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De novo protein identification in mammalian sperm using in situ cryoelectron tomography and AlphaFold2 docking.
Chen, Zhen; Shiozaki, Momoko; Haas, Kelsey M; Skinner, Will M; Zhao, Shumei; Guo, Caiying; Polacco, Benjamin J; Yu, Zhiheng; Krogan, Nevan J; Lishko, Polina V; Kaake, Robyn M; Vale, Ronald D; Agard, David A.
Afiliação
  • Chen Z; Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, CA, USA; Department of Cellular and Molecular Pharmacology, University of California, San Francisco, San Francisco, CA, USA. Electronic address: zhen.chen@ucsf.edu.
  • Shiozaki M; Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, VA, USA.
  • Haas KM; Department of Cellular and Molecular Pharmacology, University of California, San Francisco, San Francisco, CA, USA; J. David Gladstone Institutes, San Francisco, CA, USA; Quantitative Biosciences Institute (QBI), University of California, San Francisco, San Francisco, CA, USA.
  • Skinner WM; Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA, USA.
  • Zhao S; Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, VA, USA.
  • Guo C; Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, VA, USA.
  • Polacco BJ; Department of Cellular and Molecular Pharmacology, University of California, San Francisco, San Francisco, CA, USA; Quantitative Biosciences Institute (QBI), University of California, San Francisco, San Francisco, CA, USA.
  • Yu Z; Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, VA, USA.
  • Krogan NJ; Department of Cellular and Molecular Pharmacology, University of California, San Francisco, San Francisco, CA, USA; J. David Gladstone Institutes, San Francisco, CA, USA; Quantitative Biosciences Institute (QBI), University of California, San Francisco, San Francisco, CA, USA.
  • Lishko PV; Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA, USA.
  • Kaake RM; Department of Cellular and Molecular Pharmacology, University of California, San Francisco, San Francisco, CA, USA; J. David Gladstone Institutes, San Francisco, CA, USA; Quantitative Biosciences Institute (QBI), University of California, San Francisco, San Francisco, CA, USA.
  • Vale RD; Department of Cellular and Molecular Pharmacology, University of California, San Francisco, San Francisco, CA, USA; Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, VA, USA. Electronic address: valer@janelia.hhmi.org.
  • Agard DA; Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, CA, USA; Quantitative Biosciences Institute (QBI), University of California, San Francisco, San Francisco, CA, USA. Electronic address: david@agard.ucsf.edu.
Cell ; 186(23): 5041-5053.e19, 2023 11 09.
Article em En | MEDLINE | ID: mdl-37865089
ABSTRACT
To understand the molecular mechanisms of cellular pathways, contemporary workflows typically require multiple techniques to identify proteins, track their localization, and determine their structures in vitro. Here, we combined cellular cryoelectron tomography (cryo-ET) and AlphaFold2 modeling to address these questions and understand how mammalian sperm are built in situ. Our cellular cryo-ET and subtomogram averaging provided 6.0-Å reconstructions of axonemal microtubule structures. The well-resolved tertiary structures allowed us to unbiasedly match sperm-specific densities with 21,615 AlphaFold2-predicted protein models of the mouse proteome. We identified Tektin 5, CCDC105, and SPACA9 as novel microtubule-associated proteins. These proteins form an extensive interaction network crosslinking the lumen of axonemal doublet microtubules, suggesting their roles in modulating the mechanical properties of the filaments. Indeed, Tekt5 -/- sperm possess more deformed flagella with 180° bends. Together, our studies presented a cellular visual proteomics workflow and shed light on the in vivo functions of Tektin 5.
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Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Espermatozoides / Proteoma Limite: Animals Idioma: En Revista: Cell Ano de publicação: 2023 Tipo de documento: Article
Texto completo
Imprimir
XML
PubMed Links
Buscar no Google

Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Espermatozoides / Proteoma Limite: Animals Idioma: En Revista: Cell Ano de publicação: 2023 Tipo de documento: Article