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PubMed1995

Inefficient complementation activity of poliovirus 2C and 3D proteins for rescue of lethal mutations.

Poliovirus (PV) 2C protein is a nonstructural polypeptide involved in viral RNA replication, whose biochemical activity(ies) in this process has not been defined. By using site-directed mutagenesis, it was shown previously that disruption of nucleotide-binding motifs present in this protein abolished viral RNA synthesis (C. Mirzayan and E. Wimmer, Virology 189:547-555, 1992; N. L. Teterina, K. M. Kean, E. Gorbalenya, V. I. Agol, and M. Girard, J. Gen. Virol. 73:1977-1986, 1992). We have tested whether PV 2C or 2BC protein provided in trans could rescue the replication of these mutated genomes. Rescuing proteins were provided either by cotransfection with helper chimeric PV-coxsackievirus genomes or by expression in cells with a vaccinia virus-T7 RNA polymerase transient-expression system. We report here that replication of mutated RNAs genomes was poorly supported in trans both by helper genomes and by expressed 2C or 2BC proteins. Similarly, very inefficient complementation was observed for two mutated genomes with lethal lesions in 3D polymerase coding sequence. Our results indicate that poliovirus RNA replication shows marked preference for proteins contributed in cis.

Amino Acid Sequence↗
PubMed1998

DNA mutagenesis and repair in UV-irradiated E. coli K-12 under condition of mutation frequency decline.

This paper shows that mutation frequency decline (MFD) occurs in UV-irradiated and transiently starved Escherichia coli K-12 strain AB1157. This effect involves preferential repair of pre-mutagenic lesions situated on the transcribed strand, and is mfd-, and uvrA-dependent. Mutations were tested by measuring reversion of argE3(ochre) to Arg+ and the types of reversions and the mutational specificity were defined. UV-irradiation induces reversions to Arg+ predominantly by forming ochre suppressors, supB and, much more rarely, supE(ochre). In irradiated and then transiently starved cells, the ratio of supB/supE(ochre) is reversed, and supE(ochre) suppressors predominate. Back reversions at the argE3 site occurs only in bacteria defective in UvrABC-excinuclease. The introduction of a Tn10 transposon makes bacteria more sensitive to UV-irradiation and diminishes the frequency of reversions. Transformation with pGW2123, a umuD'C-bearing plasmid, recovers and enhances the frequency of reversion but has no effect on post-UV-survival of the bacteria.

Arginine↗
PubMed1999

Expression analysis of four endoglin missense mutations suggests that haploinsufficiency is the predominant mechanism for hereditary hemorrhagic telangiectasia type 1.

ENDOGLIN codes for a homodimeric membrane glycoprotein that interacts with receptors for members of the TGF-beta superfamily and is the gene mutated in the autosomal dominant vascular disorder hereditary hemorrhagic telangiectasia type 1 (HHT1). We recently demonstrated that functional endoglin was expressed at half levels on human umbilical vein endothelial cells (HUVECs) and peripheral blood activated monocytes from HHT1 patients. Two types of mutant protein were previously analyzed, the product of an exon 3 skip which was expressed as a transient intracellular species and prematurely truncated proteins that were undetectable in patient samples. Here we report the analysis of four proteins resulting from point mutations, with missense codons G52V and C53R in exon 2, W149C in exon 4 and L221P in exon 5. Metabolic labeling of activated monocytes from confirmed, clinically affected patients revealed reduced expression of fully processed normal endoglin in all cases. Pulse-chase analysis with HUVECs from a newborn with the C53R substitution indicated that mutant endoglin remained intracellular as a precursor form and did not impair processing of the normal protein. Biotinylation of cell surface proteins, metabolic labeling and pulse-chase analysis revealed that none of the engineered missense mutants was significantly expressed at the surface of COS-1 transfectants. Thus, these four HHT1 missense mutations lead to transient intracellular species which cannot interfere with normal endoglin function. These data suggest that haploinsufficiency, leading to reduced levels of one of the major surface glyco-proteins of vascular endothelium, is the predominant mechanism underlying the HHT1 phenotype.

Animals↗
PubMed2005

Insulin activates the rat sterol-regulatory-element-binding protein 1c (SREBP-1c) promoter through the combinatorial actions of SREBP, LXR, Sp-1 and NF-Y cis-acting elements.

The enhanced synthesis of fatty acids in the liver and adipose tissue in response to insulin is critically dependent on the transcription factor SREBP-1c (sterol-regulatory-element-binding protein 1c). Insulin increases the expression of the SREBP-1c gene in intact liver and in hepatocytes cultured in vitro. To learn the mechanism of this stimulation, we analysed the activation of the rat SREBP-1c promoter and its truncated or mutated congeners driving a luciferase reporter gene in transiently transfected rat hepatocytes. The rat SREBP-1c promoter contains binding sites for LXR (liver X receptor), Sp1, NF-Y (nuclear factor-Y) and SREBP itself. We have found that each of these sites is required for the full stimulatory response of the SREBP-1c promoter to insulin. Mutation of either the putative LXREs (LXR response elements) or the SRE (sterol response element) in the proximal SREBP-1c promoter reduced the stimulatory effect of insulin by about 50%. Insulin and the LXR agonist TO901317 increased the association of SREBP-1 with the SREBP-1c promoter. Ectopic expression of LXRalpha or SREBP-1c increased activity of the SREBP-1c promoter, and this effect is further enhanced by insulin. The Sp1 and NF-Y sites adjacent to the SRE are also required for full activation of the SREBP-1c promoter by insulin. We propose that the combined actions of the SRE, LXREs, Sp1 and NF-Y elements constitute an insulin-responsive cis-acting unit of the SREBP-1c gene in the liver.

Animals↗
PubMed2001

Independent and coordinate effects of ADP-ribosyltransferase and GTPase-activating activities of exoenzyme S on HT-29 epithelial cell function.

Type III-mediated translocation of exoenzyme S (ExoS) into HT-29 epithelial cells by Pseudomonas aeruginosa causes complex alterations in cell function, including inhibition of DNA synthesis, altered cytoskeletal structure, loss of readherence, microvillus effacement, and interruption of signal transduction. ExoS is a bifunctional protein having both GTPase-activating (GAP) and ADP-ribosyltransferase (ADPRT) functional domains. Comparisons of alterations in HT-29 cell function caused by P. aeruginosa strains that translocate ExoS having GAP or ADPRT mutations allowed the independent and coordinate functions of the two activities to be assessed. An E381A ADPRT mutation revealed that ExoS ADPRT activity was required for effects of ExoS on DNA synthesis and long-term cell rounding. Conversely, the R146A GAP mutation appeared to have little impact on the cellular effects of ExoS. While transient cell rounding was detected following exposure to the E381A mutant, this rounding was eliminated by an E379A-E381A ADPRT double mutation, implying that residual ADPRT activity, rather than GAP activity, was effecting transient cell rounding by the E381A mutant. To explore this possibility, E381A and R146A-E381A mutants were examined for their ability to ADP-ribosylate Ras in vitro or in vivo. While no ADP-ribosylation of Ras was detected by either mutant in vitro, both mutants were able to modify Ras when translocated by the bacteria, with the R146A-E381A mutant causing more efficient modification than the E381A mutant, in association with increased inhibition of DNA synthesis. Comparisons of Ras ADP-ribosylation by wild-type and E381A mutant ExoS by two-dimensional electrophoresis found the former to ADP-ribosylate Ras at two sites, while the latter modified Ras only once. These studies draw attention to the key role of ExoS ADPRT activity in causing the effects of bacterially translocated ExoS on DNA synthesis and cell rounding. In addition, the studies provide insight into the enhancement of ExoS ADPRT activity within the eukaryotic cell microenvironment and into possible modulatory roles that the GAP and ADPRT domains might have on the function of each other.

ADP Ribose Transferases↗
PubMed1993

Evolution in vitro: analysis of a lineage of ribozymes.

BACKGROUND: Catalytic RNAs, or ribozymes, possessing both a genotype and a phenotype, are ideal molecules for evolution experiments in vitro. A large, heterogeneous pool of RNAs can be subjected to multiple rounds of selection, amplification and mutation, leading to the development of variants that have some desired phenotype. Such experiments allow the investigator to correlate specific genetic changes with quantifiable alterations of the catalytic properties of the RNA. In addition, patterns of evolutionary change can be discerned through a detailed examination of the genotypic composition of the evolving RNA population. RESULTS: Beginning with a pool of 10(13) variants of the Tetrahymena ribozyme, we carried out in vitro evolution experiments that led to the generation of ribozymes with the ability to cleave an RNA substrate in the presence of Ca2+ ions, an activity that does not exist for the wild-type molecule. Over the course of 12 generations, a seven-error variant emerged that has substantial Ca(2+)-dependent RNA-cleavage activity. Advantageous mutations increased in frequency in the population according to three distinct dynamics--logarithmic, linear and transient. Through a comparative analysis of 31 individual variants, we infer how certain mutations influence the catalytic properties of the ribozyme. CONCLUSIONS: In vitro evolution experiments make it possible to elucidate important aspects of both evolutionary biology and structural biochemistry on a reasonable short time scale.

Animals↗
PubMedDate not supplied

Identification of two novel mutations in OCTN2 of three patients with systemic carnitine deficiency.

Systemic carnitine deficiency is a potentially lethal, autosomal recessive disorder characterized by cardiomyopathy, myopathy, recurrent episodes of hypoketotic hypoglycemia, hyperammonemia, and failure to thrive. This form of carnitine deficiency is caused by a defect in the active cellular uptake of carnitine, and the gene encoding the high affinity carnitine transporter OCTN2 has recently been shown to be mutated in patients suffering from this disorder. Here, we report the underlying molecular defect in three unrelated patients. Two patients were homozygous for the same missense mutation 632A-->G, which changes the tyrosine at amino acid position 211 into a cysteine (Y211C). The third patient was homozygous for a nonsense mutation, 844C-->T, which converts the arginine at amino acid position 282 into a stop codon (R282X). Reintroduction of wild-type OCTN2 cDNA into fibroblasts of the three patients by transient transfection restored the cellular carnitine uptake, confirming that mutations in OCTN2 are the cause of systemic carnitine deficiency.

Base Sequence↗
PubMed2004

Glycogen storage disease type Ia in Argentina: two novel glucose-6-phosphatase mutations affecting protein stability.

Glycogen storage disease type Ia (GSD-Ia) is caused by deleterious mutations in the glucose-6-phosphatase gene (G6PC). A molecular study of this gene was carried out in 11 Argentinean patients from 8 unrelated families. Four missense (p.Gln54Pro, p.Arg83Cys, p.Thr16Arg, and p.Tyr209Cys) and one deletion (c.79delC) mutations have been identified. Two novel mutations, p.Thr16Arg (c.47C>G) located within the amino-terminal domain and p.Tyr209Cys (c.626A>G) situated in the sixth transmembrane helix, were uncovered in this study. Site-directed mutagenesis and transient expression assays demonstrated that both p.Thr16Arg and p.Tyr209Cys mutations abolished enzymatic activity as well as reduced G6Pase stability.

Animals↗
PubMed1988

Fos C-terminal mutations block down-regulation of c-fos transcription following serum stimulation.

Transient accumulation of c-fos RNA following serum stimulation requires both a conserved 5' regulatory element and sequences at the 3' end of the gene. Here we show that mutations at the C terminus of Fos protein, of the type found in a virally-transduced actively transforming Fos variant, prevent the rapid down-regulation of c-fos transcription that occurs following serum-induced activation. Fos mutants that prevent down-regulation are dominant, acting in trans to prevent down-regulation of a co-transfected c-fos gene. Co-transfection experiments suggest that this effect is mediated by multiple sequence elements in the 5'-flanking region. Analysis of different Fos mutants showed that replacement of Fos sequences C-terminal to amino acid 337 with heterologous polypeptide, rather than simple truncation of the protein, is required to produce mutants defective in down-regulation. The results are discussed with reference to transformation by Fos.

Amino Acid Sequence↗
PubMed1982

Two cell division cycle mutants of Saccharomyces cerevisiae are defective in transmission of mitochondria to zygotes.

Mutations in CDC genes of S. cerevisiae disrupt the cell cycle at specific stages. The experiments reported here demonstrate that two CDC genes, CDC5 and CDC27, are necessary for mitochondrial segregation as well as for nuclear division. The defect in the transmission of mitochondria was revealed by the examination of uninucleate and binucleate progeny of transient heterokaryons generated by using the kar1-1 mutation that disrupts nuclear fusion. One of the parents lacked mitochondrial DNA (rho(0)) whereas the other parent had functional mitochondria (rho(+)). When the parents of the heterokaryon were both wild-type (CDC), nearly all progeny received mitochondria at 21 degrees and at 34 degrees . Thirty-four of the 36 cdc mutations tested had no defect in transmission of mitochondria to zygotic progeny in crosses in which one parent was a cdc mutant and the other parent was not (CDC). However, the cdc5 and cdc27 mutations prevented the transmission of mitochondria to cdc progeny at 34 degrees but not at 21 degrees ; CDC progeny received mitochondria at either temperature. This defect was observed in crosses of cdc5 or cdc27 by wild-type cells regardless of which parent donated mitochondria to the zygote. The defect in mitochondrial transmission cosegregated in meiotic tetrads with the defect in mitosis demonstrating that both are likely to be caused by the same temperature-sensitive mutation. These results indicate that the CDC5 and CDC27 gene products are essential in two motility-related processes: mitochondrial movement from the zygote to the progeny and in mitosis.-Furthermore, the results suggest that the function performed by the CDC5 and CDC27 gene products for mitochondrial transmission differ in some fundamental way from the function performed for mitosis. The function necessary for mitosis can be supplied to the cdc5 (or cdc27) nucleus by the CDC5 (or CDC27) nucleus in the same heterokaryon but the function necessary for mitochondrial transmission cannot. Perhaps the function needed for mitochondrial transmission must be performed in the cell cycle preceding the actual segregation of mitochondria whereas the function needed for nuclear segregation can be performed at the time that mitosis occurs.

Cell Cycle↗
PubMed2002

Enhanced production of IL-18 in butyrate-treated intestinal epithelium by stimulation of the proximal promoter region.

Expression of IL-18 in intestinal epithelial cells (IEC) has been implicated in Th1 cell-mediated chronic intestinal inflammation and anti-tumor immunity. However, physiological regulatory factors have not been identified. Besides their effects on proliferation and restitution, immunomodulatory functions have been attributed to short chain fatty acids (SCFA). We investigated the effect of SCFA (butyrate, propionate, acetate) on expression of IL-18 in IEC in vitro and in vivo. Expression of IL-18 mRNA and protein in human carcinoma-derived HT-29 and Caco-2 cells was analyzed by reverse transcription-PCR and Western blot. Transcriptional regulation of IL-18 gene expression was determined by transient transfection of wild-type and mutated IL-18 promoter. Further, in vivo expression of IL-18 in the intestine from butyrate-treated and untreated mice was assessed by immunohistochemistry. IL-18 mRNA and the IL-18 protein were expressed in IEC, while IL-18 secretion was not observed. Butyrate and acetate increased intracellular IL-18 content in a time- and dose-dependent fashion. In contrast to proinflammatory stimuli butyrate potently activated the IL-18 promoter, indicating that IL-18 is regulated at the transcriptional level by SCFA. Furthermore, a 108-bp sequence in the proximal region was identified to be essential for IL-18 promoter activation by butyrate. As proof of principle butyrate effects were confirmed in vivo by demonstration of increased IL-18 protein expression in IEC from butyrate-treated mice. In conclusion, SCFA up-regulate IL-18 protein expression in IEC, suggesting a potential regulatory contribution of these luminal constituents to T cell mediated inflammatory and neoplastic intestinal conditions.

Acetates↗
PubMed1998

Ah receptor and estrogen receptor-dependent modulation of gene expression by extracts of diesel exhaust particles.

The ability of a methylene chloride extract of diesel exhaust particle (EDEP) to activate the aryl hydrocarbon receptor (AhR), bind to and activate the estrogen receptor (ER), and induce gene expression mediated via these nuclear receptors was examined in Hepa1c1c7 mouse hepatoma and MCF-7 human breast cancer cells. EDEP was able to induce a protein-DNA complex by gel retardation assays using a [gamma-32P]dATP-labeled dioxin response element (DRE). This complex could be effectively competed by a 150-fold excess of unlabeled DRE but not by a 150-fold excess of unlabeled mutated DRE. In Hepa1c1c7 cells that were transiently transfected with a DRE-regulated luciferase reporter gene, 4.6 ng/microliter EDEP treatment for 24 h resulted in a 22-fold induction of luciferase activity. In the same cell line, ethoxyresorufin-O-deethylase activity was significantly induced 20-fold following 24 h treatment with 4.6 ng/microliter EDEP. Using a competitive ligand binding assay, EDEP displaced bound tritiated E2 from the rat uterine ER in a dose-dependent manner with an IC50 of approximately 100 ng/microliter compared to the IC50 of E2, which was approximately 4.4x10(-4) ng/microliter (1.6 nM). In MCF-7 human breast cancer cells transiently transfected with a Gal4-regulated luciferase reporter gene (17m5-G-Luc) and a chimeric ER (Gal4-HEG0), treatment with 4.6 ng/microliter EDEP for 24 h resulted in a three-fold increase in luciferase activity (P<0.01) compared with the seven-fold increase observed with E2. This study demonstrates that EDEP is able to activate the AhR and ER and induce transcription of reporter genes regulated by these receptors' DNA response elements. Further study is required to identify the individual compound(s) responsible for the observed activity.

Animals↗
PubMed1999

Trypanosoma cruzi: suppression of tuzin gene expression by its 5'-UTR and spliced leader addition site.

Teixeira, S. M. R., Kirchhoff, L. V., and Donelson, J. E. 1999. Trypanosoma cruzi: Suppression of tuzin gene expression by its 5'-UTR and spliced leader addition site. Experimental Parasitology 93, 143-151. The genome of the protozoan parasite Trypanosoma cruzi contains a tandemly repeated array of two alternating genes, one encoding amastin and the other encoding tuzin. Amastin is an abundant amastigote surface protein, whereas tuzin is thought to be a rare protein whose location and function are unknown. The 137-nucleotide 5' untranslated region (5'-UTR) of the tuzin mRNA has a 22-codon open translation reading frame containing 3 methionine codons followed by a stop codon that overlaps the methionine start codon of the tuzin coding region. A fragment containing the tuzin 5'-UTR and upstream intergenic region was placed in front of a luciferase reporter gene in a plasmid for transient transfection assays of luciferase activity. By mutating the three upstream ATGs in the tuzin 5'-UTR and replacing the tuzin spliced leader (SL) acceptor site with that of the amastin gene, we found that the 22-codon reading frame and the tuzin SL acceptor site combine to substantially reduce expression of the luciferase gene. These results indicate that expression of the multicopy tuzin gene is posttranscriptionally suppressed by both inefficient RNA processing and poor translation initiation, resulting in a low level of tuzin.

5' Untranslated Regions↗
PubMed1999

Structure and evolution of the human SPRR3 gene: implications for function and regulation.

SPRR3, a member of the SPRR family of cornified envelope precursor proteins, is expressed in oral and esophageal epithelia, where it is strictly linked to keratinocyte terminal differentiation. This gene is characterized by intragenic duplications that have created the characteristic proline-rich repeats in the coding sequence, an alternative noncoding exon, and a 200-bp polypyrimidine tract in the promoter region. Mutational analysis of the promoter region and transient transfection in normal human keratinocytes showed that in addition to the polypyrimidine tract, multiple regulatory elements are involved in differentiation-specific expression. These elements include a high-affinity Ets binding site bound by ESE-1, an AP-1 site (TRE) recognized by the Jun/Fos family of transcription factors, and an ATF/CRE bound by Jun/Fos and ATF factors. The repositioning of the SPRR3 Ets binding site during evolution has a major effect on the relative contribution of this site to promoter activity.

Activating Transcription Factor 2↗
PubMed2000

Induction of LPL gene expression by sterols is mediated by a sterol regulatory element and is independent of the presence of multiple E boxes.

Overexpression of the adipocyte differentiation and determination factor-1 (ADD-1) or sterol regulatory element binding protein-1 (SREBP-1) induces the expression of numerous genes involved in lipid metabolism, including lipoprotein lipase (LPL). Therefore, we investigated whether LPL gene expression is controlled by changes in cellular cholesterol concentration and determined the molecular pathways involved. Cholesterol depletion of culture medium resulted in a significant induction of LPL mRNA in the 3T3-L1 preadipocyte cell line, whereas addition of cholesterol reduced LPL mRNA expression to basal levels. Similar to the expression of the endogenous LPL gene, the activity of the human LPL gene promoter was enhanced by cholesterol depletion in transient transfection assays, whereas addition of cholesterol caused a reversal of its induction. The effect of cholesterol depletion upon the human LPL gene promoter was mimicked by cotransfection of expression constructs encoding the nuclear form of SREBP-1a, -1c (also called ADD-1) and SREBP-2. Bioinformatic analysis demonstrated the presence of 3 potential sterol regulatory elements (SRE) and 3 ADD-1 binding sequences (ABS), also known as E-box motifs. Using a combination of in vitro protein-DNA binding assays and transient transfection assays of reporter constructs containing mutations in each individual site, a sequence element, termed LPL-SRE2 (SRE2), was shown to be the principal site conferring sterol responsiveness upon the LPL promoter. These data furthermore underscore the importance of SRE sites relative to E-boxes in the regulation of LPL gene expression by sterols and demonstrate that sterols contribute to the control of triglyceride metabolism via binding of SREBP to the LPL regulatory sequences.

Adipocytes↗
PubMed2002

Interleukin-8 gene regulation in intestinal epithelial cells infected with rotavirus: role of viral-induced IkappaB kinase activation.

Rotavirus is the major etiologic agent of diarrhea in children and the most common cause of severe pediatric gastroenteritis. Rotavirus infection is limited to mature enterocytes that line the villi of the small intestine. Gut epithelial cells, upon infection and cytokine stimulation, are able to produce chemokines, a family of small chemotactic cytokines that regulate the migration and activation of leukocytes. We have previously shown that rotavirus infection of the intestinal epithelial cell line HT-29 induces increased expression of the CXC chemokine interleukin- (IL) 8. Mechanisms responsible for the transcriptional regulation of the IL-8 gene in intestinal epithelial cells during viral infections have not been fully elucidated. Therefore, the purpose of this study was to define the molecular mechanisms of IL-8 gene expression in HT-29 cells infected with rotavirus. Transient transfection analysis of 5' deletions and mutations of the IL-8 promoter driving expression of luciferase reporter gene indicates that the activating protein- (AP) 1 and nuclear factor- (NF) kappaB elements are necessary for IL-8 promoter activation during rotavirus infection. The importance of NF-kappaB activation for IL-8 gene expression was further demonstrated by the inhibition of rotavirus-induced IL-8 gene transcription and protein synthesis following blockade of degradation of the NF-kappaB cytoplasmic inhibitor IkappaB-alpha. Rotavirus infection of HT-29-induced IkappaB kinase (IKK) activation and overexpression of a dominant negative mutant of IKK-beta greatly reduced rotavirus-induced IL-8 promoter activation and NF-kappaB-driven transcription, indicating that IKK is involved in rotavirus-induced IL-8 gene expression and NF-kappaB activation.

Epithelial Cells↗
PubMed2005

DcMYB1 acts as a transcriptional activator of the carrot phenylalanine ammonia-lyase gene (DcPAL1) in response to elicitor treatment, UV-B irradiation and the dilution effect.

Expression of a carrot phenylalanine ammonia-lyase (PAL) gene (DcPAL1) in suspension-cultured carrot cells is induced by treatment with a fungal elicitor, ultraviolet B (UV-B) irradiation, and by transferring and diluting cells with fresh medium (the dilution effect). Box-L-like sequences are known as important cis-elements of genes for enzymes involved in the phenylpropanoid biosynthetic pathway. Six sequences, box-L0 to box-L5, exist in the DcPAL1 gene promoter region. In this study, we isolated cDNA encoding the R2R3 type of MYB transcription factor, DcMYB1, using yeast one-hybrid screening with box-L1 or box-L5 as target elements. DcMYB1 bound to boxes-L0, L1, L3/4, and L5 sequences (ACC(A/T)(A/T)CC) in vitro, and in yeast cells and carrot protoplasts. Transient expression of DcMYB1 could up-regulate DcPAL1 promoter activity in carrot protoplasts. Results of the transient expression experiment for the deletion-mutated promoters of boxes-L0, L1, L3, and L5 suggest that these box-L-like sequences were required for the complete activation of the DcPAL1 promoter by DcMYB1. Expression of DcMYB1 transcripts was induced 0.5 h after elicitor treatment or UV-B irradiation, and 2 h after the dilution effect. Induction of DcPAL1 expression occurred 1 h after DcMYB1 expression in all stress treatments, and repression of DcMYB1 expression by RNA interference caused cessation of the up-regulation of DcPAL1 expression in the elicitor treatment or with UV-B irradiation. These results suggest that DcMYB1 is the main regulatory factor acting on box-L sequences in the DcPAL1 gene that respond to environmental cues.

Base Sequence↗
PubMed1990

A case of Fabry's disease in a patient with no alpha-galactosidase A activity caused by a single amino acid substitution of Pro-40 by Ser.

We analyzed a male patient with Fabry's disease who had no activity of the lysosomal hydrolase alpha-galactosidase A (alpha-GalA) and female members of his family. We cloned a cDNA that encoded the mutant alpha-GalA, determined its nucleotide sequence, and found two nucleotide differences between the mutant and the wild-type cDNAs. Although one difference was silent, the other difference, a C-to-T transition at nucleotide number 118, resulted in an amino acid substitution of Pro-40 by Ser. A transient expression assay demonstrated that this missense mutation was the cause of the deficiency of alpha-GalA activity in the patient. In vitro mutagenesis experiments demonstrated that Pro-40 is critical for the appearance of alpha-GalA activity.

Base Sequence↗