C Eric Thomas

University of Michigan, Ann Arbor, MI, United States

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Publications (4)16.3 Total impact

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    ABSTRACT: Eukaryotic cells have evolved an intricate system to resolve DNA damage to prevent its transmission to daughter cells. This system, collectively known as the DNA damage response (DDR) network, includes many proteins that detect DNA damage, promote repair, and coordinate progression through the cell cycle. Because defects in this network can lead to cancer, this network constitutes a barrier against tumorigenesis. The modular BRCA1 carboxyl-terminal (BRCT) domain is frequently present in proteins involved in the DDR, can exist either as an individual domain or as tandem domains (tBRCT), and can bind phosphorylated peptides. We performed a systematic analysis of protein-protein interactions involving tBRCT in the DDR by combining literature curation, yeast two-hybrid screens, and tandem affinity purification coupled to mass spectrometry. We identified 23 proteins containing conserved BRCT domains and generated a human protein-protein interaction network for seven proteins with tBRCT. This study also revealed previously unknown components in DNA damage signaling, such as COMMD1 and the target of rapamycin complex mTORC2. Additionally, integration of tBRCT domain interactions with DDR phosphoprotein studies and analysis of kinase-substrate interactions revealed signaling subnetworks that may aid in understanding the involvement of tBRCT in disease and DNA repair.
    Science Signaling 01/2012; 5(242):rs6. · 7.65 Impact Factor
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    C Eric Thomas, Yi-Chen Chen, George A Garcia
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    ABSTRACT: tRNA-guanine transglycosylases (TGTs) are responsible for incorporating 7-deazaguanine-modified bases into certain tRNAs in eubacteria (preQ(1)), eukarya (queuine) and archaea (preQ(0)). In each kingdom, the specific modified base is different. We have found that the eubacterial and eukaryal TGTs have evolved to be quite specific for their cognate heterocyclic base and that Cys145 (Escherichia coli) is important in recognizing the amino methyl side chain of preQ(1) (Chen et al., Nuc. Acids Res. 39 (2011) 2834 [15]). A series of mutants of the E. coli TGT have been constructed to probe the role of three other active site amino acids in the differential recognition of heterocyclic substrates. These mutants have also been used to probe the differential inhibition of E. coli versus human TGTs by pteridines. The results indicate that mutation of these active site amino acids can "open up" the active site, allowing for the binding of competitive pteridine inhibitors. However, even the "best" of these mutants still does not recognize queuine at concentrations up to 50μM, suggesting that other changes are necessary to adapt the eubacterial TGT to incorporate queuine into RNA. The pteridine inhibition results are consistent with an earlier hypothesis that pteridines may regulate eukaryal TGT activity (Jacobson et al., Nuc. Acids Res. 9 (1981) 2351 [8]).
    Biochemical and Biophysical Research Communications 06/2011; 410(1):34-9. · 2.28 Impact Factor
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    ABSTRACT: The Quantitative Assay Database (QuAD), http://proteome.moffitt.org/QUAD/, facilitates widespread implementation of quantitative mass spectrometry in cancer biology and clinical research through sharing of methods and reagents for monitoring protein expression and modification. Liquid chromatography coupled to multiple reaction monitoring (LC-MRM) mass spectrometry assays are developed using SDS-PAGE fractionated lysates from cancer cell lines. Pathway maps created using GeneGO Metacore provide the biological relationships between proteins and illustrate concepts for multiplexed analysis; each protein can be selected to examine assay development at the protein and peptide levels. The coupling of SDS-PAGE and multiple reaction monitoring mass spectrometry screening has been used to detect 876 peptides from 218 cancer-related proteins in model systems including colon, lung, melanoma, leukemias, and myeloma, which has led to the development of 95 quantitative assays including stable-isotope-labeled peptide standards. Methods are published online and peptide standards are made available to the research community. Protein expression measurements for heat shock proteins, including a comparison with ELISA and monitoring response to the HSP90 inhibitor, 17-(dimethylaminoethylamino)-17-demethoxygeldanamycin (17-DMAG), are used to illustrate the components of the QuAD and its potential utility. This resource enables quantitative assessment of protein components of signaling pathways and biological processes and holds promise for systematic investigation of treatment responses in cancer.
    PROTEOMICS - CLINICAL APPLICATIONS 04/2011; 5(7-8):383-96. · 1.81 Impact Factor
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    ABSTRACT: BACKGROUND: Urine is a useful source of protein for biomarker discovery and assessment because it is readily available, can be obtained by noninvasive collection methods, and enables monitoring of a wide range of physiologic processes and diseases. Urine aliquots provide enough protein for multiple analyses, combining current protocols with new techniques. CONCLUSIONS: Standardized collection and processing protocols are now being established and new methods for protein detection and quantification are emerging to complement traditional immunoassays. The current state of urine collection, specimen processing, and storage is reviewed with regard to discovery and quantification of protein biomarkers for cancer.
    Cancer Epidemiology Biomarkers &amp Prevention 03/2010; 19(4):953-9. · 4.56 Impact Factor