Biomedical subjects

J W Dennis

Publications and source records attributed to J W Dennis.

At least 19 recordsLinked to original sources

PubMed2005

Regulation of BMP-induced ectopic bone formation by Ahsg.

alpha2-HS-glycoprotein (Ahsg), also known as fetuin is a serum and bone resident glycoprotein, which binds to TGF-beta superfamily members including bone morphogenetic proteins (BMP) and inhibits dexamethasone-induced osteogenesis in bone marrow cultures in vitro. Here we demonstrate that Ahsg reduces cytokine binding to its cognate receptor in HOS osteocyte cells and suppresses intracellular signaling, while in vivo, we test the hypothesis that Ahsg-deficient mice are hyper-responsive to BMP-induced osteogenesis. Human native BMP was implanted into the hindquarter muscles of Ahsg(+/+), Ahsg(+/-) and Ahsg(-/-) mice and 4 weeks later, ossicle formation was analyzed by radiography, bone density scanning (DEXA) and histomorphometry. Alkaline phosphatase (AP) activity was measured in ossicles as a marker for bone cell differentiation, and was significantly higher in Ahsg(-/-) versus Ahsg(+/-) and/or Ahsg(+/+) mice. Ectopic ossicle size in the Ahsg(+/-) mouse was 4-fold greater than that in the wild type (Ahsg(+/+)), and intermediate to that shown in Ahsg(-/-) mouse. Bone mineral density (BMD) was lower in the Ahsg(-/+) and Ahsg(-/-) mice compared to Ahsg(+/+) littermates. The ratio of cortical to cancellous bone was found to be >2-fold higher in Ahsg(-/-) mouse in comparison to the Ahsg(+/+) mice with no significant change in the Ahsg(-/+) mouse. Finally, a significantly higher incidence of satellite ossification; small islands of immature bone, was shown in Ahsg(-/-) mice as compared to Ahsg(+/+) mice. Although Ahsg binds to TGF-beta/BMP and blocks receptor signalling, it may also sequester cytokines in matrix, thereby acting as a reservoir of osteoinductive activity when released. This may explain the non-linear relationship between ectopic bone formation characteristics and Ahsg(+/+), Ahsg(+/-) and Ahsg(-/-) genotypes, although the increase in satellite bone formation might also explain this phenomenon. Our results suggest that Ahsg may be useful for prevention of the heterotopic ossification and the regulation of osteoinductive effects of BMP used with grafts.

Alkaline Phosphatase↗

PubMed2001

Quality control of resident operative experience: compliance with RRC criteria.

PURPOSE: Process Capability Analysis (PCA) is a quality control tool that can be applied to how resident cases are allocated. PCA measures how close an output is to its target (Cp) and its location (K) in relation to Cp. For resident cases, the statistics of Cp and K measure variability in case numbers, which is useful in planning how a program meets RRC criteria in operative experience on specialty services. METHODS: A review of 6 years of vascular surgery cases (1994-1999) using PG 5 RRC case logs and the departmental quality database as sources was done. PCA was applied to resident case numbers. RRC 1997-1998 National Program Data were used to define quality control limits. The 30th, 50th, and 70th percentiles in vascular procedures served as lower, nominal, and upper control limits, respectively. RESULTS: Cases were grouped into aortic (AO), cerebrovascular (CER), peripheral (PER), and visceral (VSC), to meet RRC definition for major reconstructions. PCA analysis of PG5 RRC submitted data, total cases reported by the graduating residents (n = 20) in their submitted RRC Case logs, and department QA log numbers over 6 years of the period revealed the following mean numbers for AO = 13.7 +/- 4.2 (Cp = 0.279, K = +70.5%), CER = 15.6 +/- 6.4 (Cp = 0.365, K = +78.6%), PER = 23.1 +/- 10.5 (Cp = 0.237, K = +80.1%), VSC = 2.5 +/- 1.5 (Cp = 0.189, K = +175.0%), and total = 55.1 +/- 20.2 (Cp = 0.305, K = +70.5%). CONCLUSIONS: Processes that are in statistical control should have a Cp > 1.0, and a small K: i.e., centered tightly and near the desired nominal limit. By PCA analysis, the vascular surgery service product of PG5 case numbers is not in statistical control, but instead exceeds the defined control limits (RRC 50th percentile levels). The low Cp values demonstrate that specific case numbers may vary from resident to resident. The high positive K percentages imply that more residents have case numbers that meet or exceed RRC 50th percentiles. Sources of potential error and variability may be partially explained by the data of total case volume and resident underreporting. More accurate RRC reporting by residents could account for up to 20% more cases, potentially offsetting individual case allocation differences within a particular resident year.

Aorta↗

PubMed2001

Late mitotic failure in mice lacking Sak, a polo-like kinase.

Polo-like kinases in yeast, flies, and mammals regulate key events in mitosis. Such events include spindle formation at G2/M, the anaphase-promoting complex (APC) at the exit from mitosis, the cleavage structure at cytokinesis, and DNA damage checkpoints in G2/M. Polo-like kinases are distinguished by two C-terminal polo box (pb) motifs, which localize the enzymes to mitotic structures. We previously identified Sak, a novel polo-like kinase found in Drosophila and mammals. Here, we demonstrate that the Sak kinase has a functional pb domain that localizes the enzyme to the nucleolus during G2, to the centrosomes in G2/M, and to the cleavage furrow during cytokinesis. To study the role of Sak in embryo development, we generated a Sak null allele, the first polo-like kinase to be mutated in mice. Sak(-/-) embryos arrested after gastrulation at E7.5, with a marked increase in mitotic and apoptotic cells. Sak(-/-) embryos displayed cells in late anaphase or telophase that continued to express cyclin B1 and phosphorylated histone H3. Our results suggest that Sak is required for the APC-dependent destruction of cyclin B1 and for exit from mitosis in the postgastrulation embryo.

3T3 Cells↗

PubMed2001

Negative regulation of T-cell activation and autoimmunity by Mgat5 N-glycosylation.

T-cell activation requires clustering of a threshold number of T-cell receptors (TCRs) at the site of antigen presentation, a number that is reduced by CD28 co-receptor recruitment of signalling proteins to TCRs. Here we demonstrate that a deficiency in beta1,6 N-acetylglucosaminyltransferase V (Mgat5), an enzyme in the N-glycosylation pathway, lowers T-cell activation thresholds by directly enhancing TCR clustering. Mgat5-deficient mice showed kidney autoimmune disease, enhanced delayed-type hypersensitivity, and increased susceptibility to experimental autoimmune encephalomyelitis. Recruitment of TCRs to agonist-coated beads, TCR signalling, actin microfilament re-organization, and agonist-induced proliferation were all enhanced in Mgat5-/- T cells. Mgat5 initiates GlcNAc beta1,6 branching on N-glycans, thereby increasing N-acetyllactosamine, the ligand for galectins, which are proteins known to modulate T-cell proliferation and apoptosis. Indeed, galectin-3 was associated with the TCR complex at the cell surface, an interaction dependent on Mgat5. Pre-treatment of wild-type T cells with lactose to compete for galectin binding produced a phenocopy of Mgat5-/- TCR clustering. These data indicate that a galectin-glycoprotein lattice strengthened by Mgat5-modified glycans restricts TCR recruitment to the site of antigen presentation. Dysregulation of Mgat5 in humans may increase susceptibility to autoimmune diseases, such as multiple sclerosis.

Animals↗

PubMed2001

Comparative expression of the mitotic regulators SAK and PLK in colorectal cancer.

BACKGROUND: Disruption of normal mechanisms for cell cycle regulation is important in carcinogenesis. SAK and PLK are members of the polo family of serine threonine kinases, which in lower organisms have been shown to be required for the precise regulation of mitosis. Studies of human polo family members have focused on PLK, which has been found to be overexpressed in several tumor types, with the degree of overexpression correlating with adverse clinical outcome. However, PLK expression had not previously been analyzed in colorectal cancer. SAK, a polo family member with unique properties, had not been systematically studied in any tumor type. METHODS: In this study, SAK expression was evaluated in a series of sporadic human colorectal cancer specimens (n = 74) and compared with that of PLK. Expression was assessed by reverse transcription-polymerase chain reaction. RESULTS: In the majority of cases, both SAK and PLK were more highly expressed in tumor tissue than in adjacent normal intestinal mucosa. Levels of SAK and PLK expression in tumor relative to paired normal mucosa correlated directly with patient age and with each other but did not correlate with tumor stage. These results suggest a mechanism for augmented disruption of mitotic regulation in older patients. CONCLUSIONS: The polo family mitotic regulators SAK and PLK are both aberrantly expressed in colorectal cancer. The potential prognostic significance of SAK and PLK expression in colorectal cancer will be evaluated in the future.

Adult↗

PubMed2001

Genetic defects in N-glycosylation and cellular diversity in mammals.

Glycoproteins in mammalian cells are modified with complex-type aspargine-linked glycans of variable chain lengths and composition. Observations of mice carrying mutations in glycosyltransferase genes imply that N-glycan structures regulate T-cell receptor clustering and hence sensitivity to agonists. We argue that the heterogeneity inherent in N-glycosylation contributes to cellular diversity and, thereby, to adaptability in the immune system.

Animals↗

PubMed2001

Complementary expression patterns of six nonessential Caenorhabditis elegans core 2/I N-acetylglucosaminyltransferase homologues.

The Caenorhabditis elegans genome contains 18 sequences related to mammalian core 2/I N-acetylglucosaminyltransferases. The six most closely related genes (gly-1 and gly-15 to gly-19) likely encode active enzymes, because are all transcribed and do not appear to be pseudogenes. Polypeptide divergence and the gene structures are both concordant with a common ancestor at the time of radiation from mammals that underwent three rounds of duplication and, most recently, a tandem duplication. Polypeptide alignments with mammalian homologues do not indicate whether the enzyme specificities are core 2, 4, or I-like or novel, but do clearly demonstrate the secondary structure characteristics of glycosyltransferases. The six homologues have essentially nonoverlapping expression patterns, unrelated by tissue type or cell lineage. The extent varies widely; gly-15 is expressed only in two gland cells, whereas gly-18 is broadly expressed in diverse cell types. gly-1, -15, -18 and -19 are expressed during adulthood; gly-16 and gly-17 appear to be restricted to embryonic or early larval stages. The parsimonious interpretation of the expression pattern and sequence data is that the catalytic activities are similar but with diverged promoters. Null alleles of three of the genes were generated without causing gross abnormality in homozygous animals. RNA-mediated interference experiments also failed to induce defects in the four genes tested. Nevertheless, the nematode has evolved six diverged core 2 GlcNAc-T-like genes, and we postulate that these arose in response to selection pressures to which C. elegans is not ordinarily subjected in the laboratory.

Amino Acid Sequence↗

PubMed2001

Long-term follow-up of prophylactic greenfield filters in multisystem trauma patients.

PURPOSE: The efficacy of prophylactic Greenfield filter (PGF) placement in multisystem trauma patients at high risk for venous thromboembolism has been established. The objective of this study is to demonstrate the long-term safety and durability of PGFs placed in young, active trauma patients. METHODS: Starting January 1992, all trauma patients at high risk for development of venous thromboembolism were identified for placement of PGFs. These included patients with ISS > 9 and severe closed head injury, spinal injury, pelvic fracture, multiple long bone fracture, or direct iliofemoral venous injury. Four patients with PGFs placed before the study protocol in 1992 were also included. Follow-up was attempted in all patients with at least 5 years' duration since placement of PGF using physical examination, duplex ultrasound (US), and plain abdominal radiograph. RESULTS: There were 108 patients who had a PGF placement during the period of January 1992 to June 1994 that were eligible for the study. Eighteen of these 108 (17%) patients died. Twelve of them (67%) had autopsies and medical records available to determine the cause of death, none from a pulmonary embolism. The average time of death was 2.7 months after injury. The remaining 90 patients and the 4 patients with PGFs placed before 1992 were sought for follow-up, but only 36 patients could be contacted, 33 of which returned for evaluation (35%). The mean time of follow-up from the time of injury to the time of examination was 67.7 months, and the mean age at follow-up was 38.1 years. Although six patients had mild to moderate lower extremity edema on physical examination, none of them had a deep venous thrombosis by US. Radiographs obtained in 19 of the 33 patients showed no migration or breakdown of the Greenfield filters in place. CONCLUSION: Prophylactic Greenfield filters in high-risk trauma patients are safe and durable. There appears to be no significant consequence in over 5 years of follow-up.

Adult↗

PubMed2000

Decreased UDP-GlcNAc levels abrogate proliferation control in EMeg32-deficient cells.

The hexosamine pathway provides UDP-N:-acetylhexosamine donor substrates used in cytosolic and Golgi-mediated glycosylation of proteins and for formation of glycosylphosphatidylinositol (GPI) anchors, which tether proteins to the outer plasma membrane. We have recently identified the murine glucosamine-6-phosphate (GlcN6P) acetyltransferase, EMeg32, as a developmentally regulated enzyme on the route to UDP-N:-acetylglucosamine (UDP-GlcNAc). Here we describe embryos and cells that have the EMeg32 gene inactivated by homologous recombination. Homozygous mutant embryos die at around embryonic day (E) 7.5 with a general proliferative delay of development. In vitro differentiated EMeg32(-/-) ES cells show reduced proliferation. Mouse embryonic fibroblasts (MEFs) deficient for EMeg32 exhibit defects in proliferation and adhesiveness, which could be complemented by stable re-expression of EMeg32 or by nutritional restoration of intracellular UDP-GlcNAc levels. Reduced UDP-GlcNAc levels predominantly translated into decreased O-GlcNAc modifications of cytosolic and nuclear proteins. Interestingly, growth-impaired EMeg32(-/-) MEFs withstand a number of apoptotic stimuli and express activated PKB/AKT. Thus, EMeg32-dependent UDP-GlcNAc levels influence cell cycle progression and susceptibility to apoptotic stimuli.

Acetyltransferases↗

PubMed2000

A homogeneous cell-based assay to identify N-linked carbohydrate processing inhibitors.

Malignant transformation is accompanied by altered cell surface glycosylation. N-Linked oligosaccharides carrying beta1-6GlcNAc branches are associated with tumor invasion and metastasis. Therefore, compounds that can enter cells and block biosynthesis of beta1-6GlcNAc-branched glycans without overt cytotoxicity are potential anticancer agents. We have developed a homogeneous cell-based assay for detection of such compounds. The method enables identification of agents that block beta1-6GlcNAc-branched glycan expression after incubation for 16-20 h with MDAY-D2 tumor cells, thereby protecting the cells from the subsequent addition of leukoagglutinin, a cytotoxic plant lectin. We observed that MDAY-D2 cell number is directly proportional to the level of endogenous alkaline phosphatase activity measured spectrophotometrically in cultures after the addition of substrate. The alkaline phosphatase assay was capable of detecting as few as 1,500 cells. The assay was readily adapted for high-throughput screening as reagent costs are low and no cell harvesting and washing steps are required. Under high-throughput operating conditions, the coefficient of variation for controls was found to be 4.2%. The results suggest that measurement of alkaline phosphatase in this cell assay format may be adapted for wider applications in high-throughput screenings for compounds that relieve cells from other growth inhibitors.

Alkaline Phosphatase↗

PubMed2000

Sak kinase gene structure and transcriptional regulation.

The Sak gene encodes a serine/threonine kinase, which is a member of the Polo family of mitotic regulators. Sak transcripts are present in S/G2/M phase cells, and in proliferating cell layers of the mouse embryo and adult tissues. In this report, we have characterized the murine Sak gene structure, the Sak chromosomal location, and identified the promoter. The murine Sak gene is located on the proximal arm of mouse chromosome 13, as determined by RFLP analysis. The murine gene comprises 15 coding exons spanning 16kb of genomic sequence, and encodes two alternately spliced transcripts. Sak-a, the predominant transcript, is encoded by 15 exons, while early termination of transcription and alternative splicing at exons 5 and 6 results in Sak-b. This truncated transcript encodes the complete kinase domain and a carboxyl end translated from 147bp of sequence contiguous with exon 5. Human Sak-a (Stk18) cDNA is reported to contain an insertion of sequence corresponding to the mouse Sak-b tail. Primer extension analysis of murine Sak revealed one major transcription start site at position -303bp relative to the start of translation. A genomic fragment of 3.5kb located 5' of the Sak transcriptional start drives expression of a luciferase-reporter gene in CHO and GC1-SPG cells in an orientation-dependent fashion. Using various Sak promoter/luciferase constructs, the core promoter region required for expression was located within 400bp of the message Cap site, and sequence further 5' strongly suppressed transcription.

Animals↗

PubMed2000

Regulation of human monocyte proMMP-9 production by fetuin, an endogenous TGF-beta antagonist.

Members of the matrix metalloproteinase family of enzymes degrade specific components of the extracellular matrix. MMP-9 is a type IV/V collagenase necessary for normal osteogenesis and is increased in inflammatory and neoplastic conditions. In such destructive diseases as emphysema and arthritis, and in epithelial cancers, MMP-9 is produced by cells of the monocyte lineage. Fetuin, a prominent serum glycoprotein, binds to and inactivates TGF-beta family members and through this mechanism regulates osteogenesis (Binkert et al., 1999, J Biol Chem 274:28514-28520.). We studied the effects of TGF-beta1 and fetuin on proMMP-9 release by the human monocyte line THP-1. Exogenous TGF-beta1 stimulated proMMP-9 release by THP-1 cells, with half-maximal stimulation at approximately 0.01 ng/ml TGF-beta1. Human fetuin (0.5-2 microM) partially inhibited this stimulation. Human fetuin alone stimulated THP-1 monocyte proMMP-9 release, with half maximal stimulation at approximately 0.25 microM fetuin. Neutralizing antibody specific for TGF-beta1 also stimulated proMMP-9 release, suggesting that endogenously-derived TGF-beta1 has an inhibitory effect. In freshly isolated human peripheral blood monocytes, fetuin stimulated proMMP-9 release with a dose-response curve similar to that observed in THP-1 cells. Human fetuin also activated proMMP-9 present in THP-1 conditioned medium. Taken together, these data suggest that under physiological conditions, fetuin facilitates matrix degradation by monocyte-derived MMP-9, both by opposing the autocrine inhibitory effect of endogenous TGF-beta1 on proMMP-9 release, and by activating proMMP-9 in the pericellular milieu. Conversely, fetuin may limit the stimulation of monocyte proMMP-9 release by high levels of exogenous TGF-beta1. Such modulation could prove important under pathological conditions where TGF-beta1 derived from paracrine sources is abundant, such as in epithelial malignancies.

Collagenases↗

PubMed2000

Suppression of tumor growth and metastasis in Mgat5-deficient mice.

Golgi beta1,6N-acetylglucosaminyltransferase V (MGAT5) is required in the biosynthesis of beta1,6GlcNAc-branched N-linked glycans attached to cell surface and secreted glycoproteins. Amounts of MGAT5 glycan products are commonly increased in malignancies, and correlate with disease progression. To study the functions of these N-glycans in development and disease, we generated mice deficient in Mgat5 by targeted gene mutation. These Mgat5-/- mice lacked Mgat5 products and appeared normal, but differed in their responses to certain extrinsic conditions. Mammary tumor growth and metastases induced by the polyomavirus middle T oncogene was considerably less in Mgat5-/- mice than in transgenic littermates expressing Mgat5. Furthermore, Mgat5 glycan products stimulated membrane ruffling and phosphatidylinositol 3 kinase-protein kinase B activation, fueling a positive feedback loop that amplified oncogene signaling and tumor growth in vivo. Our results indicate that inhibitors of MGAT5 might be useful in the treatment of malignancies by targeting their dependency on focal adhesion signaling for growth and metastasis.

Animals↗

PubMed2000

Continued experience with physical examination alone for evaluation and management of penetrating zone 2 neck injuries: results of 145 cases.

PURPOSE: Our preliminary experience with physical examination alone in the evaluation of penetrating zone 2 neck injuries for vascular trauma was previously reported in 28 patients over a 2-year period (1991-1993). The purpose of the current study was to examine the results of this approach in a much larger group of patients over an 8-year period. METHODS: The medical records for all patients admitted to our level I trauma center (all of them entered into our prospective protocol) between December 1991 and April 1999 with penetrating zone 2 neck trauma were reviewed for their initial presentation and any documented vascular injury. RESULTS: A total of 145 patients made up the study group; in 30 of these patients, the penetrating trajectory also traversed zone 1 or 3. Thirty-one patients (21%) had hard signs of vascular injury (active bleeding, expanding hematoma, bruit/thrill, pulse deficit, central neurologic deficit) and were taken immediately to the operating room; 28 (90%) of these 30 patients had either major arterial or venous injuries requiring operative repair (the false-positive rate for physical examination thus being 10%). Of the 114 patients with no hard signs, 23 underwent arteriography because of proximity of the injury to the vertebral arteries or because the trajectory included another zone. Of these 23 arteriograms, three showed abnormalities, but only one required operative repair. This case had no complications relating to the initial delay. The remaining 91 patients with no hard signs were observed without imaging or surgery for a minimum of 23 hours, and none had any evidence of vascular injury during hospitalization or during the initial 2-week follow-up period (1/114; false-negative rate for physical examination, 0.9%). CONCLUSIONS: This series confirms the earlier report indicating that patients with zone 2 penetrating neck wounds can be safely and accurately evaluated by physical examination alone to confirm or exclude vascular injury. The missed-injury rate is 0.7% (1/145) with this approach, which is comparable to arteriography in accuracy but less costly and noninvasive. Long-term follow-up is needed to confirm this management option.

Adult↗

PubMed2000

Biogenesis of multilamellar bodies via autophagy.

Transfection of Mv1Lu mink lung type II alveolar cells with beta1-6-N-acetylglucosaminyl transferase V is associated with the expression of large lysosomal vacuoles, which are immunofluorescently labeled for the lysosomal glycoprotein lysosomal-associated membrane protein-2 and the beta1-6-branched N-glycan-specific lectin phaseolis vulgaris leucoagglutinin. By electron microscopy, the vacuoles present the morphology of multilamellar bodies (MLBs). Treatment of the cells with the lysosomal protease inhibitor leupeptin results in the progressive transformation of the MLBs into electron-dense autophagic vacuoles and eventual disappearance of MLBs after 4 d of treatment. Heterologous structures containing both membrane lamellae and peripheral electron-dense regions appear 15 h after leupeptin addition and are indicative of ongoing lysosome-MLB fusion. Leupeptin washout is associated with the formation after 24 and 48 h of single or multiple foci of lamellae within the autophagic vacuoles, which give rise to MLBs after 72 h. Treatment with 3-methyladenine, an inhibitor of autophagic sequestration, results in the significantly reduced expression of multilamellar bodies and the accumulation of inclusion bodies resembling nascent or immature autophagic vacuoles. Scrape-loaded cytoplasmic FITC-dextran is incorporated into lysosomal-associated membrane protein-2-positive MLBs, and this process is inhibited by 3-methyladenine, demonstrating that active autophagy is involved in MLB formation. Our results indicate that selective resistance to lysosomal degradation within the autophagic vacuole results in the formation of a microenvironment propicious for the formation of membrane lamella.

Animals↗

PubMed2000

Minimal catalytic domain of N-acetylglucosaminyltransferase V.

UDP-GlcNAc: Manalpha1-6Manbeta-R beta1-6 N-acetylglucosaminyltransferase V (EC 2.4.1.155, GlcNAc-TV) is a Golgi enzyme that substitutes the trimannosyl core in the biosynthetic pathway for complex-type N-linked glycans. GlcNAc-TV activity is regulated by oncogenes frequently activated in cancer cells ( ras, src, and her2/neu ) and by activators of T lymphocytes. Overexpression of GlcNAc-TV in epithelial cells results in morphological transformation, while tumor cell mutants selected for loss of GlcNAc-TV products show diminished malignant potential in mice. In this report, we have expressed and characterized a series of N- and C-terminal deletions of GlcNAc-TV. Portions of GlcNAc-TV sequence were fused at the N-terminal domain to IgG-binding domains of staphylococcal Protein A and expressed in CHOP cells. The secreted fusion proteins were purified by IgG Sepharose affinity chromatography and assayed for enzyme activities. The peptide sequence S(213-740)of GlcNAc-TV was determined to be essential for the catalytic activity, the remaining amino acids comprising a 183 amino acid stem region, a 17 amino acid transmembrane domain and a 12 amino acid cytosolic moiety. Further deletion of 5 amino acids to produce peptide R(218-740)reduced enzyme activity by 20-fold. Similar K(m)and V(max)values for donor and acceptor were observed for peptide S(213-740), the minimal catalytic domain, and peptide Q(39-740), which also included the stem region. Truncation of five amino acids from the C-terminus also resulted in a 20-fold loss of catalytic activity. Secondary structure predictions suggest a high frequency of turns in the stem region, and more contiguous stretches of alpha-helix found in the catalytic domain.

Amino Acid Sequence↗

PubMed1999

Glycoprotein glycosylation and cancer progression.

Glycosylation of glycoproteins and glycolipids is one of many molecular changes that accompany malignant transformation. GlcNAc-branched N-glycans and terminal Lewis antigen sequences have been observed to increase in some cancers, and to correlate with poor prognosis. Herein, we review evidence that beta1, 6GlcNAc-branching of N-glycans contributes directly to cancer progression, and we consider possible functions for the glycans. Mgat5 encodes N-acetylglucosaminyltransferase V (GlcNAc-TV), the Golgi enzyme required in the biosynthesis of beta1,6GlcNAc-branched N-glycans. Mgat5 expression is regulated by RAS-RAF-MAPK, a signaling pathway commonly activated in tumor cells. Ectopic expression of GlcNAc-TV in epithelial cells results in morphological transformation and tumor growth in mice, and over expression in carcinoma cells has been shown to induce metastatic spread. Ectopic expression of GlcNAc-TIII, an enzyme that competes with GlcNAc-TV for acceptor, suppresses metastasis in B16 melanoma cells. Furthermore, breast cancer progression and metastasis induced by a viral oncogene expressed in transgenic mice is markedly suppressed in a GlcNAc-TV-deficient background. Mgat5 gene expression and beta1, 6GlcNAc-branching of N-glycans are associated with cell motility, a required phenotype of malignant cells.

Amino Sugars↗

PubMed1999

Overexpression of the alpha2,6-sialyltransferase, ST6Gal I, in a low metastatic variant of a murine lymphoblastoid cell line is associated with appearance of a unique ST6Gal I mRNA.

Multiple mRNA isoforms are generated from Siat1, the gene encoding ST6Gal I (beta-galactoside alpha2,6-sialyltransferase, SiaT-1, ST6N, alpha2,6ST). These isoforms, transcriptionally initiated from a number of physically distinct promoter regions, differ only in the 5'-most untranslated region and share an identical ST6Gal I coding region. W16 cells, a spontaneous mutant from MDAY-D2, the highly metastatic murine lymphoid tumor cell line, is considerably less metastatic and exhibits significantly slower tumor growth characteristics [R. Takano, E. Muchmore, and J. W. Dennis (1994) Glycobiology 4, 665-674]. Takano et al. further reported that ST6Gal I mRNA in W16 is elevated 40-fold compared to the parental cells. Here, by means of 5'-RACE analysis, we demonstrate a heretofore undocumented ST6Gal I mRNA form expressed in W16 cells. This ST6Gal I mRNA contains a novel 5'-most untranslated region with 96% sequence similarity to the retroviral-like transposable element, intracisternal particle A (IAP). This observation suggests the notion that elevated ST6Gal I expression in W16 cells is the result of DNA rearrangement in the Siat1 locus. Atypical transcriptional activation of Siat1 is the result of this IAP transposition.

Animals↗