Total synthesis of tubulysin U and V.
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Molecular Conformation↗
Biomedical subjects
Yingchun Lu
Publications and source records attributed to Yingchun Lu.
alpha-E-catenin inactivation disrupts the cardiomyocyte adherens junction, resulting in cardiomyopathy and susceptibility to wall rupture.
BACKGROUND: alpha-E-catenin is a cell adhesion protein, located within the adherens junction, thought to be essential in directly linking the cadherin-based adhesion complex to the actin cytoskeleton. Although alpha-E-catenin is expressed in the adherens junction of the cardiomyocyte intercalated disc, and perturbations in its expression are observed in models of dilated cardiomyopathy, its role in the myocardium remains unknown. METHODS AND RESULTS: To determine the effects of alpha-E-catenin on cardiomyocyte ultrastructure and disease, we generated cardiac-specific alpha-E-catenin conditional knockout mice (alpha-E-cat cKO). alpha-E-cat cKO mice displayed progressive dilated cardiomyopathy and unique defects in the right ventricle. The effects on cardiac morphology/function in alpha-E-cat cKO mice were preceded by ultrastructural defects in the intercalated disc and complete loss of vinculin at the intercalated disc. alpha-E-cat cKO mice also revealed a striking susceptibility of the ventricular free wall to rupture after myocardial infarction. CONCLUSIONS: These results demonstrate a clear functional role for alpha-E-catenin in the cadherin/catenin/vinculin complex in the myocardium in vivo. Ablation of alpha-E-catenin within this complex leads to defects in cardiomyocyte structural integrity that result in unique forms of cardiomyopathy and predisposed susceptibility to death after myocardial stress. These studies further highlight the importance of studying the role of alpha-E-catenin in human cardiac injury and cardiomyopathy in the future.
Animals↗
Design and modular parallel synthesis of a MCR derived alpha-helix mimetic protein-protein interaction inhibitor scaffold.
A terphenyl alpha-helix mimetic scaffold recognized to be capable of disrupting protein-protein interactions was structurally morphed into an easily amenable and versatile multicomponent reaction (MCR) backbone. The design, modular in-parallel library synthesis, initial cell based biological data, and preliminary in vitro screening for the disruption of the Bcl-w/Bak protein-protein interaction by representatives of the MCR derived scaffold are presented.
Apoptosis↗
Gene silencing in androgen-responsive prostate cancer cells from the tissue-specific prostate-specific antigen promoter.
The success of gene therapy using a RNA interference approach relies on small interfering RNA (siRNA) expression from a highly tissue-specific RNA polymerase II promoter rather than from ubiquitous RNA polymerase III. Accordingly, we have developed a prostate-specific vector that expresses siRNAs from the human prostate-specific antigen promoter, a RNA polymerase II promoter. Our data demonstrate androgen-dependent and tissue-specific siRNA-mediated gene silencing in the androgen-responsive prostate cancer cell line, LNCaP. The biological significance was evidenced by altered apoptotic activity through the inhibition of the apoptosis-related regulatory gene. These results demonstrate that siRNA-mediated gene silencing from a tissue-specific RNA polymerase II promoter could be a potential tool for tissue-specific gene therapy.
Androgens↗
Poly(U) and polyadenylation termination signals are interchangeable for terminating the expression of shRNA from a pol II promoter.
Most short hairpin RNA (shRNA)-expressing vectors use RNA polymerase III promoters, as expression of shRNAs from RNA polymerase II promoters is not well understood, due to the lack of defined transcription initiation sites and functional localization of the transcription termination signal. Here we describe a modified cytomegavirus (CMV) promoter, an RNA polymerase II promoter, to express shRNAs with only four overhangs at the 5' end. The expression of shRNAs from the modified CMV promoter was terminated by the transcription termination, the polyadenylation, or the poly(U) termination signal. These results demonstrated that the poly(U) and polyadenylation termination signals are interchangeable for terminating the expression of shRNAs from the CMV promoter and result in similar efficacies in inhibiting endogenous target genes in mammalian cells.
Base Sequence↗
Solution phase parallel synthesis and evaluation of MAPK inhibitory activities of close structural analogues of a Ras pathway modulator.
A solution phase parallel synthesis approach was undertaken to rapidly explore the structure-activity relationship of an inhibitor of the Ras/Raf protein interaction identified from a small molecule compound library. Evaluation of the MAPK pathway signaling inhibitory activity of the synthesized analogues as well as their antiproliferative activity and ability to inhibit soft agar growth were performed.
Animals↗
An internal control for immunoblot analysis using the blotted membrane.
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Animals↗
Inhibitors of Ras/Raf-1 interaction identified by two-hybrid screening revert Ras-dependent transformation phenotypes in human cancer cells.
The interaction of activated Ras with Raf initiates signaling cascades that contribute to a significant percentage of human tumors, suggesting that agents that specifically disrupt this interaction might have desirable chemotherapeutic properties. We used a subtractive forward two-hybrid approach to identify small molecule compounds that block the interaction of Ras with Raf. These compounds (MCP1 and its derivatives, 53 and 110) reduced serum-induced transcriptional activation of serum response element as well as Ras-induced transcription by way of the AP-1 site. They also inhibited Ras-induced Raf-1 activation in human embryonic kidney 293 cells, Raf-1 and mitogen-activated protein kinase kinase 1 activities in HT1080 fibrosarcoma cells, and epidermal growth factor-induced Raf-1 activation in A549 lung carcinoma cells. The MCP compounds caused reversion of ras-transformed phenotypes including morphology, in vitro invasiveness, and anchorage-independent growth of HT1080 cells. Decreased level of matrix metalloproteinases was also observed. Further characterization showed that MCP compounds restore actin stress fibers and cause flat reversion in NIH 3T3 cells transformed with H-Ras (V12) but not in NIH 3T3 cells transformed with constitutively active Raf-1 (RafDeltaN). Finally, we show that MCP compounds inhibit anchorage-independent growth of A549 and PANC-1 cells harboring K-ras mutation. Furthermore, MCP110 caused G(1) enrichment of A549 cells with the decrease of cyclin D level. These results highlight potent and specific effects of MCP compounds on cancer cells with intrinsic Ras activation.
3T3 Cells↗
A beta-tubulin-like cDNA expressed specifically in elongating cotton fibers induces longitudinal growth of fission yeast.
Using cDNA Representational Difference Analysis (RDA) techniques, we isolated a cDNA that was expressed specifically in cotton fibers but not in the ovules of a fuzzless-lintless mutant (fl). We designated it as Gh-BTubL for it shares high sequence identity with known plant and yeast beta-tubulins. RT-PCR and robotic cDNA dot blot analyses indicated that the expression of Gh-BTubL was correlated with the elongation pattern of cotton fibers. In situ hybridization results verified that there was no Gh-BTubL mRNA in fl ovules while it was easily detected in the elongating wild type cotton fiber cells. Overexpression of Gh-BTubL in fission yeast induced longitudinal growth of the host cells by 1.74-fold, with no apparent effect on other aspects of the host cells. We suggest that Gh-BTubL plays an important role in cotton fiber elongation and we believe that elucidation of the control mechanisms for expression of tubulin-like proteins may help improve fiber quality and productivity.
Amino Acid Sequence↗