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PubMedDate not supplied

Two novel mutations in a Japanese patient with the late-infantile form of metachromatic leukodystrophy.

Two novel mutations in the arylsulfatase A (ASA) gene from a Japanese patient with the late-infantile form of metachromatic leukodystrophy (MLD) were identified. One mutation was a G to C transversion at nucleotide 608 of the ASA gene (designated 608C) located at the 3' end of exon 2, which resulted in an amino acid substitution of Gln 153 to His. Although the 608 mutation resulted in a change in the exon-intron boundary consensus sequence, analysis of cDNA from the patient did not reveal the presence of aberrant splicing. The second mutation, a G to T transversion at nucleotide 1572 in exon 5 (designated 1572T), resulted in an amino acid substitution of Gly 308 to Val. This could potentially result in a conformational change in ASA protein structure. The patient was heterozygous for these two new mutations which were not present in 18 Japanese MLD alleles examined. A transient expression study in COS-1 cells showed no residual activity in either mutation. These results indicate that the 608C and 1572T mutations are responsible for the occurrence of the late-infantile form of MLD.

Cerebroside-Sulfatase↗

PubMed2006

[Construction and expression of various human prion proteins mutants with modified N-glycosylation sites in mammalian cells].

To study the biological function of the N-glycosylation modification of prion proteins (PrP), various eukaryotic expression vectors for the mutants with N-glycosylation modification of human PrP had been constructed and expressed. With site-direct mutation technique, human PRNP gene was mutated and the obtained mutants were subcloned into eukaryotic expressing plasmid pcDNA3.1 and transiently expressed in Hela cervical adenocarcinoma cell. The expression products of the mutated PrP were identified with Western blotting assay and the PNGase digestion assay. Several mutants with specific glycosylation modification were identified from the expressed products by Western blot, including two mutants with one glycosylation site mutated and one without any mutation at glycosylation sites. The expressed products were digested with PNGase F. The wild type proteins and those with one of glycosylation sites mutated were digested, resulting in their molecular weights reduced, while the molecular weights of products with mutations at both glycosylation sites were not changed. The mutant of wild type human PRNP gene at N-glycosylation modification sites and six modified mutants with mono- or non-N-glycosylation had been obtained successfully in the study. Moreover, the modified PrP with mono- and non-N-glycosylation were able to be expressed transitantly in Hela cells, which could be a useful means for studying prions.

Escherichia coli↗

PubMed1995

The major delayed rectifier in both Drosophila neurons and muscle is encoded by Shab.

The delayed rectifier K+ current in Drosophila is similar to the classical delayed rectifier, originally described by Hodgkin and Huxley. Drosophila provides unique tools of mutant analysis to unambiguously determine the genetic identity of this native K+ current. We identified the Shab gene as the exclusive gene underlying delayed rectifier currents in both muscle and neurons. In muscles, a genetic mutation of Shab removes virtually all the whole cell delayed rectifier current (IK), while leaving unaltered the transient A-current encoded by the Shaker gene. In neurons, the Shab mutation also removes the bulk of IK, but leaves unaltered the transient A-current encoded by the Shal gene. Although most of the delayed rectifier current is the product of the Shab gene, the Shaw gene contributes a small "leak" current to most neurons and muscle cells. Thus, in contrast to the A-currents which are encoded by different genes in muscle and neuronal cell bodies (Shaker and Shal, respectively), the predominant IK in both muscle and neurons is encoded by the same gene (Shab). With the genetic identity of IK confirmed, all of the major K+ currents in embryonic Drosophila neurons and muscle are now known.

Animals↗

PubMed2006

Selection acts on DNA secondary structures to decrease transcriptional mutagenesis.

Single-stranded DNA is more subject to mutation than double stranded. During transcription, DNA is transiently single stranded and therefore subject to higher mutagenesis. However, if local intra-strand secondary structures are formed, some bases will be paired and therefore less sensitive to mutation than unpaired bases. Using complete genome sequences of Escherichia coli, we show that local intra-strand secondary structures can, as a consequence, be used to define an index of transcription-driven mutability. At gene level, we show that natural selection has favoured a reduced transcription-driven mutagenesis via the higher than expected frequency of occurrence of intra-strand secondary structures. Such selection is stronger in highly expressed genes and suggests a sequence-dependent way to control mutation rates and a novel form of selection affecting the evolution of synonymous mutations.

Buchnera↗

PubMedDate not supplied

Two new mutations in the glucose-6-phosphatase gene cause glycogen storage disease in Hungarian patients.

Glycogen storage disease type 1a (von Gierke disease, GSD-1A) is caused by the deficiency of microsomal glucose-6-phosphatase (G6Pase) activity which catalyzes the final common step of glycogenolysis and gluconeogenesis. The cloning of the G6Pase cDNA and characterization of the human G6Pase gene enabled the identification of the mutations causing GSD-1a. This, in turn, allows the development of non-invasive DNA-based diagnosis that provides reliable carrier testing and prenatal diagnosis. Here we report on two new mutations E110Q and D38V causing GSD-1a in two Hungarian patients. The analyses of these mutations by site-directed mutagenesis followed by transient expression assays demonstrated that E110Q retains 17% of G6Pase enzymatic activity while the D38V abolishes the enzymatic activity. The patient with the E110Q has G222R as his other mutation. G222R was also shown to preserve about 4% of the G6Pase enzymatic activity. Nevertheless, the patient presented with the classical severe symptomatology of the GSD-1a.

Child↗

PubMed2003

Two novel mutations, Q1053H and C1060R, located in the D3 domain of von Willebrand factor, are responsible for decreased FVIII-binding capacity.

In type 2N von Willebrand disease (VWD), von Willebrand factor (VWF) is characterized by a markedly decreased affinity for Factor VIII (FVIII), and the mutations responsible are essentially located in the D' domain of VWF. We report the identification, in seven unrelated French families, of two novel type 2N VWD mutations, Q1053H and C1060R (Gln290His and Cys297Arg in mature VWF sequence), in exon 24 of the VWF gene. These missense mutations have been identified in the heterozygous, homozygous or hemizygous states. Using site-directed mutagenesis and transient expression in COS-7 cells, we showed that both mutations, although located in the D3 domain of VWF, outside the tryptic fragment containing the FVIII domain, dramatically decrease the binding of VWF to FVIII. In contrast, the R924Q substitution, which was identified in a patient who was heterozygous for C1060R, was shown to be a polymorphism.

Animals↗

PubMed1992

Regulation of delayed-early gene transcription by dual TATA boxes.

The 39K Autographa californica nuclear polyhedrosis virus (AcMNPV) gene is highly expressed throughout the virus life cycle and is controlled by tandem promoters that exhibit features of early and late baculovirus promoters. Late transcripts initiate at a conserved TAAG motif, while early transcripts are heterogeneous and initiate near a conserved CAGT motif. To define the nucleotide sequences that regulate early transcription of the 39K gene, a series of mutations was generated by substitution of 10-bp stretches in the 39K promoter with a BglII linker. The effects of these mutations on transcription from the early promoter were determined by transient expression and primer extension assays in the presence of the viral trans-activator IE1 gene. Mutations in the region from -15 to -44 revealed that early 39K transcription was controlled by dual TATA boxes. These TATA boxes are separated by 10 bp, which partially accounts for the heterogeneity in early 39K transcripts. Transcripts initiating at the CAGT motif (proximal transcripts) were abolished by deletion of the proximal TATA box located at -29 relative to CAGT. Proximal transcripts were not affected by alterations in the distal TATA motif located at -39 relative to the CAGT. Similarly, transcripts initiating upstream of CAGT (distal transcripts) were eliminated by mutations in the distal TATA but were unaffected by substitutions in the proximal TATA box. Proximal transcripts were not detected with a plasmid containing mutations in the CAGT motif, although the distal transcripts were unaffected by CAGT mutations. When the sequences surrounding the initiation site for the distal transcripts were altered, the start site was shifted one nucleotide, but transcription was not quantitatively affected. These results suggest that early 39K transcription is controlled by two distinct TATA elements, one that is dependent on an initiator and one in which the site of initiation is determined by the TATA element alone. Mutations in an upstream region from -45 to -68 relative to the CAGT motif had a quantitative effect but did not alter the heterogeneous pattern of early transcripts, suggesting these sequences function as an upstream regulatory region. Analysis of late transcription indicated that the TAAG element was essential, while transcription was unaffected by other mutations.

Animals↗

PubMed1999

Identification of a discrete intermediate in the assembly/disassembly of physalis mottle tymovirus through mutational analysis.

Assembly intermediates of icosahedral viruses are usually transient and are difficult to identify. In the present investigation, site-specific and deletion mutants of the coat protein gene of physalis mottle tymovirus (PhMV) were used to delineate the role of specific amino acid residues in the assembly of the virus and to identify intermediates in this process. N-terminal 30, 34, 35 and 39 amino acid deletion and single C-terminal (N188) deletion mutant proteins of PhMV were expressed in Escherichia coli. Site-specific mutants H69A, C75A, W96A, D144N, D144N-T151A, K143E and N188A were also constructed and expressed. The mutant protein lacking 30 amino acid residues from the N terminus self-assembled to T=3 particles in vivo while deletions of 34, 35 and 39 amino acid residues resulted in the mutant proteins that were insoluble. Interestingly, the coat protein (pR PhCP) expressed using pRSET B vector with an additional 41 amino acid residues at the N terminus also assembled into T=3 particles that were more compact and had a smaller diameter. These results demonstrate that the amino-terminal segment is flexible and either the deletion or addition of amino acid residues at the N terminus does not affect T=3 capsid assembly. In contrast, the deletion of even a single residue from the C terminus (PhN188Delta1) resulted in capsids that were unstable. These capsids disassembled to a discrete intermediate with a sedimentation coefficent of 19.4 S. However, the replacement of C-terminal asparagine 188 by alanine led to the formation of stable capsids. The C75A and D144N mutant proteins also assembled into capsids that were as stable as the pR PhCP, suggesting that C75 and D144 are not crucial for the T=3 capsid assembly. pR PhW96A and pR PhD144N-T151A mutant proteins failed to form capsids and were present as heterogeneous aggregates. Interestingly, the pR PhK143E mutant protein behaved in a manner similar to the C-terminal deletion protein in forming unstable capsids. The intermediate with an s value of 19.4 S was the major assembly product of pR PhH69A mutant protein and could correspond to a 30mer. It is possible that the assembly or disassembly is arrested at a similar stage in pR PhN188Delta1, pR PhH69A and pR PhK143E mutant proteins.

Animals↗

PubMed2006

Effects of voltage-gated calcium channel subunit genes on calcium influx in cultured C. elegans mechanosensory neurons.

Voltage-gated calcium channels (VGCCs) serve as a critical link between electrical signaling and diverse cellular processes in neurons. We have exploited recent advances in genetically encoded calcium sensors and in culture techniques to investigate how the VGCC alpha1 subunit EGL-19 and alpha2/delta subunit UNC-36 affect the functional properties of C. elegans mechanosensory neurons. Using the protein-based optical indicator cameleon, we recorded calcium transients from cultured mechanosensory neurons in response to transient depolarization. We observed that in these cultured cells, calcium transients induced by extracellular potassium were significantly reduced by a reduction-of-function mutation in egl-19 and significantly reduced by L-type calcium channel inhibitors; thus, a main source of touch neuron calcium transients appeared to be influx of extracellular calcium through L-type channels. Transients did not depend directly on intracellular calcium stores, although a store-independent 2-APB and gadolinium-sensitive calcium flux was detected. The transients were also significantly reduced by mutations in unc-36, which encodes the main neuronal alpha2/delta subunit in C. elegans. Interestingly, while egl-19 mutations resulted in similar reductions in calcium influx at all stimulus strengths, unc-36 mutations preferentially affected responses to smaller depolarizations. These experiments suggest a central role for EGL-19 and UNC-36 in excitability and functional activity of the mechanosensory neurons.

Amino Acid Sequence↗

PubMed2005

[Japanese CADASIL case with limited dementia who had the Notch 3 R141C mutation].

Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) is a rare hereditary disease characterized by recurrent transient ischemic attacks (TIA) and strokes, and vascular dementia caused by point mutations of the Notch 3 gene. Here, we report a Japanese CADASIL case who displayed limited dementia and had the Notch 3 R141C mutation. The relationship between pathogenesis and the mutation site in Notch 3 is discussed based on the case presented here.

CADASIL↗

PubMed2005

Unraveling a role for dopamine in Huntington's disease: the dual role of reactive oxygen species and D2 receptor stimulation.

Huntington's disease (HD), an inherited neurodegenerative disorder, results from an abnormal polyglutamine extension in the N-terminal region of the huntingtin protein. This mutation leads to protein aggregation and neurotoxicity. Despite its widespread expression in the brain and body, mutated huntingtin causes selective degeneration of striatal projection neurons. In the present study, we investigate the role of dopamine (DA) in this preferential vulnerability. Using primary cultures of striatal neurons transiently expressing GFP-tagged-exon 1 of mutated huntingtin, we show that low doses of DA (100 microM) act synergistically with mutated huntingtin to activate the proapoptotic transcription factor c-Jun. Surprisingly, DA also increases aggregate formation of mutated huntingtin in all cellular compartments, including neurites, soma, and nuclei. DA-dependent potentiation of c-Jun activation was reversed by ascorbate, a reactive oxygen species (ROS) scavenger, and SP-600125, a selective inhibitor of the c-Jun N-terminal kinase (JNK) pathway. By contrast, DA effects on aggregate formation were reversed by a selective D2 receptor antagonist and reproduced by a D2 agonist. Similarly, striatal neurons from D2 knockout mice showed no effect of DA on aggregate formation. Blocking ROS production, JNK activation, or D2 receptor stimulation significantly reversed DA aggravation of mutated huntingtin-induced striatal death. The combined treatment with the ROS scavenger and D2 antagonist totally reversed DA's effects on mutated huntingtin-induced striatal death. Thus, the present results provide insights into the cellular mechanisms that govern striatal vulnerability in HD and strongly support a dual role of JNK activation and D2 receptor signaling in this process.

Analysis of Variance↗

PubMed1996

A point mutation in the HIV-1 Tat responsive element is associated with postintegration latency.

Study of the mechanism of HIV-1 postintegration latency in the ACH2 cell line demonstrates that these cells failed to increase HIV-1 production following treatment with exogenous Tat. Reasoning that the defect in ACH2 cells involves the Tat response, we analyzed the sequence of tat cDNA and Tat responsive element (TAR) from the virus integrated in ACH2. Tat cDNA sequence is closely related to that of HIV LAI, and the encoded protein is fully functional in terms of long terminal repeat (LTR) transactivation. Cloning of a region corresponding to the 5'-LTR from ACH2, however, identified a point mutation (C37 -> T) in TAR. This mutation impaired Tat responsiveness of the LTR in transient transfection assays, and the measured defect was complemented in cells that had been treated with tetradecanoyl phorbol acetate or tumor necrosis factor type alpha (TNF-alpha). A compensatory mutation in TAR (G28 -> A), designed to reestablish base pairing in the TAR hairpin, restored wild-type Tat responsiveness. When the (C37 -> T) mutation was introduced in an infectious clone of HIV-1, no viral production was measured in the absence of TNF-alpha, whereas full complementation was observed when the infection was conducted in the presence of TNF-alpha or when a compensatory mutation (G28 -> A) was introduced into TAR. These experiments identify a novel mutation associated with HIV-1 latency and suggest that alterations in the Tat-TAR axis can be a crucial determinant of the latent phenotype in infected individuals.

Base Sequence↗

PubMed2005

In vitro resistance to interferon of hepatitis B virus with precore mutation.

AIM: Chronic hepatitis B virus (HBV) infection is predominantly treated with interferon alpha (IFN-alpha), which results in an efficient reduction of the viral load only in 20-40% of treated patients. Mutations at HBV precore prevail in different clinical status of HBV infection. The roles of precore mutation in the progression of chronic hepatitis and interferon sensitivity are still unknown. The aim of this study was to explore if there was any relationship between HBV precore mutation and sensitivity to interferon in vitro. METHODS: HBV replication-competent recombinant constructs with different patterns of precore mutations were developed. Then the recombinants were transiently transfected into hepatoma cell line (Huh7) by calcium phosphate transfection method. With or without IFN, viral products in culture medium were collected and quantified 3 d after transfection. RESULTS: We obtained 4 recombinant constructs by orientation-cloning 1.2-fold-overlength HBV genome into pUC18 vector via the EcoRI and Hind III and PCR mediated site-directed mutagenesis method. All the recombinants contained mutations within precore region. Huh7 cells transfected with recombinants secreted HBsAg and HBV particles into the cell culture medium, indicating that all the recombinants were replication-competent. By comparing the amount of HBV DNA in the medium, we found that HBV DNA in medium reflecting HBV replication efficiency was different in different recombinants. Recombinants containing precore mutation had fewer HBV DNAs in culture medium than wild type. This result showed that recombinants containing precore mutation had lower replication efficiency than wild type. HBV DNA was decreased in pUC18-HBV1.2-WT recombinants after IFN was added while others with precore mutations were not, indicating that HBV harboring precore mutation was less sensitive to IFN in cell culture system. CONCLUSION: These data indicate that HBV harboring precore mutation may be resistant to IFN in vitro.

Antiviral Agents↗

PubMed2004

Core promoter structure in the oomycete Phytophthora infestans.

We have investigated the core promoter structure of the oomycete Phytophthora infestans. The transcriptional start sites (TSS) of three previously characterized P. infestans genes, Piexo1, Piexo3, and Piendo1, were determined by primer extension analyses. The TSS regions were homologous to a previously identified 16-nucleotide (nt) core sequence that overlaps the TSS in most oomycete genes. The core promoter regions of Piexo1 and Piendo1 were investigated by using a transient protoplast expression assay and the reporter gene beta-glucuronidase. Mutational analyses of the promoters of Piexo1 and Piendo1 showed that there is a putative core promoter element encompassing the TSS (-2 to +5) that has high sequence and functional homology to a known core promoter element present in other eukaryotes, the initiator element (Inr). Downstream and flanking the Inr is a highly conserved oomycete promoter region (+7 to +15), hereafter referred to as FPR (flanking promoter region), which is also important for promoter function. The importance of the 19-nt core promoter region (Inr and FPR) in Piexo1 and Piendo1 was further investigated through electrophoretic mobility shift assays (EMSA). The EMSA studies showed that (i) both core promoters were able to specifically bind a protein or protein complex in a P. infestans whole-cell protein extract and (ii) the same mutations that reduced binding of the EMSA complex also reduced beta-glucuronidase (GUS) levels in transient expression assays. The consistency of results obtained using two different assays (GUS transient assays [in vivo] and EMSA studies [in vitro]) supports a convergence of inference about the relative importance of specific nucleotides within the 19-nt core promoter region.

Base Sequence↗

PubMed2005

[Construction of eukaryotic expression vectors of two mutants of hypoxia-inducible factor-1 and their expressions in human microvascular endothelial cells].

OBJECTIVE: To construct eukaryotic expression vectors of two mutants of hypoxia-inducible factor-1 (HIF-1alpha) and study their expressions in human microvascular endothelial cells (HMVECs). METHODS: Site-directed mutagenesis was performed to induce the mutation of the codons for the residue Pro564 (ccc) in HIF-1alpha into gcc (Ala) in pcDNA3.1(+)-HIF-1alphato obtain single-site-mutated vector pcDNA3.1(+)-HIF-1alpha-564Ala, which was then subjected to a second site-directed mutagenesis to convert the codons for Asn803 into that of Ala (gct) to acquire double-site-mutated pcDNA3.1(+)-HIF-1alpha-564Ala-803Ala. After lipofectin-mediated transient transformation of HMVECs with the 3 recombinant plasmids including the two plasmids containing the mutations and the one without mutation, respectively, the expression levels of HIF-1alpha mRNA and protein were determined using RT-PCR, immunofluorescent staining and Western blotting. RESULTS: DNA sequence analysis demonstrated success of the two-step mutagenesis and the two plasmids of pcDNA3.1+-HIF-1alpha-564Ala and pcDNA3.1(+)-HIF-1alpha-564Ala- 803Ala were obtained, both of which could produce HIF-1alpha protein resistant to oxidation degradation in HMVECs as compared with the non-mutated one. CONCLUSION: The recombinant plasmids pcDNA3.1(+)-HIF-1alpha-564Ala and pcDNA3.1(+)-HIF-1alpha- 564Ala-803Ala have been successfully constructed with efficient expressions in HMVECs.

Cloning, Molecular↗

PubMed2003

Inhibiting expression of specific genes in mammalian cells with 5' end-mutated U1 small nuclear RNAs targeted to terminal exons of pre-mRNA.

Reducing or eliminating expression of a given gene is likely to require multiple methods to ensure coverage of all of the genes in a given mammalian cell. We and others [Furth, P. A., Choe, W. T., Rex, J. H., Byrne, J. C., and Baker, C. C. (1994) Mol. Cell. Biol. 14, 5278-5289] have previously shown that U1 small nuclear (sn) RNA, both natural or with 5' end mutations, can specifically inhibit reporter gene expression in mammalian cells. This inhibition occurs when the U1 snRNA 5' end base pairs near the polyadenylation signal of the reporter gene's pre-mRNA. This base pairing inhibits poly(A) tail addition, a key, nearly universal step in mRNA biosynthesis, resulting in degradation of the mRNA. Here we demonstrate that expression of endogenous mammalian genes can be efficiently inhibited by transiently or stably expressed 5' end-mutated U1 snRNA. Also, we determine the inhibitory mechanism and establish a set of rules to use this technique and to improve the efficiency of inhibition. Two U1 snRNAs base paired to a single pre-mRNA act synergistically, resulting in up to 700-fold inhibition of the expression of specific reporter genes and 25-fold inhibition of endogenous genes. Surprisingly, distance from the U1 snRNA binding site to the poly(A) signal is not critical for inhibition, instead the U1 snRNA must be targeted to the terminal exon of the pre-mRNA. This could reflect a disruption by the 5' end-mutated U1 snRNA of the definition of the terminal exon as described by the exon definition model.

3' Untranslated Regions↗

PubMed2004

Genome-wide phenotype analysis in ES cells by regulated disruption of Bloom's syndrome gene.

The chief limitation of phenotype-based genetic screening in mammalian systems is the diploid nature of the genome. Cells deficient in the Bloom's syndrome gene (Blm) show an increased rate of loss of heterozygosity. Here we have used a tetracycline-regulated Blm allele (Blm(tet)) to introduce bi-allelic mutations across the genome in mouse embryonic stem (ES) cells. Transient loss of Blm expression induces homologous recombination not only between sister chromatids but also between homologous chromosomes. We considered that the phenotype of ES cells bearing bi-allelic mutations would be maintained after withdrawal of the tetracycline analogue doxycycline. Indeed, a combination of N-ethyl-N-nitrosourea mutagenesis and transient loss of Blm expression enabled us to generate an ES cell library with genome-wide bi-allelic mutations. The library was evaluated by screening for mutants of glycosylphosphatidylinositol-anchor biosynthesis, which involves at least 23 genes distributed throughout the genome. Mutants derived from 12 different genes were obtained and two unknown mutants were simultaneously isolated. Our results indicate that phenotype-based genetic screening with Blm(tet) is very efficient and raises possibilities for identifying gene functions in ES cells.

Adenosine Triphosphatases↗

PubMed1998

Transactivation of the human apolipoprotein CII promoter by orphan and ligand-dependent nuclear receptors. The regulatory element CIIC is a thyroid hormone response element.

The regulatory elements CIIC (-159/-116) and CIIB (-102/-81) of the apolipoprotein CII (apoCII) promoter have distinct specificities for orphan nuclear receptors (Vorgia, P., Zannis, V. I., and Kardassis, D. (1998) J. Biol. Chem. 273, 4188-4199). In this communication we investigated the contribution of ligand-dependent and orphan nuclear receptors on the transcriptional regulation of the human apoCII gene. It was found that element CIIC in addition to ARP-1 and EAR-2 binds RXRalpha/T3Rbeta heterodimers strongly, whereas element CIIB binds hepatic nuclear factor 4 (HNF-4) exclusively. Binding is abolished by mutations that alter the HRE binding motifs. Transient cotransfection experiments showed that in the presence of T3, RXRalpha/T3Rbeta heterodimers transactivated the -205/+18 apoCII promoter 1.6- and 11-fold in HepG2 and COS-1 respectively. No transactivation was observed in the presence of 9-cis-retinoic acid. Transactivation requires the regulatory element CIIC, suggesting that this element contains a thyroid hormone response element. HNF-4 did not affect the apoCII promoter activity in HepG2 cells. However, mutations in the HNF-4 binding site on element CIIB and inhibition of HNF-4 synthesis in HepG2 cells by antisense HNF-4 constructs decreased the apoCII promoter activity to 25-40% of the control, indicating that HNF-4 is a positive regulator of the apoCII gene. ARP-1 repressed the -205/+18 but not the -104/+18 apoCII promoter activity in HepG2 cells, indicating that the repression depends on the regulatory element CIIC. In contrast, combination of ARP-1 and HNF-4 transactivated different apoCII promoter segments as well as a minimal adenovirus major late promoter driven by the regulatory element CIIB. Mutagenesis or deletion of elements CIIB or CIIC established that the observed transactivation requires DNA binding of one of the two factors and may result from HNF-4-ARP-1 interactions that elicit the transactivation functions of HNF-4. The combined data indicate that RXRalpha/T3Rbeta in the presence of T3 and HNF-4 can upregulate the apoCII promoter activity by binding to the regulatory elements CIIC and CIIB, respectively. In addition, ARP-1 can either have inhibitory or stimulatory effects on the apoCII promoter activity via different mechanisms.

Animals↗