295 research outputs found
SMYD3-mediated lysine methylation in the PH domain is critical for activation of AKT1
No full textAKT1 is a cytosolic serine/threonine kinase that is overexpressed in various types of cancer and has a central role in human tumorigenesis. Although it is known that AKT1 is post-translationally modified in various ways including phosphorylation and ubiquitination, methylation has not been reported so far. Here we demonstrate that the protein lysine methyltrasnferase SMYD3 methylates lysine 14 in the PH domain of AKT1 both in vitro and in vivo. Lysine 14-substituted AKT1 shows significantly lower levels of phosphorylation at threonine 308 than wild-type AKT1, and knockdown of SMYD3 as well as treatment with a SMYD3 inhibitor significantly attenuates this phosphorylation in cancer cells. Furthermore, substitution of lysine 14 diminishes the plasma membrane accumulation of AKT1, and cancer cells overexpressing lysine 14-substiuted AKT1 shows lower growth rate than those overexpressing wild-type AKT1. These results imply that SMYD3-mediated methylation of AKT1 at lysine 14 is essential for AKT1 activation and that SMYD3-mediated AKT1 methylation appears to be a good target for development of anti-cancer therapy
Abstract 342: WHSC1L1-mediated EGFR mono-methylation enhances the cytoplasmic and nuclear oncogenic activity of EGFR in head and neck cancer
No full textAbstract
While multiple post-translational modifications have been reported to regulate the function of epidermal growth factor receptor (EGFR), the effect of protein methylation on its function has not been well characterized. In this study, we show that WHSC1L1 mono-methylates lysine 721 in the tyrosine kinase domain of EGFR, and that this methylation leads to enhanced activation of its downstream ERK cascade without EGF stimulation. We also show that EGFR K721 mono-methylation not only affects the function of cytoplasmic EGFR, but also that of nuclear EGFR. WHSC1L1-mediated methylation of EGFR in the nucleus enhanced its interaction with PCNA in squamous cell carcinoma of the head and neck (SCCHN) cells and resulted in enhanced DNA synthesis and cell cycle progression. Overall, our study demonstrates the multifaceted oncogenic function of the protein lysine methyltransferase WHSC1L1 in SCCHN, which is mediated through direct non-histone methylation of the EGFR protein with effects both in its cytoplasmic and nuclear functions.
Citation Format: Vassiliki Saloura, Theodore Vougiouklakis, Makda Zewde, Xiaolan Deng, Kazuma Kiyotani, Jae-Hyun Park, Yo Matsuo, Mark Lingen, Naoshi Dohmae, Takehiro Suzuki, Ryuji Hamamoto, Yusuke Nakamura. WHSC1L1-mediated EGFR mono-methylation enhances the cytoplasmic and nuclear oncogenic activity of EGFR in head and neck cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 342. doi:10.1158/1538-7445.AM2017-342</jats:p
The COP9 complex is conserved between plants and mammals and is related to the 26S proteasome regulatory complex
No full textThe COP9 complex, genetically identified in Arabidopsis as a repressor of photomorphogenesis, is composed of multiple subunits including COP9, FUS6 (also known as COP11) and the Arabidopsis JAB1 homolog 1 (AJH1) ([1-3]; unpublished observations). We have previously demonstrated the existence of the mammalian counterpart of the COP9 complex and purified the complex by conventional biochemical and immunoaffinity procedures [4]. Here, we report the molecular identities of all eight subunits of the mammalian COP9 complex. We show that the COP9 complex is highly conserved between mammals and higher plants, and probably among most multicellular eukaryotes. It is not present in the single-cell eukaryote Saccharomyces cerevisiae, however. All of the subunits of the COP9 complex contain structural features that are also present in the components of the proteasome regulatory complex and the translation initiation factor eIF3 complex. Six subunits of the COP9 complex have overall similarity with six distinct non-ATPase regulatory subunits of the 26S proteasome, suggesting that the COP9 complex and the proteasome regulatory complex are closely related in their evolutionary origin. Subunits of the COP9 complex include regulators of the Jun N-terminal kinase (JNK) and c-Jun, a nuclear hormone receptor binding protein and a cell-cycle regulator. This suggests that the COP9 complex is an important cellular regulator modulating multiple signaling pathway
Remodeling of the postsynaptic proteome in male mice and marmosets during synapse development
No full textAbstract
Postsynaptic proteins play crucial roles in synaptic function and plasticity. During brain development, alterations in synaptic number, shape, and stability occur, known as synapse maturation. However, the postsynaptic protein composition changes during development are not fully understood. Here, we show the trajectory of the postsynaptic proteome in developing male mice and common marmosets. Proteomic analysis of mice at 2, 3, 6, and 12 weeks of age shows that proteins involved in synaptogenesis are differentially expressed during this period. Analysis of published transcriptome datasets shows that the changes in postsynaptic protein composition in the mouse brain after 2 weeks of age correlate with gene expression changes. Proteomic analysis of marmosets at 0, 2, 3, 6, and 24 months of age show that the changes in the marmoset brain can be categorized into two parts: the first 2 months and after that. The changes observed in the first 2 months are similar to those in the mouse brain between 2 and 12 weeks of age. The changes observed in marmoset after 2 months old include differential expression of synaptogenesis-related molecules, which hardly overlap with that in mice. Our results provide a comprehensive proteomic resource that underlies developmental synapse maturation in rodents and primates
Real-Time Control of Nanoscale Protein Assembly for Further Crystallization Using a Solution Circulating Nanoaggregation Control Apparatus
No full textExamination of an absolute quantity of less than a hundred nanograms of proteins by amino acid analysis
No full textAutomated Protein Hydrolysis Delivering Sample to a Solid Acid Catalyst for Amino Acid Analysis
No full textCharacterization of post‐translational modifications on lysine 9 of histone H3 variants in mouse testis using matrix‐assisted laser desorption/ionization in‐source decay
No full textReal-Time Control of Nanoscale Protein Assembly for Further Crystallization Using a Solution Circulating Nanoaggregation Control Apparatus
No full textIn this study, a solution circulating apparatus was employed
to
control the aggregation of protein molecules in situ. The association
state of protein molecules was controlled by injecting protein solutions,
buffers, and solutes into the device while monitoring the heterodispersity
of the molecular nanoaggregates by an apparatus of high-performance
heterodyne-based dynamic light scattering. Further, a highly ordered
association of protein nanoaggregates was induced that lead to the
generation and growth of protein crystals suitable for structural
analysis by the X-ray protein crystallography. We could also dissociate
protein molecules that had already formed aggregates into mono- or
bimodal states and reassociate them to form protein crystals, which
contribute to minimizing the amount of protein required for crystallization.
Our nanotechnological approach allowed us to carefully monitor and
control crystal growth with a focus on the association state of the
target proteins. The high-performance DLS device monitors the in situ
association process of the target proteins allowing a dynamic response
to modify the solution properties and leading to a deeper understanding
of protein crystallization and achieving rational control of the processes
Global mapping of post-translational modifications on histone H3 variants in mouse testes
No full textMass spectrometry (MS)-based characterization is important in proteomic research for verification of structural features and functional understanding of gene expression. Post-translational modifications (PTMs) such as methylation and acetylation have been reported to be associated with chromatin remodeling during spermatogenesis. Although antibody- and MS-based approaches have been applied for characterization of PTMs on H3 variants during spermatogenesis, variant-specific PTMs are still underexplored. We identified several lysine modifications in H3 variants, including testis-specific histone H3 (H3t), through their successful separation with MS-based strategy, based on differences in masses, retention times, and presence of immonium ions. Besides methylation and acetylation, we detected formylation as a novel PTM on H3 variants in mouse testes. These patterns were also observed in H3t. Our data provide high-throughput structural information about PTMs on H3 variants in mouse testes and show possible applications of this strategy in future proteomic studies on histone PTMs
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