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1.  Mammalian WTAP is a regulatory subunit of the RNA N6-methyladenosine methyltransferase 
Cell Research  2014;24(2):177-189.
The methyltransferase like 3 (METTL3)-containing methyltransferase complex catalyzes the N6-methyladenosine (m6A) formation, a novel epitranscriptomic marker; however, the nature of this complex remains largely unknown. Here we report two new components of the human m6A methyltransferase complex, Wilms' tumor 1-associating protein (WTAP) and methyltransferase like 14 (METTL14). WTAP interacts with METTL3 and METTL14, and is required for their localization into nuclear speckles enriched with pre-mRNA processing factors and for catalytic activity of the m6A methyltransferase in vivo. The majority of RNAs bound by WTAP and METTL3 in vivo represent mRNAs containing the consensus m6A motif. In the absence of WTAP, the RNA-binding capability of METTL3 is strongly reduced, suggesting that WTAP may function to regulate recruitment of the m6A methyltransferase complex to mRNA targets. Furthermore, transcriptomic analyses in combination with photoactivatable-ribonucleoside-enhanced crosslinking and immunoprecipitation (PAR-CLIP) illustrate that WTAP and METTL3 regulate expression and alternative splicing of genes involved in transcription and RNA processing. Morpholino-mediated knockdown targeting WTAP and/or METTL3 in zebrafish embryos caused tissue differentiation defects and increased apoptosis. These findings provide strong evidence that WTAP may function as a regulatory subunit in the m6A methyltransferase complex and play a critical role in epitranscriptomic regulation of RNA metabolism.
PMCID: PMC3915904  PMID: 24407421
WTAP; m6A methyltransferase; METTL3; METTL14; mRNA
2.  Synthesis of Binucleating Macrocycles and their Nickel(II) Hydroxo- and Cyano-Bridged Complexes with Divalent Ions: Anatomical Variation of Ligand Features 
Inorganic chemistry  2012;51(20):11017-11029.
The planar NNN-pincer complexes [MII(pyN Me2)(OH)]1− (MII = Ni, Cu) fix CO2 in η1-OCO2H complexes; results for the copper system are described. MnII, FeII, CoII, and ZnII behave differently, forming [MII(pyN2Me)2]2− with N4O2 coordination. Incorporation of the NiII pincer into binucleating macrocycle 2 containing a triamino MII locus connected by two 1,3-biphenylene groups affords proximal NiII and MII sites for investigation of the synthesis, structure, and reactivity of Ni-X-M bridge units. This ligand structure is taken as a reference for variations in MII atoms and binding sites and bridges X = OH− and CN− to produce additional members of the macrocyclic family with improved properties. Macrocycle 2 with a 22-membered ring is shown to bind MII = Mn, Fe, and Cu with hydroxo bridges. Introduction of the 4-BuiO group (macrocycle 3) improves the solubility of neutral complexes such as those with NiII-OH-CuII and NiII-CN-FeII bridges. The syntheses of macrocycle 5 with a 7-Me-[12]aneSN3 and macrocycle 6 with a 1,8-Me2-[14]aneN4 MII binding site are described to together with hydoxo-bridged Ni-Cu and cyano-bridged Ni-Fe complexes. This work was motivated by the presence of a Ni⋯(HO)-Fe bridge grouping in a reactive state of carbon monoxide dehydrogenase. Attempted decrease in Ni-(OH)-M distances (3.70-3.87 Å) to smaller values observed in the enzyme by use of macrocycle 4 having 1,2-biphenylene connectors led to a mononuclear octahedral NiII complex. Bridge structural units are summarized and the structures of fourteen macrocyclic complexes including eight with bridges are described.
PMCID: PMC3483644  PMID: 23030366
3.  Risk Factors for Small Airway Obstruction among Chinese Island Residents: A Case-Control Study 
PLoS ONE  2013;8(7):e68556.
We investigated the prevalence of and risk factors for small airway obstruction (SAO) among Chinese island residents to establish means to prevent and treat SAO.
From October 17, 2011 to November 1, 2011, a total of 2,873 residents aged >20 years who lived on the Huangqi Peninsula of Fujian were recruited by random cluster sampling. They were asked to complete a Burden of Obstructive Lung Disease (BOLD) questionnaire and underwent physical examinations and lung function evaluations. SAO was defined as a forced expiratory flow at 50% of vital capacity, Vmax50%, of less than 70% of predicted. Risk factors for SAO were assessed from among demographic and anthropometric variables, blood chemistry results, and questionnaire response items.
A total of 216 (7.52%) Chinese island residents were identified as having SAO (95 males; 121 females). Their survey and test results were compared with 432 age and sex-matched healthy controls (192 males; 240 females) for SAO risk factors. Among numerous factors investigated, only diabetes mellitus (p = 0.039), smoking index (SI, p<0.001 for SI>600), second hand smoke (p = 0.002), and lack of regular exercise (p<0.001) were significant risk factors for SAO.
The risk factors for SAO among Chinese island residents appeared to be similar to those among people who live in high-density urban environments and impoverished rural areas. Public health policies and medical practices directed toward improving respiratory health for island residents should be comparable to those used for urban and rural dwellers.
PMCID: PMC3715483  PMID: 23874670
4.  Receptor-Mediated Activation of a Proinsulin-Transferrin Fusion Protein in Hepatoma Cells 
A proinsulin-transferrin (ProINS-Tf) recombinant fusion protein was designed and characterized for the sustained release of an active form of insulin (INS) by hepatoma cells. During incubation with H4IIE hepatoma cells, a gradual decline of ProINS-Tf concentration, with a concomitant generation of the immuno-reactive insulin-transferrin (irINS-Tf), was detected in the culture medium by using INS- or proinsulin (ProINS)-specific radioimmunoassay (RIA) system. Further studies indicated that the conversion of ProINS-Tf to irINS-Tf was a transferrin receptor (TfR) mediated process that was pH-sensitive, and temperature- and microtubule-dependent. These results suggest that the conversion occurred during the slow recycling route of transferrin (Tf)-TfR pathway, possibly processed by proteases in the slow recycling compartments juxtaposed to the trans-Golgi network (TGN). ProINS-Tf exhibited little activity in the short-term promotion of glucose uptake in adipocytes, indicating that it was in an inactive form similar to ProINS. Stimulation of Akt phosphorylation by ProINS-Tf was detected only after prolonged incubation with H4IIE cells. On the other hand, ProINS-Tf pre-incubated with H4IIE cells for 24 h acquired an immediate activity of stimulating Akt phosphorylation. Furthermore, ProINS-Tf elicited a strong activity in inhibition of glucose production following 24 h incubation with H4IIE cells. Based on these findings, we conclude that the Tf-TfR endocytosis and recycling pathway enables the conversion and release of ProINS-Tf in an active form of irINS-Tf. Results from this study suggest that the Tf-TfR pathway can be exploited for the design of prohormone-Tf fusion proteins as protein prodrugs for their sustained and targeted activation.
PMCID: PMC3196267  PMID: 21756950
Proinsulin; transferrin; insulin; prohormone conversion; receptor-mediated endocytosis and recycling; bifunctional fusion protein
5.  Local injection of lentivirus-delivered livinshRNA suppresses lung adenocarcinoma growth by inducing a G0/G1 phase cell cycle arrest 
The inhibitor of apoptosis protein (IAP) plays an important role in tumorigenesis and may be a potential target for cancer therapy. Livin, which belongs to this family, is highly expressed in various tumors. The previous study demonstrated that silencing Livin gene promoted lung cancer cell apoptosis; however, the effects on tumor growth suppression by targeting this gene in vivo, to thereby determine the efficacy of targeting Livin for patient therapy, have not been determined. This study injected lentivirus-delivered livinshRNA into established xenograft tumors derived from the lung adenocarcinoma cell line SPC-A-1 in BALB/C nude mice, the result showed that LivinshRNA down-regulated Livin expression effectively, induced tumor cell apoptosis, reduced tumor cell proliferation, and suppressed tumor growth dramatically, with a tumor volume inhibitory rate of (58.65±4.82)% and a tumor weight inhibitory rate of (47.44±1.64)%, but with less severe adverse reaction to the mouse. This study further demonstrated that Livin gene silencing induced a G0/G1-phase cell cycle arrest and cyclin D1 downregulation, which is a key regulator of the G0/G1- to S-phase transition. These findings suggest that LivinshRNA local injection may serve as a therapeutic method for patient treatment, and that LivinshRNA may suppress tumor growth by arresting the cell cycle in the G0/G1-phase.
PMCID: PMC3466986  PMID: 23071862
Livin gene; RNA interference; lung adenocarcinoma; xenograft tumor model; cell cycle
6.  Optimizing the accuracy and precision of the single-pulse Laue technique for synchrotron photo-crystallography 
Journal of Synchrotron Radiation  2010;17(Pt 4):479-485.
The accuracy achieved in single-pulse pump-probe Laue experiments at beamline 14-ID at APS is estimated to be 3–4%.
The accuracy that can be achieved in single-pulse pump-probe Laue experiments is discussed. It is shown that with careful tuning of the experimental conditions a reproducibility of the intensity ratios of equivalent intensities obtained in different measurements of 3–4% can be achieved. The single-pulse experiments maximize the time resolution that can be achieved and, unlike stroboscopic techniques in which the pump-probe cycle is rapidly repeated, minimize the temperature increase due to the laser exposure of the sample.
PMCID: PMC2891491  PMID: 20567080
single-pulse diffraction; accuracy; Laue method; RATIO method; photo-crystallography
7.  Time-resolved synchrotron diffraction and theoretical studies of very short-lived photo-induced molecular species 
Excited-state geometries determined by time-resolved synchrotron diffraction are summarized with emphasis on their comparison with a series of theoretical results. The relative merits of monochromatic and polychromatic (Laue) techniques are discussed.
Definitive experimental results on the geometry of fleeting species are at the time of writing still limited to monochromatic data collection, but methods for modifications of the polychromatic Laue data to increase their accuracy and their suitability for pump–probe experiments have been implemented and are reviewed. In the monochromatic experiments summarized, excited-state conversion percentages are small when neat crystals are used, but are higher when photoactive species are embedded in an inert framework in supramolecular crystals. With polychromatic techniques and increasing source brightness, smaller samples down to tenths of a micrometre or less can be used, increasing homogeneity of exposure and the fractional population of the excited species. Experiments described include a series of transition metal complexes and a fully organic example involving excimer formation. In the final section, experimental findings are compared with those from theoretical calculations on the isolated species. Qualitative agreement is generally obtained, but the theoretical results are strongly dependent on the details of the calculation, indicating the need for further systematic analysis.
PMCID: PMC2824528  PMID: 20164641
pump–probe experiments; time-resolved diffraction; excited-state molecular geometries; excimers

Results 1-7 (7)