolytic activities were detected, we report here the evidence for the production of a high molecular mass cellulosomal complex in C. acetobutylicum.