Quantitative Phosphoproteome Profiling of Wnt3a-mediated Signaling Network

Tang, Liu-Ya; Deng, Ning; Wang, Lian-Shui; Dai, Jie; Wang, Zhenglong; Jiang, Xueying; Li, Su-Jun; Li, Long; Sheng, Quanhu; Wu, Dianqing; Li, Lin; Zeng, Rong

doi:10.1074/mcp.m700120-mcp200

Cited by 57 publications

(17 citation statements)

References 84 publications

(89 reference statements)

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“…The biological role of USP10 is not fully understood, but it is known to interact with DNA-bound AR complexes 57 . USP10 has been detected as a phosphoprotein in Hela cells 35, 58 , HEK293 cells 59 , human embryonic kidney 293T cells 60 , and consistent with our data, also in LNCaP cells 20 . Physical interaction between USP10 and G3BP, a RasGAP interacting protein overexpressed in tumors, was shown by a two-hybrid screen to suggest a link between the ubiquitination pathway and Ras-mediated signaling 61 …”

Section: Discussionsupporting

confidence: 91%

Large scale phosphoproteome analysis of LNCaP human prostate cancer cells

Myung

Sadar

2012

Mol. BioSyst.

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Prostate cancer is the most frequently diagnosed cancer among men in the western world. The androgen receptor, a phosphoprotein, is suspected to be involved in all stages of the prostate cancer. Androgen receptor activity can be modulated by various kinases such as PKA, MAPK, AKT, and Src. Phosphorylation is an important post-translational modification and serves as a molecular on/off switch to regulate signaling. Disruptions of cellular phosphorylation are associated with various diseases such as cancer and kinases provide important drug targets. Here we present an analysis of the phosphoproteome in LNCaP human prostate cancer cells. The analytical strategy employed used proteomics based methodologies with a combination of detergent and chaotropic reagent during trypsin digestion followed by titanium dioxide enrichment of phosphopeptides. Over the course of multiple analyses by mass spectrometry we identified a total of 746 phosphorylation sites in 540 phosphopeptides corresponding to 116 phosphoproteins, of which 56 have not been previously reported. Phosphoproteins identified included transcription factors, co-regulators of the androgen receptor, and cancer-related proteins that include β-catenin, USP10, and histone deacetylase-2. The information of signaling pathways, motifs of phosphorylated peptides, biological processes, molecular functions, cellular components, and protein interactions from the identified phosphoproteins established a map of phosphoproteome and signaling pathways in LNCaP cells.

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Section: Discussionsupporting

confidence: 91%

Large scale phosphoproteome analysis of LNCaP human prostate cancer cells

Myung

Sadar

2012

Mol. BioSyst.

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“…4B). A previously identified single phosphorylation site on C8␤ Thr 364 is located at the end of the TMH2 region (38). The N-linked carbohydrates in C8 used for crystallization were enzymatically removed.…”

Section: Resultsmentioning

confidence: 99%

Structure of Human C8 Protein Provides Mechanistic Insight into Membrane Pore Formation by Complement

Lovelace

Cooper

Sodetz

et al. 2011

Journal of Biological Chemistry

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C8 is one of five complement proteins that assemble on bacterial membranes to form the lethal pore-like "membrane attack complex" (MAC) of complement. The MAC consists of one C5b, C6, C7, and C8 and 12-18 molecules of C9. C8 is composed of three genetically distinct subunits, C8␣, C8␤, and C8␥. The C6, C7, C8␣, C8␤, and C9 proteins are homologous and together comprise the MAC family of proteins. All contain N-and C-terminal modules and a central 40-kDa membrane attack complex perforin (MACPF) domain that has a key role in forming the MAC pore. Here, we report the 2.5 Å resolution crystal structure of human C8 purified from blood. This is the first structure of a MAC family member and of a human MACPF-containing protein. The structure shows the modules in C8␣ and C8␤ are located on the periphery of C8 and not likely to interact with the target membrane. The C8␥ subunit, a member of the lipocalin family of proteins that bind and transport small lipophilic molecules, shows no occupancy of its putative ligand-binding site. C8␣ and C8␤ are related by a rotation of ϳ22°with only a small translational component along the rotation axis. Evolutionary arguments suggest the geometry of binding between these two subunits is similar to the arrangement of C9 molecules within the MAC pore. This leads to a model of the MAC that explains how C8-C9 and C9-C9 interactions could facilitate refolding and insertion of putative MACPF transmembrane ␤-hairpins to form a circular pore. Assembly of the "membrane attack complex" (MAC)3 of complement on the surface of Gram-negative bacteria and other pathogenic organisms involves the sequential interaction of complement proteins C5b, C6, C7, C8, and C9 (1-3). Association of the first four components produces a membranebound tetrameric C5b-8 complex, which then initiates the recruitment and sequential binding of 12-18 C9 molecules to form a cylindrical transmembrane pore (supplemental Fig. S1). Pore formation leads to loss of membrane integrity and lysis of the cell under attack.The sequence of interactions leading to MAC formation is well defined; however, the mechanism by which the MAC disrupts membrane organization is poorly understood. C6, C7, the C8␣ and C8␤ subunits, and C9 are homologous and together comprise the "MAC family" of proteins (4, 5). Until now, structures have not been determined for any of these proteins. All contain N-and C-terminal modules and a central 40-kDa "membrane attack complex/perforin" (MACPF) domain. The MACPF domain was named as such because of sequence similarity between the MAC family proteins and perforin. The modules range in number from three to eight; all are small domains of 40 -60 amino acids that contain multiple disulfide bonds (supplemental Fig. S2). One type of module, thrombospondin type 1 (TSP1), contains several mannosylated tryptophans (6).Several hundred MACPF-containing proteins have been identified; however, functions are known for only a few. MACPF proteins exhibit limited sequence similarity, but all contain the MACPF signature motif ((Y/...

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“…All cysteine residues were searched as carboxamidomethycystein (+57.02 Da). Homemade Buildsummary software was used to delete the redundant data as previously reported (Tang et al, 2007). Additionally, to estimate the rate of incorrect identifications (false positives), all the filtered spectra were subjected to database searching against a composite database containing human protein sequences in both the forward (correct) and reverse (incorrect) orientation.…”

Section: Methodsmentioning

confidence: 99%