Deletion Mapping of the λ REX Gene

Deletion mapping has been used to order 12 λ rex^- mutants. Correlation of recombination data with physically-determined positions of deletion end-points (Szybalski 1971; Blattner et al. 1972) suggests that the left-most rex^- mutation, rex209, is located about 260-300 nucleotide pairs from the p_L mutation sex1 and about 475 nucleotide pairs from the left end-point of the region of nonhomology with λimm434.     

Investigation of Gene Insertion–Deletion Polymorphism of the Gene for Angiotensin-Converting Enzyme in Populations of the Volga–Ural Region

Insertion–deletion polymorphism at the angiotensin I-converting enzyme (ACE) gene in populations of the Volga–Ural region was examined by means of polymerase chain reaction. The populations studied belong to the Finno-Ugric (Komis, Maris, Mordovians, and Udmurts), Turkic (Chuvashes, Tatars, and Bashkirs), and Eastern-Slavic (Russians) ethnic groups. Distribution patterns of allele and genotype frequencies of this polymorphic system in the examined region were characterized. Comparison of the obtained results with the literature data on the ACE gene polymorphism in other Caucasoid and Mongoloid populations revealed some trends in the ACE genotype frequency dynamics depending on the ethnicity of the populations.     

Polymorphism and Linkage of the αa-Crystallin Gene in t-Haplotypes of the Mouse

Restriction fragment polymorphisms were used to order the alpha A-crystallin locus (Crya-1) relative to other genes in mouse t-chromatin and to investigate the relatedness of alpha-A-crystallin sequences among different t-haplotypes. Analysis of DNA from t-recombinant mice mapped Crya-1 to the K end of the H-2 complex and within the distal inverted region characteristic of t-haplotypes. Hybridization with Crya-1 cDNA revealed three distinct phenotypic groups among the 17 different t-haplotypes studied. A majority (9 of 17) of the t-haplotypes were classified into a novel group (Crya-1t) characterized by restriction fragments apparently unique to t-chromosomes and therefore thought to contain alpha A-crystallin sequences descended from the original t-chromosome. A second group of t-haplotypes had restriction fragment patterns indistinguishable from those observed among many common inbred strains of mice of the Crya-1a type, and a third restriction fragment pattern, observed only in the tw121 haplotype, was indistinguishable from the fragment pattern for C3H/DiSn (Crya-1b) and several other inbred strains of mice. Thus, with respect to sequences around the Crya-1 locus, different t-haplotypes show restriction fragment polymorphisms, some of which are comparable to those found in wild-type chromosomes and provide further evidence for genetic heterogeneity in DNA from the distal region of t-haplotypes.     

Cloning of the Xuhuai Goat PPARγ Gene and the Preparation of Transgenic Sheep

This study aimed to clone the peroxisome proliferator-activated receptor γ (PPARγ) gene of the Xuhuai goat and to make transgenic sheep using intratesticular injection, so as to improve the meat quality and flavor by increasing the intramuscular fat content. The coding sequence of the goat PPARγ gene was 1,428 bp, encoding 475 amino acids. Its similarity with other species was 81 (chicken), 89 (mouse), 92 (pig), 98 (cow), and 99% (sheep). The similarity of the corresponding amino acid sequences was 92.9, 97.3, 98.3, 99.6, and 99.8%, respectively. The signal peptide region of the PPARγ protein was not found in this study, demonstrating that the protein is not secreted. RT-PCR and western blot revealed that PPARγ was expressed in vitro, and the protein was localized in the cytoplasm. The PPARγ gene was expressed in F₁ transgenic sheep at both the mRNA and the protein levels; the positive ratio was 13.7%.     

Genetic Characterization of the Drosophila Birt-Hogg-Dubé Syndrome Gene

Folliculin (FLCN) is a conserved tumor suppressor gene whose loss is associated with the human Birt-Hogg-Dubé (BHD) syndrome. However, its molecular functions remain largely unknown. In this work, we generated a Drosophila BHD model through genomic deletion of the FLCN gene (DBHD⁻). The DBHD mutant larvae grew slowly and stopped development before pupation, displaying various characteristics of malnutrition. We found the growth delay was sensitive to the nutrient supplies. It became more severe upon restrictions of the dietary yeast; while high levels of yeast significantly restored the normal growth, but not viability. We further demonstrated that leucine was able to substitute for yeast to provide similar rescues. Moreover, the human FLCN could partially rescue the DBHD⁻ phenotypes, indicating the two genes are involved in certain common mechanisms. Our work provides a new animal model of the BHD syndrome and suggests that modulation of the local nutrient condition might be a potential treatment of the BHD lesions.     

Characterization of Mutant Alleles of Myospheroid, the Gene Encoding the β Subunit of the Drosophila PS Integrins

This paper presents the characterization of nine alleles of myospheroid, which encodes the beta PS subunit of the Drosophila PS integrins. On Southern blots, the mysXB87, mysXN101 and mysXR04 genes yield restriction digest patterns similar to that seen for wild-type chromosomes, however the mys1 and mysXG43 genes contain detectable deletions. mys1, mysXB87 and mysXG43 make little or no stable protein product, and genetically behave as strong lethal alleles. For the mysXN101 mutation, protein product is seen on immunoblots and a reduced amount of beta PS protein is seen at muscle attachment sites of embryos; this mutant protein retains some wild-type function, as revealed by complementation tests with weak alleles. Protein is also seen on immunoblots from mysXR04 embryos, and this allele behaves as an antimorph, being more deleterious in some crosses than the complete deficiency for the locus. mysts2 and mysnj42 are typically lethal in various combinations with other alleles at high temperatures only, but even at high physiological temperatures, neither appears to eliminate gene function completely. The complementation behaviors of mysts1 and mysts3 are quite unusual and suggest that these mutations involve regulatory phenomena. For mysts3, the data are most easily explained by postulating transvection effects at the locus. The results for mysts1 are less straightforward, but point to the possibility of a chromosome pairing-dependent negative interaction.     

Cloning and Molecular Characterization of the First Innexin of the Phylum Annelida—Expression of the Gene During Development

A novel member of the innexin family (cv-inx) has been isolated from the annelid polychaete worm Chaetopterus variopedatus using a PCR approach on genomic DNA and sequence analysis on genomic DNA clones. The gene is present in a HindIII-HindIII segment of 2250 bp containing an uninterrupted open reading frame of 1196 bp encoding a protein of 399 amino acids. The predicted protein shows the typical structural features of innexins and consensus sites for phosphorylation. Analyses on genomic DNA demonstrate that cv-inx is a single copy gene with no introns in the coding region, exactly corresponding to the cDNA sequence. The gene expression is regulated during development as shown by Northern blots analyses of the RNA and by immunoreaction with antibodies against the protein at several embryonic stages. The finding of an innexin in the phylum Annelida, outside of the Ecdysozoa clade, and its peculiar gene structure suggest the necessity to reconsider the current hypothesis on the origin and evolution of gap junctional proteins. Received: 15 December 2000 / Accepted: 27 August 2001     

Functional Identification of a Putative β-Galactosidase Gene in the Special lac Gene Cluster of Lactobacillus acidophilus

The putative β-galactosidase gene (lacZ) of Lactobacillus acidophilus has a very low degree of homology to the Escherichia coli β-galactosidase gene (lacZ) and locates in a special lac gene cluster which contains two β-galactosidase genes. No functional characteristic of the putative β-galactosidase has been described so far. In this study, the lacZ gene of L. acidophilus was hetero-expressed in E. coli and the recombinant protein was purified by a three-step procedure. The product of the lacZ gene was also extracted from L. acidophilus ATCC 4356 and active staining was carried out. The enzymatic properties of the purified recombinant LacZ were assayed. The results of hetero-expression showed the recombinant LacZ without tag had β-galactosidase activity. The purified recombinant LacZ had a specific activity of 43.2 U/mg protein. The result of active staining showed that the functional product of the lacZ gene did exist in L. acidophilus. The L. acidophilus β-galactosidase (LacZ) had an optimal pH of 6, an optimal temperature of 37°C and could hydrolyze 73% of lactose in milk in 30 h at 10°C. The L. acidophilus β-galactosidase (LacZ) was identified as cold-adapted β-galactosidase in this study for the first time, and may be useful for lactose removal from dairy products at low temperatures.     

Polymorphism of Human Glutatione S-Transferase Gene in the Populations of the Volga–Ural Region

The frequency of the GstM1 gene deletion homozygotes in eight populations of the Volga–Ural region belonging according to linguistic classification to Turkic (Bashkirs, Tatars, and Chuvashs), Finno–Ugric (Maris, Komis, Mordovians, and Udmurts), and Eastern–Slavic (Russians) ethnic groups, was examined by means of PCR technique. The frequency of the deletion homozygotes varied from 41.4% in Bashkirs to 61.3% in Mordovians. The mean deletion frequency comprised 50.1%, which was consistent with the data for European populations (²= 0.009).     

Analysis of Polymorphisms of the Huntington Disease Gene in Ethnic Populations of the Volga–Ural Region

Eleven populations of the Volga–Ural region were analyzed with respect to three intragenic polymorphisms of the Huntington disease gene (IT15), including highly polymorphic (CAG)_n and moderately polymorphic (CCG)_n of exon 1 and neutral del2642 of exon 58. In the case of (CAG)_n, 101 genotypes were observed, with genotype number varying from 15 in Southeastern Bashkirs to 34 in Mari. Allele diversity R_S ranged from 9.70 in Southeastern Bashkirs to 18.00 in Chuvash, averaging 13.79 ± 2.12. The (CAG)_n allele frequency distribution was unimodal and had a maximum at (CAG)₁₇. In the case of (CCG)_n, six alleles with 6–12 repeats were observed. R_S was 4.13 ± 0.44, ranging from 3.73 in Udmurts to 4.99 in Tatars. In the case of del2642, allele del– was detected at a frequency 0.830 in Mari to 0.932 in Udmurts. Of all Volga–Ural ethnic populations, Finno-Ugric ones proved to be most heterogeneous with respect to the three polymorphisms, whereas Turkic populations and, in particular, Bashkirs were homogeneous. Microdifferentiation of the Volga–Ural populations corresponded to the European type.     

Mutations of the Microsomal Triglyceride-Transfer–Protein Gene in Abetalipoproteinemia

Elevated plasma levels of apolipoprotein B (apoB)–containing lipoproteins constitute a major risk factor for the development of coronary heart disease. In the rare recessively inherited disorder abetalipoproteinemia (ABL) the production of apoB-containing lipoproteins is abolished, despite no abnormality of the apoB gene. In the current study we have characterized the gene encoding a microsomal triglyceride-transfer protein (MTP), localized to chromosome 4q22-24, and have identified a mutation of the MTP gene in both alleles of all individuals in a cohort of eight patients with classical ABL. Each mutant allele is predicted to encode a truncated form of MTP with a variable number of aberrant amino acids at its C-terminal end. Expression of genetically engineered forms of MTP in Cos-1 cells indicates that the C-terminal portion of MTP is necessary for triglyceride-transfer activity. Deletion of 20 amino acids from the carboxyl terminus of the 894-amino-acid protein and a missense mutation of cysteine 878 to serine both abolished activity. These results establish that defects of the MTP gene are the predominant, if not sole, cause of hereditary ABL and that an intact carboxyl terminus is necessary for activity.     

Host–Parasite Interactions and the Evolution of Gene Expression

Interactions between hosts and parasites provide an ongoing source of selection that promotes the evolution of a variety of features in the interacting species. Here, we use a genetically explicit mathematical model to explore how patterns of gene expression evolve at genetic loci responsible for host resistance and parasite infection. Our results reveal the striking yet intuitive conclusion that gene expression should evolve along very different trajectories in the two interacting species. Specifically, host resistance loci should frequently evolve to co-express alleles, whereas parasite infection loci should evolve to express only a single allele. This result arises because hosts that co-express resistance alleles are able to recognize and clear a greater diversity of parasite genotypes. By the same token, parasites that co-express antigen or elicitor alleles are more likely to be recognized and cleared by the host, and this favours the expression of only a single allele. Our model provides testable predictions that can help interpret accumulating data on expression levels for genes relevant to host−parasite interactions.     

Is There Selection for the Pace of Successive Inactivation of the arpAT Gene in Primates?

Pseudogenes have classically been considered inactive sequences evolving under neutrality. In recent years, however, a growing body of evidence is favoring the appearance of hypotheses attributing a functional role to pseudogenes. One of these hypotheses is that the silencing of a gene could produce a loss of function that could have been favored by natural selection. Here, we analyzed the pace of pseudogenization of arpAT, an L-DOPA transporter related to the neurotransmitter function of this amino acid in the brain. While active in rodent, dog, and chicken, arpAT has been silenced during primate evolution. Given the high number of inactivating mutations described in humans, it is possible that there have been selective pressures favoring this silencing. Through analysis of orthologous sequences in several primate species, we show that the silencing of arpAT occurred approximately 77 million to 90 million years ago, and that the observed mutation pattern is likely a consequence of its antiquity.     

LEA Gene Introns: is the Intron of Dehydrin Genes a Characteristic of the Serine-Segment?

Dehydrins, which belong to group 2 LEA proteins, are a family of intrinsically unstructured plant proteins that accumulate during the late stages of embryogenesis and in response to abiotic stresses. We have previously reported that the OpsDHN1 gene, encoding an SK₃-type acidic dehydrin protein from Opuntia streptacantha, contains an intron inserted within the sequence encoding the S-motif. Herein, we present an in silico analysis of intron sequences in dehydrin genes from mono- and dicotyledonous plants that reveals a preference for insertion within the nucleotide sequence encoding the S-motif. Sequence comparison of ten Dhn genes from Arabidopsis thaliana and the orthologous genes in Arabidopsis lyrata revealed that introns maintain considerable sequence identity and conserve the insertion pattern. Furthermore, syntenic regions were identified among eight orthologous genes of A. thaliana and A. lyrata, showing that correlated gene arrangements are conserved between these Arabidopsis species. Our study shows that most SKn-type dehydrins contain one intron that is conserved in phase and location; this intron is linked to the nucleotide sequence that encodes the S-motif.