Biomedical subjects

L Li

Publications and source records attributed to L Li.

At least 793 records · Page 44Linked to original sources

PubMed1999

Coniferyl aldehyde 5-hydroxylation and methylation direct syringyl lignin biosynthesis in angiosperms.

A central question in lignin biosynthesis is how guaiacyl intermediates are hydroxylated and methylated to the syringyl monolignol in angiosperms. To address this question, we cloned cDNAs encoding a cytochrome P450 monooxygenase (LsM88) and a caffeate O-methyltransferase (COMT) from sweetgum (Liquidambar styraciflua) xylem. Mass spectrometry-based functional analysis of LsM88 in yeast identified it as coniferyl aldehyde 5-hydroxylase (CAld5H). COMT expressed in Escherichia coli methylated 5-hydroxyconiferyl aldehyde to sinapyl aldehyde. Together, CAld5H and COMT converted coniferyl aldehyde to sinapyl aldehyde, suggesting a CAld5H/COMT-mediated pathway from guaiacyl to syringyl monolignol biosynthesis via coniferyl aldehyde that contrasts with the generally accepted route to sinapate via ferulate. Although the CAld5H/COMT enzyme system can mediate the biosynthesis of syringyl monolignol intermediates through either route, k(cat)/K(m) of CAld5H for coniferyl aldehyde was approximately 140 times greater than that for ferulate. More significantly, when coniferyl aldehyde and ferulate were present together, coniferyl aldehyde was a noncompetitive inhibitor (K(i) = 0.59 microM) of ferulate 5-hydroxylation, thereby eliminating the entire reaction sequence from ferulate to sinapate. In contrast, ferulate had no effect on coniferyl aldehyde 5-hydroxylation. 5-Hydroxylation also could not be detected for feruloyl-CoA or coniferyl alcohol. Therefore, in the presence of coniferyl aldehyde, ferulate 5-hydroxylation does not occur, and the syringyl monolignol can be synthesized only from coniferyl aldehyde. Endogenous coniferyl, 5-hydroxyconiferyl, and sinapyl aldehydes were detected, consistent with in vivo operation of the CAld5H/COMT pathway from coniferyl to sinapyl aldehydes via 5-hydroxyconiferyl aldehyde for syringyl monolignol biosynthesis.

Cloning, Molecular↗

PubMed1999

Axin and Frat1 interact with dvl and GSK, bridging Dvl to GSK in Wnt-mediated regulation of LEF-1.

Wnt proteins transduce their signals through dishevelled (Dvl) proteins to inhibit glycogen synthase kinase 3beta (GSK), leading to the accumulation of cytosolic beta-catenin and activation of TCF/LEF-1 transcription factors. To understand the mechanism by which Dvl acts through GSK to regulate LEF-1, we investigated the roles of Axin and Frat1 in Wnt-mediated activation of LEF-1 in mammalian cells. We found that Dvl interacts with Axin and with Frat1, both of which interact with GSK. Similarly, the Frat1 homolog GBP binds Xenopus Dishevelled in an interaction that requires GSK. We also found that Dvl, Axin and GSK can form a ternary complex bridged by Axin, and that Frat1 can be recruited into this complex probably by Dvl. The observation that the Dvl-binding domain of either Frat1 or Axin was able to inhibit Wnt-1-induced LEF-1 activation suggests that the interactions between Dvl and Axin and between Dvl and Frat may be important for this signaling pathway. Furthermore, Wnt-1 appeared to promote the disintegration of the Frat1-Dvl-GSK-Axin complex, resulting in the dissociation of GSK from Axin. Thus, formation of the quaternary complex may be an important step in Wnt signaling, by which Dvl recruits Frat1, leading to Frat1-mediated dissociation of GSK from Axin.

Adaptor Proteins, Signal Transducing↗

PubMed1999

Effects of bilateral electrical stimulation of the ventral pallidum on acoustic startle.

The ventral pallidum (VP) is believed to occupy a critical position between the limbic and the motor systems, for transferring motive information into motor commands. To estimate the time course of signaling from the VP to motor outputs, in the present study we examined the effects of bilateral electrical stimulation of the VP on the acoustic startle reflex in awake rats. When the interstimulus interval (ISI) between VP stimulation and acoustic stimulation was shorter than 5 ms, VP stimulation potentiated acoustic startle. When the ISI was longer than 5 ms, VP stimulation inhibited acoustic startle over a large range of ISIs with the maximum inhibition at ISIs between 15 and 25 ms. In contrast, bilateral electrical stimulation of the amygdala did not have a significant inhibitory effect on acoustic startle, but strongly augmented acoustic startle at shorter ISIs (0-10 ms). Compared to unilateral electrical stimulation of the inferior colliculus (IC), bilateral stimulation of the VP gave rise to a rightward shift of the ISI curve, indicating that the neural pathways conveying the inhibitory influence from the VP to the acoustic startle circuit are longer than those from the IC.

Acoustic Stimulation↗

PubMed1999

Human tryptases alpha and beta/II are functionally distinct due, in part, to a single amino acid difference in one of the surface loops that forms the substrate-binding cleft.

Tryptases alpha and beta/II were expressed in insect cells to try to ascertain why human mast cells express these two nearly identical granule proteases. In contrast to that proposed by others, residue -3 in the propeptide did not appear to be essential for the three-dimensional folding, post-translational modification, and/or activation of this family of serine proteases. Both recombinant tryptases were functional and bound the active-site inhibitor diisopropyl fluorophosphate. However, they differed in their ability to cleave varied trypsin-susceptible chromogenic substrates. Structural modeling analyses revealed that tryptase alpha differs from tryptase beta/II in that it possesses an Asp, rather than a Gly, in one of the loops that form its substrate-binding cleft. A site-directed mutagenesis approach was therefore carried out to determine the importance of this residue. Because the D215G derivative of tryptase alpha exhibited potent enzymatic activity against fibrinogen and other tryptase beta/II-susceptible substrates, Asp215 dominantly restricts the substrate specificity of tryptase alpha. These data indicate for the first time that tryptases alpha and beta/II are functionally different human proteases. Moreover, the variation of just a single amino acid in the substrate-binding cleft of a tryptase can have profound consequences in the regulation of its enzymatic activity and/or substrate preference.

Amino Acid Sequence↗

PubMed1999

Gene therapy for canine alpha-L-iduronidase deficiency: in utero adoptive transfer of genetically corrected hematopoietic progenitors results in engraftment but not amelioration of disease.

Canine alpha-L-iduronidase (iduronidase) deficiency is a model of the human lysosomal storage disorder mucopolysaccharidosis type I (MPS I). We used this canine model to evaluate the therapeutic potential of hematopoietic stem cell (HSC) gene therapy for enzyme deficiencies. In previous studies, iduronidase-deficient dogs infused with autologous marrow cells genetically modified to express iduronidase had long-term engraftment with provirally marked cells, but there was no evidence of proviral iduronidase expression or clinical improvement. The presence of humoral and cellular immune responses against iduronidase apparently abrogated the therapeutic potential of HSC gene therapy in these experiments. To evaluate HSC gene therapy for canine MPS I in the absence of a confounding immune response, we have now performed in utero adoptive transfer of iduronidase-transduced MPS I marrow cells into preimmune fetal pups. In three separate experiments, 17 midgestation fetal pups were injected with 0.5-1.5 x 10(7) normal or MPS I allogeneic long-term marrow culture (LTMC) cells transduced with neo(r)- or iduronidase-containing retroviral vectors. Nine normal and three MPS I pups survived the neonatal period and demonstrated engraftment of provirally marked progenitors at levels of up to 12% for up to 12 months. However, the proportion of provirally marked circulating leukocytes was approximately 1%. Neither iduronidase enzyme nor proviral-specific transcripts were detected in blood or marrow leukocytes of any MPS I dog. Humoral immune responses to iduronidase were not detected in neonates, even after "boosting" with autologous iduronidase-transduced LTMC cells. All MPS I dogs died at 8-11 months of age from complications of MPS I disease with no evidence of amelioration of MPS I disease. Our results suggest that iduronidase-transduced primitive hematopoietic progenitors can engraft in fetal recipients, contribute to hematopoiesis, and induce immunologic nonresponsiveness to iduronidase in MPS I dogs. However, the therapeutic potential of HSC gene transfer in this model of iduronidase deficiency appears to be limited by poor maintenance of proviral iduronidase gene expression and relatively low levels of genetically corrected circulating leukocytes.

Adoptive Transfer↗

PubMed1999

Prostaglandin E2 enhancement of interferon-gamma production by antigen-stimulated type 1 helper T cells.

Prostaglandin E2 (PGE2) is a potent mediator generated in immune tissues by cyclooxygenation of arachidonic acid. PGE2 affects T cell functions through four homologous G protein-coupled receptors termed EP1R, EP2R, EP3R, and EP4R that differ in tissue distribution and signaling. Antigen-evoked secretion of interferon-gamma (IFN-gamma) by sperm whale myoglobin-specific Th1 cells of DBA/2 mouse I-Ed-restricted clones, that express EP3Rs and EP4Rs, was enhanced a maximum of 3-fold by 10(-10) to 10(-8) M PGE2 and 2.5-fold each for the EP1R/EP3R-directed agonist sulprostone (10(-8) and 10(-7) M) and for the EP4R/EP3R/EP2R agonist misoprostol (10(-9) M). Neither PGE2 nor the synthetic analogs affected secretion of IFN-gamma by PMA plus ionomycin-stimulated clones of Th1 cells. Antigen-evoked secretion of IFN-gamma by influenza hemagglutinin-specific mouse lymph node Th1 cells, that also express EP3Rs and EP4Rs, was increased a maximum of 12-fold by 10(-9) to 10(-8) M PGE2, 14-fold by 10(-9) M sulprostone, and 10-fold by 10(-9) M misoprostol. Production of IFN-gamma by either type of Th1 cell was not affected significantly by 10(-6) M PGE2 alone. The generation of IFN-gamma by antigen-stimulated Th1 cells thus is significantly enhanced by physiologically relevant concentrations of PGE2.

Animals↗

PubMed1999

Decay accelerating factor (CD55): a target for cancer vaccines?

The 791Tgp72 antigen has been used successfully as a target for tumor imaging and T-cell immunotherapy. We have characterized this antigen using the monoclonal antibody 791T/36 as a 72/66 kDa doublet. NH2-terminal protein sequencing of the two bands revealed identity with the complement regulatory protein CD55. Antibodies recognizing different domains of CD55 were also shown to bind to the purified 791Tgp72, although sequence analysis of the cDNA cloned from 791T tumor cells showed 100% homology with CD55 and transfection of the cDNA into antigen-negative CHO cells resulted in binding of 791T/36. This identifies the tumor antigen 791Tgp72 as CD55. This protein protects cells from complement attack; however, it can also transduce signals in lymphocytes and is a ligand for CD97, expressed by activated T cells. These results suggest that CD55 plays a role in signaling between the innate and adaptive immune responses. It is therefore a very intriguing target, because absence of the molecule makes the tumor cells susceptible to complement, whereas protective overexpression results in the antigen being a target for T-cell immunotherapy.

Amino Acid Sequence↗

PubMed1999

Coupling of M2 muscarinic receptors to membrane ion channels via phosphoinositide 3-kinase gamma and atypical protein kinase C.

We report a novel signaling pathway linking M2 muscarinic receptors to metabotropic ion channels. Stimulation of heterologously expressed M2 receptors, but not other Gi/Go-associated receptors (M4 or alpha2c), activates a calcium- and voltage-independent chloride current in Xenopus oocytes. We show that the stimulatory pathway linking M2 receptors to these chloride channels consists of Gbeta gamma stimulation of phosphoinositide 3-kinase gamma (PI-3Kgamma), formation of phosphatidylinositol 3,4,5-trisphosphate (PIP3), and activation of atypical protein kinase C (PKC). The chloride current is activated in the absence of M2 receptor stimulation by the injection of PIP3, and PIP3 current activation is blocked by a pseudosubstrate inhibitory peptide of atypical PKC but not other PKCs. Moreover, the current is activated by injection of recombinant PKCzeta at concentrations as low as 1 nM. M2 receptor-current coupling was disrupted by inhibiton of PI-3K and by injection of beta gamma binding peptides, but it was not affected by expression of dominant negative p85 cRNA. We also show that this pathway mediates M2 receptor coupling to metabotropic nonselective cation channels in mammalian smooth muscle cells, thus demonstrating the broad relevance of this signaling cascade in neurotransmitter signaling.

Animals↗

PubMed1999

Effects of gel material on fluorescence lifetime detection of dyes and dye-labeled DNA primers in capillary electrophoresis.

Investigations of fluorescence lifetimes of the dye 6-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino]hexanoic acid (NBD-HA) and of DNA M13 primers labeled with NBD-HA, Cy3, rhodamine green and 4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacene++ +-3-propionic acid (BODIPY-FL) dyes in polyacrylamide gels of various degrees of crosslinking and different crosslinkers, and in a cellulose sieving buffer with different organic modifiers, are described. The dependence of fluorescence lifetime on gel matrix and on experimental conditions was studied in order to identify which factors may be important for optimization of multiplex fluorescence lifetime detection. Lifetimes were determined in both batch solution and on-the-fly, on-column in CE. Results show that lifetimes of the primer-attached dyes remain constant in gels of different composition. Additionally, multiexponential fluorescence decays are observed for primer-attached dyes in batch solutions of the cellulose sieving buffers but are reduced to monoexponential decays when measured on-the-fly, on-column in CE. Lifetime detectability can be improved by addition of an organic modifier to the gel matrix.

Acrylic Resins↗

PubMed1999

Emergence of regulatory CD4+ T cell response to repetitive stimulation with antigen-presenting cells in vitro: implications in designing antigen-presenting cell-based tumor vaccines.

Because APCs play a crucial role in the generation of T cell-mediated immune responses, numerous clinical trials with APC-based vaccines have been initiated in different types of human cancers. Encouraging results have emerged from some of these initial studies. Thus far, APC-based vaccinations usually include multiple rounds of immunization. With this approach, although we and others have detected induction of Ag-specific CTL responses in vaccinated patients after stimulation with the same APC-based immunogen, in vitro we also find that repetitive in vitro stimulation with Ag-loaded APC can, at times, lead to the emergence of noncytolytic CD4+ T cells exhibiting the characteristic phenotype of Th2 cells. These noncytolytic CD4+ T cells synthesize large quantities of type 2 cytokines such as IL-4 and IL-10 on stimulation with the autologous APC or tumor cells in an MHC class II-restricted manner. Further, these CD4+ T cells and a cell-free supernatant factor block the activation of fresh T lymphocytes. The supernatant factor also exhibits a marked inhibitory effect on the expression of the costimulatory molecules, CD80 and CD86, by APC. The inhibitory effect of the supernatant factor can be abrogated by neutralizing IL-10 in the supernatant. These observations therefore have implications in the APC-based tumor vaccine protocol design.

Antigen-Presenting Cells↗

PubMed1999

Key role of the diglucosyldiacylglycerol synthase for the nonbilayer-bilayer lipid balance of Acholeplasma laidlawii membranes.

In the single membrane of Acholeplasma laidlawii, a specific glucosyltransferase (DGlcDAG synthase) synthesizes the major, bilayer-forming lipid diglucosyldiacylglycerol (DGlcDAG) from the preceding major, nonbilayer-prone monoglucosyldiacylglycerol (MGlcDAG). This is crucial for the maintenance of phase equilibria close to a potential bilayer-nonbilayer transition and a nearly constant spontaneous curvature for the membrane bilayer lipid mixture. The glucolipid pathway is also balanced against the phosphatidylglycerol (PG) pathway to maintain a certain lipid surface charge density. The DGlcDAG synthase was purified approximately 5000-fold by three chromatographic techniques and identified as a minor 40 kDa membrane protein. In CHAPS mixed micelles, a cooperative dependence on anionic lipid activators was confirmed, with PG as the best. The dependence of the enzyme on the soluble UDP-glucose substrate followed Michaelis-Menten kinetics, while the kinetics for the other (lipid) substrate MGlcDAG exhibited cooperativity, with Hill coefficients in the range of 3-5. Vmax and the Hill coefficient, but not Km, for the MGlcDAG substrate were increased by increased PG concentrations, but above 3 mol % MGlcDAG, the rate of synthesis was constant. Hence, the DGlcDAG synthase is more affected by the lipid activator than by the lipid substrate at physiological lipid concentrations. The enzyme was shown to be sensitive to curvature "stress" changes, i.e., was stimulated by various nonbilayer lipids but inhibited by certain others. Certain phosphates were also stimulatory. With the two purified MGlcDAG and DGlcDAG synthases reconstituted together in the presence of a potent nonbilayer lipid, the strong responses in the amounts of MGlcDAG and DGlcDAG synthesized mimicked the responses in vivo. This supports the important regulatory functions of these enzymes.

Acholeplasma laidlawii↗

PubMed1999

Neuron-specific and developmental regulation of the synapsin II gene expression in transgenic mice.

Synapsin II, a major phosphoprotein of synaptic vesicles, is believed to function in neurotransmitter release as well as in synapse formation. The expression of the synapsin II gene is neuron-specific, and correlates temporally with synaptogenesis. To understand the mechanisms by which the expression of the synapsin II gene is regulated in vivo, we generated transgenic mice carrying a 5.1-kb 5'-flanking sequence of the murine synapsin II gene fused to the firefly luciferase reporter gene. The synapsin II-luciferase transgene is specifically expressed in neural tissues, such as brain and spinal cord, but not in non-neural tissues. Throughout the brain, the expression of the transgene is widely distributed, and restricted only to neuronal cells. Moreover, the expression of the transgene is developmentally regulated, with a temporal profile similar to that of endogenous synapsin II expression. These results indicate that the 5.1-kb flanking sequence of the murine synapsin II gene contains cis-regulatory elements that are required for directing neuron-specific and synaptogenesis-regulated expression in vivo.

Age Factors↗

PubMed1999

Regulation of maternal behavior and offspring growth by paternally expressed Peg3.

Imprinted genes display parent-of-origin-dependent monoallelic expression that apparently regulates complex mammalian traits, including growth and behavior. The Peg3 gene is expressed in embryos and the adult brain from the paternal allele only. A mutation in the Peg3 gene resulted in growth retardation, as well as a striking impairment of maternal behavior that frequently resulted in death of the offspring. This result may be partly due to defective neuronal connectivity, as well as reduced oxytocin neurons in the hypothalamus, because mutant mothers were deficient in milk ejection. This study provides further insights on the evolution of epigenetic regulation of imprinted gene dosage in modulating mammalian growth and behavior.

Animals↗

PubMed1999

Differential susceptibility of brain areas to cyanide involves different modes of cell death.

We have demonstrated that cyanide (KCN) induces selective degeneration of dopaminergic neurons in mice and apoptotic cell death in cultured neurons. In the present study the mode of cyanide-induced cell death was determined in the susceptible brain areas. Mice were treated with KCN (6 mg/kg ip) or vehicle (saline) twice daily for 1 to 12 days. After 3 days of KCN treatment, two separate lesions were observed in coronal brain sections. Widespread DNA fragmentation in parietal and suprarhinal regions of the motor cortex was observed by the in situ terminal deoxynucleotide transferase nick-end labeling (TUNEL) technique. Pyknosis and chromatin condensation, morphological hallmarks of apoptotic cells, were observed in TUNEL-positive regions. On the other hand, in the substantia nigra (SN), KCN produced a progressive, bilateral necrotic lesion that was evident by 3 days of treatment. The SN lesion was circumscribed by a prominent ring of glial infiltration, as determined by glial-acidic fibrillary protein (GFAP) immunostaining. The extent of the SN lesion steadily increased with treatment duration, and DNA fragmentation was not observed over the 1- to 12-day period. On the other hand, cortical apoptosis was not associated with necrotic cell loss or astrogliosis. Pretreatment of animals with the antioxidant alpha-phenyl-tert-butyl nitrone (PBN) for 7 days prior to and during 3 days of KCN administration markedly reduced cortical DNA fragmentation whereas the PBN treatment did not influence the SN necrosis or astrocytic gliosis. Except for moderate GFAP immunostaining in corpus callosum, other brain areas were not affected by cyanide. It is concluded that KCN-induced neuronal loss involves selective activation of necrosis or apoptosis in different neuronal populations, and involves divergent mechanisms and sensitivity to antioxidants.

Animals↗

PubMed1999

Intestinal absorption of dietary cholesteryl ester is decreased but retinyl ester absorption is normal in carboxyl ester lipase knockout mice.

Carboxyl ester lipase (CEL; EC 3.1.1.13) hydrolyzes cholesteryl esters and retinyl esters in vitro. In vivo, pancreatic CEL is thought to liberate cholesterol and retinol from their esters prior to absorption in the intestine. CEL is also a major lipase in the breast milk of many mammals, including humans and mice, and is thought to participate in the processing of triglycerides to provide energy for growth and development while the pancreas of the neonate matures. Other suggested roles for CEL include the direct facilitation of the intestinal absorption of free cholesterol and the modification of plasma lipoproteins. Mice with different CEL genotypes [wild type (WT), knockout (CELKO), heterozygote] were generated to study the functions of CEL in a physiological system. Mice grew and developed normally, independent of the CEL genotype of the pup or nursing mother. Consistent with this was the normal absorption of triglyceride in CELKO mice. The absorption of free cholesterol was also not significantly different between CELKO (87 +/- 26%, mean +/- SD) and WT littermates (76 +/- 10%). Compared to WT mice, however, CELKO mice absorbed only about 50% of the cholesterol provided as cholesteryl ester (CE). There was no evidence for the direct intestinal uptake of CE or for intestinal bacterial enzymes that hydrolyze it, suggesting that another enzyme besides CEL can hydrolyze dietary CE in mice. Surprisingly, CELKO and WT mice absorbed similar amounts of retinol provided as retinyl ester (RE). RE hydrolysis, however, was required for absorption, implying that CEL was not the responsible enzyme. The changes in plasma lipid and lipoprotein levels to diets with increasing lipid content were similar in mice of all three CEL genotypes. Overall, the data indicate that in the mouse, other enzymes besides CEL participate in the hydrolysis of dietary cholesteryl esters, retinyl esters, and triglycerides.

Animals↗

PubMed1999

Carboxyl ester lipase overexpression in rat hepatoma cells and CEL deficiency in mice have no impact on hepatic uptake or metabolism of chylomicron-retinyl ester.

To study the role of carboxyl ester lipase (CEL) in hepatic retinoid (vitamin A) metabolism, we investigated uptake and hydrolysis of chylomicron (CM)-retinyl esters (RE) by rat hepatoma (McArdle-RH7777) cells stably transfected with a rat CEL cDNA. We also studied tissue uptake of CM-RE in CEL-deficient mice generated by targeted disruption of the CEL gene. CEL-transfected cells secreted active enzyme into the medium. However, both control and CEL-transfected cells accumulated exogenously added CM-RE or CM remnant (CMR)-derived RE in equal amounts. Serum clearance of intravenously injected CM-RE and cholesteryl ester were not different between wild-type and CEL-deficient mice. Also, the uptake of the two compounds by the liver and other tissues did not differ. These data indicate that the lack of CEL expression does not affect the uptake of dietary CM-RE by the liver or other tissues. Moreover, the percentage of retinol formed in the liver after CM-RE uptake, the levels of retinol and retinol-binding protein in serum, and retinoid levels in various tissues did not differ, indicating that CEL deficiency does not affect hepatic retinoid metabolism and retinoid distribution throughout the body. Surprisingly, in both pancreas and liver of wild-type, heterozygous, and homozygous CEL-deficient mice, the levels of bile salt-dependent retinyl ester hydrolase (REH) activity were similar. This indicates that in the mouse pancreas and liver an REH enzyme activity, active in the presence of bile salt and distinct from CEL, is present, compatible with the results from our accompanying paper that the intestinal processing and absorption of RE were unimpaired in CEL-deficient mice.

Animals↗

PubMed1999

SMRTe, a silencing mediator for retinoid and thyroid hormone receptors-extended isoform that is more related to the nuclear receptor corepressor.

SMRT (silencing mediator for retinoid and thyroid hormone receptors) and N-CoR (nuclear receptor copressor) mediate transcriptional repression of important regulators that are involved in many signaling pathways. SMRT and N-CoR are related proteins that form complexes with mSin3A/B and histone deacetylases to induce local chromatin condensation and transcriptional repression. However, SMRT is substantially smaller than N-CoR, lacking an N-terminal domain of approximately 1,000 aa that are present in N-CoR. Here, we report the identification of SMRT-extended (SMRTe), which contains an N-terminal sequence that shows striking similarity with N-CoR. As in N-CoR, this SMRTe-N-terminal domain also represses basal transcription. We find that SMRTe expression is regulated during cell cycle progression and SMRTe transcripts are present in many embryonic tissues. These data redefine a structurally and functionally more related nuclear receptor corepressor family and suggest an additional role for SMRTe in the regulation of cycle-specific gene expression in diverse signaling pathways.

Amino Acid Sequence↗

PubMed1999

Identification of the block in targeted retroviral-mediated gene transfer.

A chimeric retroviral vector (33E67) containing a CD33-specific single-chain antibody was generated in an attempt to target cells displaying the CD33 surface antigen. The chimeric envelope protein was translated, processed, and incorporated into viral particles as efficiently as wild-type envelope protein. The viral particles carrying the 33E67 envelope protein could bind efficiently to the CD33 receptor on target cells and were internalized, but no gene transfer occurred. A unique experimental approach was used to examine the basis for this postbinding block. Our data indicate that the chimeric envelope protein itself cannot participate in the fusion process, the most reasonable explanation being that this chimeric protein cannot undergo the appropriate conformational change that is thought to be triggered by receptor binding, a suggested prerequisite to subsequent fusion and core entry. These results indicate that the block to gene transfer in this system, and probably in most of the current chimeric retroviral vectors to date, is the inability of the chimeric envelope protein to undergo this obligatory conformational change.

3T3 Cells↗