Li-Jyun Syu - Academia.edu (original) (raw)

Papers by Li-Jyun Syu

Journal of Molecular Biology, 1997

The fate of the genome of the polyoma (Py) tumor virus following integration in the chromosomes o... more The fate of the genome of the polyoma (Py) tumor virus following integration in the chromosomes of transformed rat FR3T3 cells was re-examined. The viral sequences were integrated at a single transformant-speci®c chromosomal site in each of 22 transformants tested. In situ ampli®cation of the viral sequences was observed in 24 of 34 transformants analyzed. Large T antigen, the unique viral function involved in initiating DNA replication from the viral origin, was essential for the ampli®cation process. There was an absolute requirement for a reiteration of viral sequences and the extent of the reiteration affected the degree of ampli®cation. The reiteration may be important for homologous recombination-mediated resolution of in situ ampli®ed sequences. Among 11 transformants harboring a 1 to 2 kb repeat, the degree of ampli®cation was transformant-speci®c and varied over a wide range. At the high end of the spectrum, the genome copy number increased 1300-fold at steady state, while at the low end, ampli®cation was below twofold. Some aspect of the host chromatin at the site integration that affected viral gene expression, also directly or indirectly modulated the ampli®cation.

Oncotarget, Jan 3, 2016

Gastric adenocarcinoma is the third most common cause of cancer-related death worldwide. Here we ... more Gastric adenocarcinoma is the third most common cause of cancer-related death worldwide. Here we report a novel, highly-penetrant mouse model of invasive gastric cancer arising from deregulated Hedgehog/Gli2 signaling targeted to Lgr5-expressing stem cells in adult stomach. Tumor development progressed rapidly: three weeks after inducing the Hh pathway oncogene GLI2A, 65% of mice harbored in situ gastric cancer, and an additional 23% of mice had locally invasive tumors. Advanced mouse gastric tumors had multiple features in common with human gastric adenocarcinomas, including characteristic histological changes, expression of RNA and protein markers, and the presence of major inflammatory and stromal cell populations. A subset of tumor cells underwent epithelial-mesenchymal transition, likely mediated by focal activation of canonical Wnt signaling and Snail1 induction. Strikingly, mTOR pathway activation, based on pS6 expression, was robustly activated in mouse gastric adenocarcinom...

PLoS ONE, 2012

Chronic inflammation in the stomach can lead to gastric cancer. We previously reported that gastr... more Chronic inflammation in the stomach can lead to gastric cancer. We previously reported that gastrin-deficient (Gast 2/2 ) mice develop bacterial overgrowth, inflammatory infiltrate, increased Il-1b expression, antral hyperplasia and eventually antral tumors. Since Hedgehog (Hh) signaling is active in gastric cancers but its role in precursor lesions is poorly understood, we examined the role of inflammation and Hh signaling in antral hyperplasia. LacZ reporter mice for Sonic hedgehog (Shh), Gli1, and Gli2 expression bred onto the Gast 2/2 background revealed reduced Shh and Gli1 expression in the antra compared to wild type controls (WT). Gli2 expression in the Gast 2/2 corpus was unchanged. However in the hyperplastic Gast 2/2 antra, Gli2 expression increased in both the mesenchyme and epithelium, whereas expression in WT mice remained exclusively mesenchymal. These observations suggested that Gli2 is differentially regulated in the hyperplastic Gast 2/2 antrum versus the corpus and by a Shh ligand-independent mechanism. Moreover, the proinflammatory cytokines Il-1b and Il-11, which promote gastric epithelial proliferation, were increased in the Gast 2/2 stomach along with Infc. To test if inflammation could account for elevated epithelial Gli2 expression in the Gast 2/2 antra, the human gastric cell line AGS was treated with IL-1b and was found to increase GLI2 but decrease GLI1 levels. IL-1b also repressed human GAST gene expression. Indeed, GLI2 but not GLI1 or GLI3 expression repressed gastrin luciferase reporter activity by ,50 percent. Moreover, chromatin immunoprecipitation of GLI2 in AGS cells confirmed that GLI2 directly binds to the GAST promoter. Using a mouse model of constitutively active epithelial GLI2 expression, we found that activated GLI2 repressed Gast expression but induced Il-1b gene expression and proliferation in the gastric antrum, along with a reduction of the number of G-cells. In summary, epithelial Gli2 expression was sufficient to stimulate Il-1b expression, repress Gast gene expression and increase proliferation, leading to antral hyperplasia.

Nucleic Acids Research, 2005

At the molecular level, members of the NKx2.2 family of transcription factors establish neural co... more At the molecular level, members of the NKx2.2 family of transcription factors establish neural compartment boundaries by repressing the expression of homeobox genes specific for adjacent domains [Muhr et al. (2001) Cell, 104, 861-873; Weiss et al. (1998) Genes Dev., 12, 3591-3602]. The Drosophila homologue, vnd, interacts genetically with the high-mobility group protein, Dichaete, in a manner suggesting co-operative activation ) Development, 129, 1165-1174. However, evidence for direct interactions and transcriptional activation is lacking. Here, we present molecular evidence for the interaction of Vnd and Dichaete that leads to the activation of target gene expression. Two-hybrid interaction assays indicate that Dichaete binds the Vnd homeodomain, and additional Vnd sequences stabilize this interaction. In addition, Vnd has two activation domains that are typically masked in the intact protein. Whether vnd can activate or repress transcription is contextdependent. Full-length Vnd, when expressed as a Gal4 fusion protein, acts as a repressor containing multiple repression domains. A divergent domain in the N-terminus, not found in vertebrate Vnd-like proteins, causes the strongest repression. The corepressor, Groucho, enhances Vnd repression, and these two proteins physically interact. The data presented indicate that the activation and repression domains of Vnd are complex, and whether Vnd functions as a transcriptional repressor or activator depends on both intra-and inter-molecular interactions.

Nature Genetics, 2008

Constitutive Hedgehog (Hh) signaling underlies several human tumors 1 , including basal cell carc... more Constitutive Hedgehog (Hh) signaling underlies several human tumors 1 , including basal cell carcinoma (BCC) and basaloid follicular hamartoma in skin 2,3 . Intriguingly, superficial BCCs arise as de novo epithelial buds resembling embryonic hair germs 4-6 , collections of epidermal cells whose development is regulated by canonical Wnt/β-catenin signaling 7,8 . Similar to embryonic hair germs, human BCC buds showed increased levels of cytoplasmic and nuclear β-catenin, and expressed early hair follicle lineage markers. We also detected canonical Wnt/β-catenin signaling in epithelial buds and hamartomas from mice expressing an oncogene, M2SMO 9 , leading to constitutive Hh signaling in skin. Conditional overexpression of the Wnt pathway antagonist Dkk1 in M2SMO-expressing mice potently inhibited epithelial bud and hamartoma development without affecting Hh signaling. Our findings uncover a hitherto unknown requirement for ligand-driven, canonical Wnt/β-catenin signaling for Hh pathway-driven tumorigenesis, identify a new pharmacological target for these neoplasms, and establish the molecular basis for the well-known similarity between early superficial BCCs and embryonic hair germs.

Molecular Cell, 1999

In addition, studies † Department of Physiology using endotoxin proteases and endosome ablation h... more In addition, studies † Department of Physiology using endotoxin proteases and endosome ablation have University of Michigan School of Medicine implicated VAMP2 as the relevant v-SNARE partici-Ann Arbor, Michigan 48109 pating in insulin-stimulated GLUT4 vesicle exocytosis In regulated exocytosis, the vesicular v-SNAREs directly interact with plasma membrane t-SNAREs for docking and subsequent fusion. Syntaxin 4 is the major Summary plasma membrane t-SNARE for insulin-stimulated GLUT4 translocation in adipocytes (Cheatham et al., 1996; Tim-Insulin-stimulated glucose transport and GLUT4 transmers et al., 1996; Volchuk et al., 1996; Olson et al., 1997).

Mechanisms of Development, 2007

The transcription factor, Vnd, is a dual regulator that specifies ventral neuroblast identity in ... more The transcription factor, Vnd, is a dual regulator that specifies ventral neuroblast identity in Drosophila by both repressing and activating target genes. Vnd and its homologues have a conserved amino acid sequence, the Nk-2 box or Nk specific domain, as well a conserved DNA-binding homeodomain and an EhI-type Groucho interaction domain. However, the function of the conserved Nk-2 box has not been fully defined. To explore its function, we deleted the Nk-2 box and compared the regulatory activity of mutant Vnd in transgenic over-expression assays to that of the wild-type protein. We were unable to assign regulatory activity to the Nk-2 box using an over-expression assay, because the mutant protein activated expression of endogenous Vnd, masking a requirement for the Nk-2 box. However, in transgenic rescue assays, Vnd lacking the Nk-2 box repressed ind expression at 30% lower levels than the wild-type protein. Moreover, in transient transfection assays using Gal4 DNA-binding domain-Vnd chimeras, the repression activity of Vnd lacking the Nk-2 box was compromised. Because Vnd represses target gene expression in conjunction with Groucho, we asked whether the Nk-2 box affects Vnd's ability to interact with this co-repressor. Vnd lacking the Nk-2 box binds Groucho 30% less efficiently than wild-type Vnd in co-immunoprecipitations. These data suggest that the Nk-2 box contributes to the repression activity of Vnd by stabilizing its interaction with the co-repressor, Groucho.

J Neurochem, 2002

Activation of protein kinase C (PKC) regulates the processing of Alzheimer amyloid precursor prot... more Activation of protein kinase C (PKC) regulates the processing of Alzheimer amyloid precursor protein (APP) into its soluble form (sAPP) and amyloid beta-peptide (A beta). However, little is known about the intermediate steps between PKC activation and modulation of APP metabolism. Using a specific inhibitor of mitogen-activated protein (MAP) kinase kinase activation (PD 98059), as well as a dominant negative mutant of MAP kinase kinase, we show in various cell lines that stimulation of PKC by phorbol ester rapidly induces sAPP secretion through a mechanism involving activation of the MAP kinase cascade. In PC12-M1 cells, activation of MAP kinase by nerve growth factor was associated with stimulation of sAPP release. Conversely, M1 muscarinic receptor stimulation, which is known to act in part through a PKC-independent pathway, increased sAPP secretion mainly through a MAP kinase-independent pathway. A beta secretion and its regulation by PKC were not affected by PD 98059, supporting the concept of distinct secretory pathways for A beta and sAPP formation.

Journal of Clinical Investigation, 2011

Uncontrolled Hedgehog (Hh) signaling leads to the development of basal cell carcinoma (BCC), the ... more Uncontrolled Hedgehog (Hh) signaling leads to the development of basal cell carcinoma (BCC), the most common human cancer, but the cell of origin for BCC is unclear. While Hh pathway dysregulation is common to essentially all BCCs, there exist multiple histological subtypes, including superficial and nodular variants, raising the possibility that morphologically distinct BCCs may arise from different cellular compartments in skin. Here we have shown that induction of a major mediator of Hh signaling, GLI2 activator (GLI2ΔN), selectively in stem cells of resting hair follicles in mice, induced nodular BCC development from a small subset of cells in the lower bulge and secondary hair germ compartments. Tumorigenesis was markedly accelerated when GLI2ΔN was induced in growing hair follicles. In contrast, induction of GLI2ΔN in epidermis led to the formation of superficial BCCs. Expression of GLI2ΔN at reduced levels in mice yielded lesions resembling basaloid follicular hamartomas, which have previously been linked to low-level Hh signaling in both mice and humans. Our data show that the cell of origin, tissue context (quiescent versus growing hair follicles), and level of oncogenic signaling can determine the phenotype of Hh/Gli-driven skin tumors, with high-level signaling required for development of superficial BCC-like tumors from interfollicular epidermis and nodular BCC-like tumors from hair follicle stem cells.

Journal of Cellular Biochemistry, 1997

We conditionally overexpressed a MEK1 mutant that contains triple mutations in the regulatory and... more We conditionally overexpressed a MEK1 mutant that contains triple mutations in the regulatory and kinase domains, and investigated its effects on the MAP kinase cascade in Swiss 3T3 cells. Expression of the mutant produced a 60% blockade in MAP kinase activity. However, only a modest blockade in DNA synthesis was observed, without any reductions in the phosphorylation of two proteins known to be substrates of MAP kinase. Moreover, the overexpression of MEK1(3A) failed to block endogenous MEK1 activation, although MEK1(3A) formed complexes with both c-Raf and B-Raf as well as p42/p44 MAPK. These results suggest that there may be multiple biochemical inputs into the MEK/MAPK pathway.

Journal of Biological Chemistry, 1998

The insulin-stimulated uptake of 2-(methylamino) isobutyric acid (MeAIB), a nonmetabolizable subs... more The insulin-stimulated uptake of 2-(methylamino) isobutyric acid (MeAIB), a nonmetabolizable substrate for system A, in 3T3-L1 adipocytes was investigated. As cells took on a more adipogenic phenotype, the insulinstimulated versus the saturable basal MeAIB uptake increased by 5-fold. The induced transport activity showed properties characteristic of system A, with a K m value of 190 M. The half-life of the induced system A activity was independent of de novo mRNA and protein synthesis and was not accelerated by ambient amino acids, therefore, it was mechanistically distinct from the previously described adaptive and hormonal regulation of system A. Inhibition of mitogen-activated protein kinase kinase by PD98059, Ras farnesylation by PD152440 and B581, p70 S6K by rapamycin, and phosphatidylinositol 3-kinase (PI 3-K) by wortmannin and LY294002 revealed that only wortmannin and LY294002 inhibited the insulin-induced MeAIB uptake with IC 50 values close to that previously reported for inhibition of PI 3-K. These results suggest that the Ras/mitogenactivated protein kinase and pp70 S6K insulin signaling pathways are neither required nor sufficient for insulin stimulation of MeAIB uptake, and activation of PI 3-K or a wortmannin/LY294002-sensitive pathway may play an important role in regulation of system A transport by insulin in 3T3-L1 cells.

Journal of Biological Chemistry, 1995

increase Ras-GTP loading by targeting SOS to the plasma membrane location of Ras through the smal... more increase Ras-GTP loading by targeting SOS to the plasma membrane location of Ras through the small adaptor protein Grb2. However, despite the continuous stimulation of receptor tyrosine kinase activity, Ras activation is transient and, in the case of insulin, begins returning to the GDP-bound state within 5 min. We report here that the cascade of serine kinases activated directly by Ras results in a mitogen-activated protein kinase kinase (MEK)-dependent phosphorylation of SOS and subsequent disassociation of the Grb2-SOS complex, thereby interrupting the ability of SOS to catalyze nucleotide exchange on Ras. These data demonstrate a molecular feedback mechanism accounting for the desensitization of Ras-GTP loading following insulin stimulation.

Endocrinology, 1997

To investigate the mechanism of action of the placental angiogenic hormone proliferin (PLF), we a... more To investigate the mechanism of action of the placental angiogenic hormone proliferin (PLF), we analyzed the signaling components in endothelial cells that are required for PLF-induced chemotaxis. Pertussis toxin, which inactivates Gi proteins, inhibited PLF-induced chemotaxis of endothelial cells. Gi proteins can lead to activation of the mitogen-activated protein kinase (MAPK) pathway; PLF was found to stimulate MAPK activity, and this induction was blocked by both pertussis toxin and a specific inhibitor of MAPK kinase, PD 098059. Furthermore, a blockade of MAPK activation prevented endothelial cell movement in response to PLF. As PLF functionally interacts with the insulin-like growth factor II (IGF-II)/mannose 6-phosphate receptor, we also examined the effects of pertussis toxin and PD 098059 on another ligand for this receptor, a mutant form of IGF-II; both inhibitors also block the action of this factor on endothelial cells. These data suggest that chemotaxis initiated by PLF and mediated by the IGF-II/mannose 6-phosphate receptor occurs through a G protein-coupled pathway, and that MAPK activation is necessary for the chemotactic response.

Brain Research, 2009

Consistent with the common role of Nkx6 family members in specifying motor neuron identity, we sh... more Consistent with the common role of Nkx6 family members in specifying motor neuron identity, we show that over-expression of Drosophila Nkx6 results in an increase in the number of Fasiclin II expressing motor neurons in the intersegmental nerve B branch. Our dissection of the regulatory domains of Nkx6 using chimeric cell culture assays revealed the presence of two repression domains and a single activation domain within this transcription factor. As well as its conserved homeodomain, Nkx6 also has a candidate Engrailed homology 1 (Eh1) domain that is conserved amongst all NKx6 family members, through which vertebrate NKx6-type proteins bind the co-repressor, Groucho (Muhr, J., et al., 2001. Groucho-mediated transcriptional repression establishes progenitor cell pattern and neuronal fate in the ventral neural tube. Cell 104, 861-73). Paralleling our previous reports that the Eh1 domain of Vnd and Ind are ineffective in Gal4 chimeric assays (Von Ohlen, T., Syu, L.J., Mellerick, D.M., 2007. Conserved properties of the Drosophila homeodomain protein.

The American Journal of Pathology, 2012

Gastric adenocarcinoma is one of the leading causes of cancer mortality worldwide. It arises thro... more Gastric adenocarcinoma is one of the leading causes of cancer mortality worldwide. It arises through a stepwise process that includes prominent inflammation with expression of interferon-␥ (IFN-␥) and multiple other pro-inflammatory cytokines. We engineered mice expressing IFN-␥ under the control of the stomach-specific H ؉ /K ؉ ATPase ␤ promoter to test the potential role of this cytokine in gastric tumorigenesis. Stomachs of H/K-IFN-␥ transgenic mice exhibited inflammation, expansion of myofibroblasts, loss of parietal and chief cells, spasmolytic polypeptide expressing metaplasia, and dysplasia. Proliferation was elevated in undifferentiated and metaplastic epithelial cells in H/K-IFN-␥ transgenic mice, and there was increased apoptosis. H/K-IFN-␥ mice had elevated levels of mRNA for IFN-␥ target genes and the pro-inflammatory cytokines IL-6, IL-1␤, and tumor necrosis factor-␣. Intracellular mediators of IFN-␥ and IL-6 signaling, pSTAT1 and pSTAT3, respectively, were detected in multiple cell types within stomach. H/K-IFN-␥ mice developed dysplasia as early as 3 months of age, and 4 of 39 mice over 1 year of age developed antral polyps or tumors, including one adenoma and one adenocarcinoma, which expressed high levels of nuclear ␤-catenin. Our data identified IFN-␥ as a pivotal secreted factor that orchestrates complex changes in inflammatory, epithelial, and mesenchymal cell populations to drive pre-neoplastic progression in stomach; however, additional alterations appear to be required for malignant conversion. Am J Pathol 2012, 181:2114 -2125; http://dx.doi.org/10.1016/j.ajpath.2012.08.017)