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At least 253 records · Page 14Linked to original sources

Closely linked H2B genes in the marine copepod, Tigriopus californicus indicate a recent gene duplication or gene conversion event.

Two nonallelic histone gene clusters were characterized in the marine copepod, Tigriopus californicus. The DNA sequence of one of the clusters reveals six genes in the contiguous arrangement of H2B, H1, H3, H4, H2B and H2A. The order of genes within the second cluster is H3, H4, H2B and H2A. There is no evidence for the presence of an H1 gene in this cluster. Comparison of the three copepod H2B genes reveals a high degree of similarity between the 5' upstream regions and between the amino terminal halves of the two H2B genes found within the same cluster. From these data we infer that gene duplication and/or gene conversion events occurred within this cluster in the recent past.

Alleles↗

[Expression and assessment of double genes of tyrosine hydroxylase gene and aromatic L-amino acid decarboxylase gene in vitro].

The characteristic pathological changes of Parkinson s disease (PD) include a severe loss of dopamine neurons in the substantia nigra and a severe decrease in dopamine in the striatum. Since the expression of tyrosine hydroxylase (TH) and aromatic L-amino acid decarboxylase (AADC) in the biosynthetic pathway for dopamine are low, a promising approach to the gene therapy of PD is to augment the gene expression of the enzymes in the biosynthetic pathway for dopamine. In the present study, human TH and AADC genes were reconstructed into retrovirous vectors pLHCX and pLNCX(2) respectively. Then pLHCX/TH and pLNCX(2)/AADC were transfected into packaging cell line PA317 with liposome. PA317/TH and PA317/AADC were selected by different antibiotics. Gene expression was examined by methods of immunohistochemistry and in situ hybridization. The catalytic activity of two cloned gene enzymes was assessed in vitro by HPLC-EC. Immunocytochemical staining showed that TH and AADC were expressed efficiently in vitro. Both TH and AADC mRNA were transcripted in PA317 cell lines by using in situ hybridazation. HPLC-EC experiments revealed that the transfected cells produced a significantly higher level of dopamine and L-dopa than the untransfected cells. The two genetically modified cells could improve the production of L-dopa and dopamine in response to suitable substrate. The present results suggest that not only recombinant TH and AADC genes are successfully expressed in vitro, but also the enzymes have respective functional activities. These results have set up a way for in vivo gene therapy of PD with TH and AADC genes.

Aromatic-L-Amino-Acid Decarboxylases↗

Expansion of the complement receptor gene family. Identification in the mouse of two new genes related to the CR1 and CR2 gene family.

Human cDNA probes encoding the C3b/C4b complement receptor, CR1, have been used to identify, in the mouse, two new genes which are related to CR1 but which appear to encode a different protein product. These new mouse genes, arbitrarily designated mouse genes X and Y, hybridize specifically to three different cDNA probes derived from human CR1. The degree of hybridization homology between the mouse X and Y genes suggests they are very closely related to one another; however, the chromosomal localization of the mouse X gene to chromosome 8 and the mouse Y gene to chromosome 1 indicates they are distinct gene sequences. The mRNA species detected with the X and/or Y (X/Y) sequences are approximately 2000 bases in length, but vary in both quantity and size depending upon the tissue analyzed. DNA sequence analysis of a cDNA specific for the X and Y sequences indicates the mature protein(s) will contain the 60 amino acid consensus repeat characteristic of a group of other proteins including CR1, the C3d receptor (CR2), H, C4 binding protein (C4bp), the interleukin 2 (Il 2) receptor and others. The identity of the mouse X and Y genes, and the function of the proteins which they encode, is not known; however, the small size of the mRNA and the tissue specific expression suggests they do not encode mouse CR1 or CR2 but instead encode a related protein (or proteins) which is expressed in a wide variety of mouse tissues.

Amino Acid Sequence↗

[A member of the SRC gene family, the c-erbB-1 gene, is closely related to the EGF receptor gene].

An avian erythroblastosis virus, AEV-H, induces both erythroblastosis and sarcomas in susceptible chickens. Since AEV-H carries the v-erbB as a sole oncogene, the erbB gene was suggested to be responsible for the induction of these tumors. Analysis of the amino acid sequence predicted from the nucleotide sequence of the v-erbB gene revealed that the gene product has a domain characteristic for tyrosine kinase. Recently in has been suggested has the v-erbB protein is a part of the chicken EGF (epidermal growth factor) receptor. Using antibody against either v-erbB protein or EGF receptor, we also demonstrated close similarity between the two proteins. Further studies on human genomic DNA revealed that the c-erbB-1 gene, a proto-oncogene of the v-erbB gene, is the EGF receptor gene. We were also able to identify the c-erbB-2 gene that seems to code for a EGF receptor-like protein with a domain for tyrosine kinase. Finally, we would like to show that cell lines established from human squamous cell carcinom are frequently associated with amplification of the c-erbB-1/EGF receptor gene.

Alpharetrovirus↗

Two large clusters with thirty-seven transfer RNA genes adjacent to ribosomal RNA gene sets in Bacillus subtilis. Sequence and organization of trrnD and trrnE gene clusters.

Sequences of two large tRNA gene clusters (trrnD and trrnE) in Bacillus subtilis 168 revealed 16 and 21 tRNA genes, respectively, as identified by anticodon assignments. Each cluster contains upstream flanking 23 and 5 S rRNA sequences. The 23-5 S intergenic space in trrnE corresponds exactly to the analogous space in trrnB, which was previously sequenced (Wawrousek, E.F., and Hansen, J.N. (1983) J.Biol. Chem. 258, 291-298). The 5 S rRNA genes in trrnB and trrnE are B. subtilis major species; but trrnD possesses a minor species (Raue, H.A., and Planta, R. J. (1977) Mol. Gen. Genet. 156, 185-193) gene with a putative promoter that may allow differential expression with respect to the upstream rRNA gene set. Most of the tRNA genes are probably expressed as large transcriptional units, except for a LeuTTG tRNA in the trrnD cluster that appears to constitute its own operon with putative promoter and terminator sequences. Although all the amino acids are represented among the tRNA anticodons, there are few repeats of amino acid types within clusters; trrnD with 16 tRNA genes has anticodons corresponding to 15 amino acids. About two-thirds of the tRNA genes encode a 3'-terminal-CCA, and these are intermingled with those that do not, with no apparent pattern.

Anticodon↗

Cloning and characterization of a mouse gene with homology to the human von Hippel-Lindau disease tumor suppressor gene: implications for the potential organization of the human von Hippel-Lindau disease gene.

The human von Hippel-Lindau disease (VHL) gene has recently been identified and, based on the nucleotide sequence of a partial cDNA clone, has been predicted to encode a novel protein with as yet unknown functions [F. Latif et al., Science (Washington DC), 260: 1317-1320, 1993]. The length of the encoded protein and the characteristics of the cellular expressed protein are as yet unclear. Here we report the cloning and characterization of a mouse gene (mVHLh1) that is widely expressed in different mouse tissues and shares high homology with the human VHL gene. It predicts a protein 181 residues long (and/or 162 amino acids, considering a potential alternative start codon), which across a core region of approximately 140 residues displays a high degree of sequence identity (98%) to the predicted human VHL protein. High stringency DNA and RNA hybridization experiments and protein expression analyses indicate that this gene is the most highly VHL-related mouse gene, suggesting that it represents the mouse VHL gene homologue rather than a related gene sharing a conserved functional domain. These findings provide new insights into the potential organization of the VHL gene and nature of its encoded protein.

Amino Acid Sequence↗

Induction of keratinocyte proliferation and lymphocytic infiltration by in vivo introduction of the IL-6 gene into keratinocytes and possibility of keratinocyte gene therapy for inflammatory skin diseases using IL-6 mutant genes.

To understand biological function of IL-6 in the skin in vivo, we constructed a vector that strongly expressed human IL-6 in keratinocytes and introduced it into rat keratinocytes in vivo by the naked DNA method. The overexpression of IL-6 induced macroscopic erythema and histologically evident keratinocyte proliferation and lymphocytic infiltration in the treated area of rat skin. Since previous studies using IL-6 transgenic mice have not shown skin inflammation of these mice, our result provides the first evidence that IL-6 is related to the pathogenesis of inflammatory skin diseases. ELISA suggested that a certain degree of transgenic IL-6 expression in keratinocytes was required for inducing skin inflammation. Cytokine profile in rat keratinocytes after the gene introduction was examined by reverse transcriptase-PCR assay and revealed that gene expression of rat IL-1alpha and TNF-alpha showed no marked change until 24 h, whereas that of rat IL-6 and TGF-alpha increased with time. We then introduced and expressed the IL-6 mutant genes, which were designed to behave as IL-6Ralpha antagonists, and found that their ability to induce erythema was lower than that of the wild-type gene. Furthermore, preintroduction of some mutant genes delayed the erythema induced by postintroduction of the wild-type IL-6 gene, suggesting that the mutant forms of IL-6 prevent wild-type IL-6 from binding to IL-6Ralpha. This result indicates that keratinocyte gene therapy may be possible for inflammatory skin diseases using IL-6 mutant genes.

Animals↗

Frequent rearrangements of retinoic acid receptor alpha gene and myl gene, and rare mutations of RAS and FMS genes in acute promyelocytic leukemia.

To investigate leukemogenesis of acute promyelocytic leukemia (APL), we studied the involvements of retinoic acid receptor alpha (RAR alpha) and myl genes, and also the frequency of N-RAS, K-RAS, H-RAS, and FMS point mutations in sixteen patients with APL. By Southern blot analysis, the rearrangements of RAR alpha gene were detected in 13 patients (81.2%), and myl gene in 14 (87.5%). Either RAR alpha or myl gene rearrangements were found in all patients including one with normal karyotype. Breakpoints of both genes were clustered. By direct sequencing, no point mutations were found at codons 12, 13, and 61 of N-, K-, and H-RAS genes, and at codons 301 and 969 of FMS gene. These data indicate that myl-RAR alpha translocation occurs frequently in APL, whereas RAS and FMS mutations are rare in APL. It may be suggested that leukemogenesis of APL is different from other subtypes of acute myelogenous leukemia, and multistep leukemogenesis may not be a prevalent feature in APL.

Adolescent↗

The Saccharomyces cerevisiae SPT14 gene is essential for normal expression of the yeast transposon, Ty, as well as for expression of the HIS4 gene and several genes in the mating pathway.

To investigate the role of the trans-acting transcription factor encoded by the essential SPT14 (SPT = Suppressor of Ty insertion mutations) gene, we have cloned, mapped and sequenced the gene. From the analysis of the effect of spt14 mutations on expression of various genes, we conclude that the SPT14 product has an important role in activation of Ty transcription as well as in the regulation of other genes including HIS4 and several of the a- and alpha-specific mating type genes. Similarities in the phenotypes of spt14 and spt13 mutants (suppression of Ty insertion mutations but not delta insertion mutations), lead to the suggestion that the SPT14 gene and the previously characterized SPT13/GAL11 gene might encode transcriptional regulators with related functions. Our current findings show that in contrast to SPT13/GAL11, which appears negatively to regulate Ty transcription, SPT14 plays a role in the activation of Ty transcription. Thus, despite the similarities in the suppression phenotype exhibited by spt13 and spt14 mutants, SPT13/GAL11 and SPT14 probably differ in their transcriptional roles.

Amino Acid Sequence↗

Gene within gene configuration and expression of the Drosophila melanogaster genes lethal(2) neighbour of tid [l(2)not] and lethal(2) relative of tid[l(2)rot].

In this paper, we describe the structure and temporal expression pattern of the Drosophila melanogaster genes l(2)not and l(2)rot located at locus 59F5 vis à vis the tumor suppressor gene l(2)tid described previously and exhibiting a gene within gene configuration. The l(2)not protein coding region, 1530 nt, is divided into two exons by an intron, 2645 nt, harboring the genes l(2)rot, co-transcribed from the same DNA strand, and l(2)tid, co-transcribed from the opposite DNA strand, located vis à vis. To determine proteins encoded by the genes described in this study polyclonal rabbit antibodies (Ab), anti-Not and anti-Rot, were generated. Immunostaining of developmental Western blots with the anti-Not Ab resulted in the identification of a 45-kDa protein, Not45, which is smaller than the Not56 protein predicted from the sequence. Its localization in endoplasmic reticulum (ER) was established by immunoelectron microscopy of Drosophila melanogaster Schneider 2 cells. Not45 shows significant homology to yeast ALG3 protein acting as a dolichol mannosyltransferase in the asparagine-linked glycosylation. It is synthesized ubiquitously throughout embryonic life. The protein predicted from the l(2)rot sequence, Rot57, shows a homology to the NS2B protein of the yellow fever virus1 (yefv1). The results of l(2)rot RNA analysis by developmental Northern blot and by in situ RNA localization, as well as the results of the protein analysis via Western blot and immunohistochemistry suggest that l(2)rot is transcribed but not translated. Since RNAs encoded by the genes l(2)tid and l(2)rot are complementary and l(2)rot is presumably not translated we performed preliminary experiments on the function of the l(2)rot RNA as a natural antisense RNA (asRNA) regulator of l(2)tid expression, expressed in the same temporal and spatial manner as the l(2)tid- and l(2)not RNA. l(2)tid knock-out by antisense RNA yielded late embryonic lethality resulting from multiple morphogenetic defects.

Amino Acid Sequence↗

A set of human putative lymphocyte G0/G1 switch genes includes genes homologous to rodent cytokine and zinc finger protein-encoding genes.

Lymphocyte G0/G1 switch genes (G0S genes) are potential oncogenes and may regulate, be regulated by, or be coordinately regulated with, latent lymphotropic viruses. To identify these genes, a cDNA library was prepared from blood mononuclear cells that had been cultured for 2 hr with a T-cell mitogen (lectin) and cycloheximide. Eight differentially hybridizing recombinants were characterized by RNA and DNA blotting and sequencing. One cDNA (G0S7) corresponded to the oncogene c-fos. Another cDNA (G0S19) was homologous (70%) to a cDNA encoding a murine inhibitor of stem cell proliferation (the cytokine MIP1 alpha) and, less closely, to other members of the "small inducible" secreted protein-encoding gene family. Whereas cDNA hybridization to genomic DNA blots indicated a small subfamily of G0S19 genes, simple patterns of bands indicated that most cDNAs, including G0S30 cDNA, corresponded to single-copy genes. The 3' noncoding sequence of G0S30 cDNA was homologous (87-89%) to the 3' noncoding sequences of certain rodent genes (NGFI-A, Krox24, EGR1) that encode zinc finger proteins (putative transcriptional regulators). This degree of evolutionary conservation suggests an important function for the 3' noncoding region. The 3' noncoding regions of some cDNAs contained the TTATTTAT (mRNA destabilization) element. The corresponding RNAs each formed doublets in agarose gels. Previous studies of c-fos RNA from HeLa cells indicate that this is due to cycloheximide-dependent stabilization of poly(A) tails. Our results reveal the power of cycloheximide enrichment in isolating what would appear to be significant low-abundance mRNAs.

Base Sequence↗

Gene expression patterns during swarming in Salmonella typhimurium: genes specific to surface growth and putative new motility and pathogenicity genes.

Swarming is a specialized form of surface motility displayed by several flagellated bacterial genera, which shares features with other surface phenomenon such as biofilm formation and host invasion. Swarmer cells are generally more flagellated and longer than vegetative cells of the same species propagated in liquid media, and move within an encasement of polysaccharide 'slime'. Signals and signalling pathways controlling swarm cell differentiation are largely unknown. In order to test whether there is a genetic programme specific to swarming, we have determined global gene expression profiles of Salmonella typhimurium over an 8 h time course during swarming, and compared the microarray data with a similar time course of growth in liquid media as well as on harder agar where the bacteria do not swarm. Our data show that bacteria growing on the surface of agar have a markedly different physiology from those in broth, as judged by differential regulation of nearly one-third of the functional genome. The large number of genes showing surface-specific upregulation included those for lipopolysaccharide synthesis, iron metabolism and type III secretion. Although swarming-specific induction of flagellar gene expression was not generally apparent, genes for iron metabolism were strongly induced specifically on swarm agar. Surface-dependent regulation of many virulence genes suggests that growth on an agar surface could serve as a model for gene expression during the initial stages of host infection. Based on cluster analysis of distinctive expression patterns, we report here the identification of putative new genes involved in motility and virulence.

Bacterial Proteins↗

Fructosyltransferase and invertase genes evolved by gene duplication and rearrangements: rice, perennial ryegrass, and wheat gene families.

The invertase enzyme family is responsible for carbohydrate metabolism in rice, perennial ryegrass, and wheat. Fructan molecules accumulate in cell vacuoles of perennial ryegrass and wheat and are associated with abiotic stress tolerance. High levels of amino acid similarity between the fructosyltransferases responsible for fructan accumulation indicates that they may have evolved from invertase-like ancestral genes. In this study, we have applied comparative genomics to determine the mechanisms that lead to the evolution of fructosytransferase and invertase genes in rice, perennial ryegrass, and wheat. Duplications and rearrangements have been inferred to generate variant forms of the rice invertases since divergence from a common grass progenitor. The occurrence of multiple copies of fructosyltransferase genes indicated that duplication events continued during evolution of the wheat and perennial ryegrass lineages. Further gene rearrangements were evident in perennial ryegrass genes, albeit at a reduced level compared with the rice invertases. Gene orthologs were largely static after duplication during evolution of the wheat lineage. This study details evolutionary events that contribute to fructosyltransferase and invertase gene variation in grasses.

Amino Acid Sequence↗

Expression of alpha-fetoprotein and albumin genes in human hepatocellular carcinomas: limitations in the application of the genes for targeting human hepatocellular carcinoma in gene therapy.

For an approach of gene therapy for hepatocellular carcinoma (HCC), transcriptional regulatory sequence (TRS) of either alpha-fetoprotein (AFP) or albumin has been used for targeting cancer cells. To examine the feasibility of using TRSs of these genes for possible gene therapy of HCCs, the cellular distribution of AFP and albumin gene transcripts was studied in 25 cases of surgically removed human HCCs. AFP gene expression was observed in HCC nodules of 13 cases. The expression in HCC was heterogeneous, and the distribution of the transcripts was mostly sparse and spotty. The higher the serum AFP levels, the larger population of the AFP-expressing HCC cells tended to reflect. In noncancerous liver, a slight AFP expression was found by Northern blot analysis, but the transcripts were not detected in the liver sections. In contrast, albumin expression was found in all HCCs as well as in noncancerous hepatocytes. In HCC, the transcripts for albumin were distributed in cancer cells, and the expression varied with nodules. There were more albumin-expressing cancer cells than the AFP-expressing cells. Albumin expression was retained even in poorly differentiated HCC, although the intensity of the signal was not as strong as in more-differentiated HCCs. Metastatic HCC nodules revealed transcripts for both AFP and albumin genes, and those were clearly recognized in the lung tissue. These results suggest that, for gene therapy for HCCs, neither AFP nor albumin are ideal options for targeting HCC cells. AFP-TRS may be used as a transcriptional regulator in selected cases in which AFP gene expression is observed in the cancer nodules. The serum AFP level appears to be an important indicator in selecting cases. Albumin-TRS in conjunction with retroviral vector might be used in limited cases such as HCCs with no AFP expression. However, careful consideration must be taken, because albumin is constitutively expressed in normal hepatocytes, and AFP-expressing nonmalignant progenitor cells possibly exist.

Adult↗

Hepatoma-specific gene therapy through retrovirus-mediated and targeted gene transfer using an adenovirus carrying the ecotropic receptor gene.

Tissue-specific transcriptional regulatory sequences have been inserted in retrovirus vectors for therapeutic gene expression in cancer gene therapy. However, the transcriptional activity of these sequences is generally low, and the proviral DNA integration appears to increase the possibility of genomic DNA alteration in nontarget cells. Therefore, retrovirus-mediated targeted gene transduction into human hepatoma cells was evaluated using transient expression of an ecotropic receptor gene, mouse cationic amino acid transporter-1 (MCAT-1). Two recombinant adenoviruses, AxCAMCAT and AxAFMCAT, carrying the MCAT-1 gene under the control of the CAG and human alpha-fetoprotein (AFP) promoter, respectively, were generated. The preinfection with AxCAMCAT allowed highly efficient ecotropic retroviral infection of human cells. In addition, after AxAFMCAT infection, retroviral infection occurred only in AFP-producing hepatoma cells, resulting in selective cytotoxicity induced by the herpes simplex virus thymidine kinase (HSV-tk)/ganciclovir (GCV) system. Transient expression of the MCAT-1 gene under the control of the AFP promoter permits ecotropic retrovirus-mediated gene transduction specifically in AFP-producing human hepatoma cells, resulting in selective induction of the suicide killing effect while the therapeutic gene is driven from ubiquitously expressed promoters.

3T3 Cells↗

Molecular characterization of the mouse p47-phox (Ncf1) gene and comparative analysis of the mouse p47-phox (Ncf1) gene to the human NCF1 gene.

The cytosolic factor p47-phox (NCF1) is a key component of the phagocyte NADPH-oxidase system, critical for microbicidal activity. The human p47-phox gene has been well characterized and resides on chromosome 7q11. Here we describe the molecular characterization of the mouse ortholog (Ncf1), which maps to distal chromosome 5, and compare the structure of the genes, commenting on the degree of homology. The mouse and human genes contain the same number of exons and introns, but the mouse gene is more compact (7.8 kb versus 15.2 kb). A percentage identity plot analysis comparing the human and mouse genes indicates that sequence homology is generally restricted to exons and does not include any large segment of introns or the 5' flanking sequence. The mouse gene also contains notably fewer repetitive elements than its human counterpart (34% versus 50%). The start of transcription of the mouse gene has been localized to within 12 nucleotides of the translation start site, similar to the human ortholog. Our findings provide an important foundation for investigating the evolutionary history of the p47-phox gene, particularly as it relates to understanding the molecular basis of the p47-phox-deficient autosomal recessive form of chronic granulomatous disease.

5' Untranslated Regions↗

Molecular evolution of daphnia immunity genes: polymorphism in a gram-negative binding protein gene and an alpha-2-macroglobulin gene.

Studies of DNA polymorphism have shown that some immune system genes of mammals and plants are exceptionally diverse, indicating that coevolution between these taxa and their parasites mediates positive selective sweeps and/or balancing selection. The genes of the arthropod immune system remain comparatively unstudied. We isolated two putative immune system genes from the cladoceran crustacean Daphnia and examined DNA sequence diversity. For one gene, encoding a putative gram-negative binding protein, we found evidence of only purifying selection, indicating that this gene is under strong functional constraint and that selection acts to eliminate amino acid variation. For another gene, encoding a putative alpha-2-macroglobulin, we found evidence of positive selection, indicating the possible involvement of this gene in a host-parasite arms race. We discuss the assumed function of these genes and offer speculation regarding which components of the arthropod immune system might experience diversifying adaptive evolution.

Amino Acid Sequence↗

Gene structure of rat BAF60b, a component of mammalian SW1/SNF complexes, and its physical linkage to the growth hormone gene and transcription factor SUG/proteasome p45 gene.

In the +27.6 to +36.7 kb downstream region from the transcriptional start site of the rat growth hormone (GH) gene, a gene encoding BAF60b, a component of mammalian SWI/SNF complexes, was found to have the same transcriptional orientation as the GH gene. The 5' end of the BAF60b gene was heterogeneous and the longest gene was 9060 bp long with 13 exons. The largest of all exons was estimated to be 2774 bases. Deduced rat BAF60b protein was made of 531 amino acids and its amino acid sequence was 97% identical with the human counterpart. No TATA box was found up to the -100 bp region but five GC boxes corresponding to the Sp1 binding site were observed up to 640 bp upstream from the transcriptional start site. Sixty-three bases downstream from the BAF60b gene, the polyadenylation site of the gene encoding transcription factor SUG/proteasome p45, whose expression is constant in many tissues, was identified. The BAF60b gene was expressed as 3.0 kb poly(A)-rich RNA in seven tissues and one cell line from rat but its expression varied considerably according to the tissue.

ATPases Associated with Diverse Cellular Activitie↗