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    Home > Biochemistry News > Biotechnology News > A protein facility user reveals the unique composition and structure of the mammalian radiation axis head complex

    A protein facility user reveals the unique composition and structure of the mammalian radiation axis head complex

    • Last Update: 2022-01-07
    • Source: Internet
    • Author: User
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    On January 26, 2021, the international academic journal PNAS published the latest collaborative research between the team of Cong Yao and the team of Zhu Xueliang, the National Protein Science Research (Shanghai) Facility User, Chinese Academy of Sciences Center for Excellence in Molecular Cell Science (Institute of Biochemistry and Cell Biology) The paper "Distinct architecture and composition of mouse axonemal radial spoke head revealed by cryo-EM"

    .
    The study comprehensively applied cryo-electron microscopy, cell biology and biochemical methods to analyze the 3.
    2 angstrom-resolution cryo-electron microscopy structure of the rodent radiation shaft head complex for the first time, revealing the differences in the evolution of higher animal radiation shaft complexes.
    And the structural mechanism of the radiation axis in the process of cilia swinging

    .


    Radial spoke (RS) is a super-macromolecular complex formed by more than twenty subunits extending from axon-filament two-linked microtubules

    .
    In the motor cilia or flagella, the RS head contacts the central pair (CP) accessory structure, which transmits mechanical and chemical signals from the central pair to the axon dynein arm, thereby coordinating the movement of the cilia

    .
    Although the RS component has been preserved in evolution, the morphology of the RS head complex is very different between protozoa and metazoa

    .
    Moreover, gene mutations in the head component of RS are closely related to primary ciliary dyskinesia (PCD), which manifests as recurrent bronchitis, infertility, and visceral inversion

    .
    Previous research on RS mainly applied cryo-electron tomography (cryo-ET) technology, which had a low resolution and mainly focused on lower organisms

    .
    Recent progress has been made in the study of the RS structure of the protist Chlamydomonas, but so far there is no high-resolution RS structure of higher animals, which restricts the understanding of the molecular mechanism of ciliary swing

    .


    The researchers detected the cilia location of the RS homologous protein in primary cultured mouse cells, combined with in vitro biochemical experiments, found that the mouse RS head complex is composed of Rsph1, Rsph3b, Rsph4a and Rsph9 subunits, but lacks Rsph6a and Rsph6a.
    Rsph10b (its ortholog is present in the RS head of protozoa) is very different from the composition in Chlamydomonas, and has tissue-specific expression (Figure A)

    .
    It shows that in the process of evolution, organisms have produced different RSs in order to adapt to the sports environment under different conditions, and then produced different cilia swinging methods

    .


    The researchers analyzed the cryo-EM structure of the mouse-derived RS head complex with a resolution of 3.
    2 angstroms and performed de novo modeling.
    This is the first higher animal RS head complex atomic structure (Figure BC)

    .
    The structure is a two-fold symmetrical "brake pad"-like structure, in which Rsph4a and Rsph9 form a compact "body" with two twisted long arms formed by Rsph1, and may be connected to the stem of RS through the back of Rsph3b.
    Department

    .
    It was found that a variety of PCD pathogenic mutations occur at sites that may be critical to the assembly or stability of the complex (Figure D), which may cause abnormal assembly of the head of the radiation axis, which in turn causes abnormal cilia swinging

    .
    In addition, the modeling analysis combined with multi-scale structural information shows that the RS head can either rigidly contact the periodic protrusions of the CP through the toothed Rsph4a region or elastically through its extended arms (Figure E), thereby optimizing the RS- The interaction of CP and mechanical signal transduction, and then precisely control the swing of cilia

    .


    This study is the first to analyze the high-resolution cryo-EM structure of the RS head complex, which plays a key role in the movement of higher animal cilia, and clarifies the interaction between the radiation axis and the central microtubule accessory structure, and is used to study the molecular mechanism of cilia swing It provides an important structural basis and reveals the possible causes of PCD caused by related gene mutations

    .


    The National Protein Science Research (Shanghai) Facility Electron Microscopy Analysis System provided important technical support for the screening of tomography data in the early stage of the project, and provided important technical support and machine-time support for the collection of massive single-particle electron microscopy data and three-dimensional reconstruction calculations; The mass spectrometry system provided important help for this result through the analysis of protein complex composition and cross-linked mass spectrometry; the database and computational analysis system provided a stable computing platform and environment for the research, and ensured the rapid output of massive data calculations

    .



    (A) Immunofluorescence of mEPC shows that Rsph1, Rsph3b, Rsph4a, Rsph9 and Rsph10b (but not Rsph6a) are located in ciliated axons
    .
    (BC) 3.
    2 Angstrom resolution cryo-electron microscope structure of RS head complex and its atomic model

    .
    (D) The mutations known to cause PCD are mapped to the atomic model

    .
    The green balls represent mutations associated with PCD

    .
    (E) Two possible modes of interaction between the RS head and the central microtubule accessory structure

    .

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