Biochemistry (Moscow)

, Volume 78, Issue 5, pp 445–454 | Cite as

Topology of mRNA chain in isolated eukaryotic double-row polyribosomes

  • Zh. A. Afonina
  • A. G. Myasnikov
  • N. F. Khabibullina
  • A. Yu. Belorusova
  • J. -F. Menetret
  • V. D. Vasiliev
  • B. P. Klaholz
  • V. A. Shirokov
  • A. S. Spirin
Article

Abstract

In the process of protein synthesis, the translating ribosomes of eukaryotic cells form polyribosomes that are found to be multiplex functional complexes possessing elements of ordered spatial organization. As revealed by a number of electron microscopy studies, the predominant visible configurations of the eukaryotic polyribosomes are circles (circular polyribosomes) and two-stranded formations (so-called double-row polyribosomes). The “long” (i.e. heavy loaded) polyribosomes are usually represented by double-row structures, which can be interpreted as either topologically circular (“col-lapsed rings”), or topologically linear (zigzags or helices). In the present work we have analyzed the mRNA path within the eukaryotic polyribosomes, isolated from a wheat germ cell-free translation system, by integrating two approaches: the visualization of mRNA ends in polyribosomes by marking them with gold nanoparticles (3′-end) and initiating 40S subunits (5′-end), as well as by the cryoelectron tomography. Examination of the location of the mRNA markers in polyribosomes and mutual orientation of ribosomes in them has shown that the double-row polyribosomes of the same sample can have both circular and linear arrangements of their mRNA.

Key words

eukaryotic polyribosomes mRNA circular translation cryoelectron tomography 

Abbreviations

CECF

continuous exchange cell-free system

CET

cryoelectron tomography

eIF

eukaryotic initiation factor

EM

electron microscopy

FITC

fluorescein isothiocyanate

FTSC

fluorescein-5-thiosemicarbazide

“MAFITC 10 nm gold”

anti-fluorescein isothiocyanate antibody conjugated with 10 nm gold particles

PABP

poly(A)-binding protein

UTR

untranslated region of mRNA

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Copyright information

© Pleiades Publishing, Ltd. 2013

Authors and Affiliations

  • Zh. A. Afonina
    • 1
  • A. G. Myasnikov
    • 2
    • 3
    • 4
    • 5
  • N. F. Khabibullina
    • 1
  • A. Yu. Belorusova
    • 1
  • J. -F. Menetret
    • 2
    • 3
    • 4
    • 5
  • V. D. Vasiliev
    • 1
  • B. P. Klaholz
    • 2
    • 3
    • 4
    • 5
  • V. A. Shirokov
    • 1
  • A. S. Spirin
    • 1
  1. 1.Institute of Protein ResearchRussian Academy of SciencesPushchino, Moscow RegionRussia
  2. 2.Department of Integrated Structural BiologyInstitute of Genetics and Molecular and Cellular Biology (IGBMC)IllkirchFrance
  3. 3.INSERM, U596IllkirchFrance
  4. 4.CNRS, UMR7104IllkirchFrance
  5. 5.Universite de StrasbourgStrasbourgFrance
  6. 6.Research and Education Center “Bionanophysics”Moscow Institute of Physics and TechnologyDolgoprudnyiRussia
  7. 7.Institute of Genetics and of Molecular and Cellular Biology (IGBMC)IllkirchFrance

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