Congenital disorder(s) of glycosylation (CDG) covers a wide range of disorders affecting glycoconjugates. A number of cases due to defects in the biosynthetic pathway of N-glycosylation have been identified in the last 30 years. MS has considerably helped to promote our knowledge of this emerging disease, by characterizing the molecular abnormalities of glycoproteins from the patients. ESI MS of transferrin detects a lack of N-glycans due to defects in the early glycosylation pathway in ER or allows profiling of glycoforms including immature structures derived from defects in the Golgi apparatus. MALDI MS of apolipoprotein C-III is a simple method of elucidating the profiles of mucin-type core 1 O-glycans including site occupancy and glycoforms. A preliminary result of CDG screening in Japan suggested an incidence of approximately one percent in 1,200 Japanese patients with developmental delay of unknown cause.

Ribosome, a ribonucleoprotein complex huge enough to be visible with light microscopy, is essential for protein synthesis and for the survival of almost all cells. The cells consume 70–80% of the total substances and energy used during their multiplication just for ribosome biogenesis. It has currently shown that ribosome biogenesis is closely linked to cell proliferation, cell growth/cell cycle control, aging/stress response, and actions of oncogenes and growth factors. Analysis of human ribosome biosynthesis was hampered by the lack of the technologies analyzing very complicated processes involving hundreds of proteins and RNAs until recent years. Along with the progress of proteomics, the technique for isolating protein complexes, and that for large-scale identification and quantification of proteins have dramatically improved. Identification of components of ribosome synthesis intermediates in human cells and their functional analyses have rapidly progressed, and the biosynthesis pathway of human cells and its regulatory mechanism are becoming clear. In this review, we will outline the current status of the analyses of the ribosome biosynthesis and of its control mechanism in human cells.

Protein function is primarily determined by genomic cording, but further precisely regulated by co- and post-translational modifications. For instance, phosphorylation may be involved in regulation of enzymatic activity, protein interaction with various molecules, cellular localization and so on. Ubiquitination may have significant roles both in protein degradation through proteasome process, and in signal transduction through protein-protein interaction. Therefore, abnormal post-translational modification may disrupt normal cell functions and result in disease states. Actually, phosphorylation of proteins involved in cell growth signaling is upregulated in most cancer cells. Comprehensive proteome analysis based on mass spectrometric technologies are also applicable to detection and identification of post-translational modifications in disease states. Thus, in this review article, I give a brief overview of the recent knowledge about relationship between post-translational modifications and disease states.

I started my research on proteomics of the glomerulus of kidney almost 15 years ago when proteomics was at its initial stage and in a frenzy of anticipation. Through the research I have experienced a lot of frustration as well as pleasures in working with mass spectrometers. In this short review, I would like to summarize history and achievements of our research. This review, however, start with overview and perspective of proteomics field in the sight of a researcher deeply committed to proteomics for long years. It includes ambiguity of protein identification, tremendous heterogeneity of proteome, brief history of proteomics and current challenge confronting proteomics. I then briefly summarize our research including recent works. Finally I would like to depict my personal view on prospects of proteomics.

Aberrant phosphorylation state of proteins is one of the major causes of tumorigenesis and cancer malignancy. Here, we performed a quantitative phosphoproteomic study aimed at revealing the molecular mechanisms underlying the pathology of ovarian clear cell carcinoma (OCCC), a highly malignant tumor among various histological subtypes of epithelial ovarian carcinoma. Comparative phosphoproteomic analysis using OCCC-derived cell lines indicated OCCC-specific downregulation of phosphopeptides derived from five components of SWI/SNF chromatin remodeling complex, including ARID1A, a tumor suppressor with frequent somatic mutations (46–57%) in OCCC tumors and cell lines. Native phosphorylation levels of ARID1A and BRG1, core components of SWI/SNF complex, were also analyzed by multiple reaction monitoring (MRM)-MS analysis, which we developed to perform parallel detection of phosphopeptide and its cognate nonphosphopeptide of these proteins. As a result, we detected significant downregulation of the phosphorylation level of BRG1, whereas diminished phosphopeptide level of ARID1A was likely to reflect decreased ARID1A level. These results indicated that not only decreased protein level of ARID1A but also downregulation of phosphorylation level of BRG1 might be related to the high malignancy of OCCC, which is induced by the impaired chromatin remodeling activity of SWI/SNF complex.

Antibodies are usually developed using purified proteins or synthetic peptides. We have exhaustively generated monoclonal antibodies against various tumor-associated proteins using lung cancer cell lines or tissues as antigens with the random immunization method, and have obtained over 2,000 monoclonal antibodies. This method is expected to generate antibodies against proteins with tumor-specific post-translational modifications that are difficult to obtain by conventional immunization methods. Moreover, random immunization method can select antibodies which react with only tumor cells at the time of screening and is useful to obtain many potential immunostaining antibodies in a short time. Actually, monoclonal antibodies generated by this random immunization method are useful as diagnostic and prognostic markers for lung cancers. In a study to search for the marker which is useful for differential diagnosis and prognosis, the use of the random immunization is expected.