Expression of the Clustered NeuAcα2–3Galβ O-Glycan Determines the Cell Differentiation State of the Cells

Higashi, Kei; Asano, Koji; Yagi, Masahide; Yamada, Keita; Arakawa, Tatsuhiko; Ehashi, Tomo; Mori, Takeo; Sumida, Kayo; Kushida, Masahiko; Ando, Satoshi; Kinoshita, Mitsuhiro; Kakehi, Kazuaki; Tachibana, Taro; Saito, Koichi

doi:10.1074/jbc.m114.550848

Cited by 14 publications

(21 citation statements)

References 37 publications

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“…(16) Recently, we have demonstrated that a mouse monoclonal antibody recognizing the clustered NeuAca2-3Galb O-Glycan marks undifferentiated cells such as hESCs and cancer stem-like cells. (4) In the present study, monoclonal antibodies against undifferentiated hESCs were generated, and MAb2 was selected for further characterization. Biochemical analyses revealed that MAb2 antigen is distributed on the cell surface, and MAb2 detected three diffused bands with molecular mass between 30 and 60 kDa in the lysates from hESCs and human lung cancer cells.…”

Section: Discussionmentioning

confidence: 99%

“…After incubation with primary antibodies, the proteins of interest on immunoblots were detected using a NBT/BCIP (14) or an enhanced chemiluminescence detection system. (4) …”

Section: Immunoprecipitation and Microsequencingmentioning

confidence: 97%

“…Monoclonal antibodies were purified from the desired hybridoma supernatants. (4) Flow cytometry hESCs were dissociated to single cells, incubated with antibody at 4°C, and counted and sorted with FACSAria (Becton Dickinson, Franklin Lakes, NJ).…”

Section: Generation Of Monoclonal Antibodiesmentioning

confidence: 99%

“…Desired bands were excised, in-gel digested with trypsin, and solvent extracted, and the resulting MS/MS spectra were searched using the MASCOT search engine. (4) Western blot analysis Whole cell extracts were resolved by 10% SDS-PAGE, transferred to a PVDF membrane, and blocked with 5% skim milk for 30 min at room temperature. After incubation with primary antibodies, the proteins of interest on immunoblots were detected using a NBT/BCIP (14) or an enhanced chemiluminescence detection system.…”

Section: Immunoprecipitation and Microsequencingmentioning

confidence: 99%

“…We have recently demonstrated that the clustered NeuAca2-3Galb O-Glycan on the plasma membrane is specifically expressed in undifferentiated cells such as hESCs and cancer stem-like cells, and plays a possible role in the undifferentiated cells. (4) In addition, a recent study suggested the involvement of the terminal sialic acid in the maintenance of embryonic stem cell pluripotency and undifferentiation. (5) Insights into the developmental roles of cell surface molecules ultimately impact the understanding of mechanisms governing the pluripotency and self-renewal capacity of hESCs.…”

Section: Introductionmentioning

confidence: 99%

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A Novel Marker for Undifferentiated Human Embryonic Stem Cells

Higashi

Yagi²,

Arakawa³

et al. 2015

Monoclonal Antibodies in Immunodiagnosis and Immunotherapy

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Human embryonic stem cells (hESCs) are pluripotent stem cells from early embryos, and their self-renewal capacity depends on the sustained expression of hESC-specific molecules and the suppressed expression of differentiation-associated genes. To discover novel molecules expressed on hESCs, we generated a panel of monoclonal antibodies against undifferentiated hESCs. The antigen recognized by MAb2 is expressed on the cell surface of undifferentiated hESCs; three diffused bands with molecular mass between 30 and 60 kDa in the lysates of hESCs were diminished during hESC differentiation into neural cells. The expression of MAb2 antigen was also observed on the plasma membrane of lung cancer cells, and MAb2 detected 55, 50, and 35 kDa protein bands in the cell lysates. Immunoprecipitation followed by proteomics analyses identified CD147/basigin as a MAb2 antigen. Finally, the positive expression of CD147/basigin protein in undifferentiated hESCs was confirmed. These results suggested that CD147/basigin could be another undifferentiated hESC marker.

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Section: Discussionmentioning

confidence: 99%

“…After incubation with primary antibodies, the proteins of interest on immunoblots were detected using a NBT/BCIP (14) or an enhanced chemiluminescence detection system. (4) …”

Section: Immunoprecipitation and Microsequencingmentioning

confidence: 97%

Section: Generation Of Monoclonal Antibodiesmentioning

confidence: 99%

Section: Immunoprecipitation and Microsequencingmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 3 more Smart Citations

A Novel Marker for Undifferentiated Human Embryonic Stem Cells

Higashi

Yagi²,

Arakawa³

et al. 2015

Monoclonal Antibodies in Immunodiagnosis and Immunotherapy

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Chemical Remodeling of Cell‐Surface Sialic Acids through a Palladium‐Triggered Bioorthogonal Elimination Reaction

Wang

Cheng

et al. 2015

Angewandte Chemie

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We herein report achemical decaging strategy for the in situ generation of neuramic acid (Neu), au nique type of sialic acid, on live cells by the use of ap alladium-mediated bioorthogonal elimination reaction. Palladium nanoparticles (Pd NPs) were found to be ah ighly efficient and biocompatible depropargylation catalyst for the direct conversion of metabolically incorporated N-(propargyloxycarbonyl)neuramic acid (Neu5Proc) into Neu on cell-surface glycans.T his conversion chemically mimics the enzymatic de-N-acetylation of N-acetylneuramic acid (Neu5Ac), ap roposed mechanism for the natural occurrence of Neu on cell-surface glycans.The bioorthogonal elimination was also exploited for the manipulation of cell-surface charge by unmasking the free amine at C5 to neutralizethe negatively charged carboxyl group at C1 of sialic acids.

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Chemical Remodeling of Cell‐Surface Sialic Acids through a Palladium‐Triggered Bioorthogonal Elimination Reaction

et al. 2015

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We herein report a chemical decaging strategy for the in situ generation of neuramic acid (Neu), a unique type of sialic acid, on live cells by the use of a palladium-mediated bioorthogonal elimination reaction. Palladium nanoparticles (Pd NPs) were found to be a highly efficient and biocompatible depropargylation catalyst for the direct conversion of metabolically incorporated N-(propargyloxycarbonyl)neuramic acid (Neu5Proc) into Neu on cell-surface glycans. This conversion chemically mimics the enzymatic de-N-acetylation of N-acetylneuramic acid (Neu5Ac), a proposed mechanism for the natural occurrence of Neu on cell-surface glycans. The bioorthogonal elimination was also exploited for the manipulation of cell-surface charge by unmasking the free amine at C5 to neutralize the negatively charged carboxyl group at C1 of sialic acids.

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Expression of the Clustered NeuAcα2–3Galβ O-Glycan Determines the Cell Differentiation State of the Cells

Cited by 14 publications

References 37 publications

A Novel Marker for Undifferentiated Human Embryonic Stem Cells

A Novel Marker for Undifferentiated Human Embryonic Stem Cells

Chemical Remodeling of Cell‐Surface Sialic Acids through a Palladium‐Triggered Bioorthogonal Elimination Reaction

Chemical Remodeling of Cell‐Surface Sialic Acids through a Palladium‐Triggered Bioorthogonal Elimination Reaction

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