A cytoskeletal structure with associated polyribosomes obtained from HeLa cells

Lenk, Robert; Ransom, Laura J.; Kaufmann, Yael; Penman, Sheldon

doi:10.1016/0092-8674(77)90141-6

Cited by 534 publications

(290 citation statements)

References 26 publications

Supporting

Mentioning

270

Contrasting

Unclassified

Order By: Relevance

“…It is estimated that 70 -80% of mRNA in the cell is associated with, but not bound directly to, microfilaments or microtubules (Lenk et al, 1977;Cervera et al, 1981;van Venrooij et al, 1981;Taneja et al, 1992;Bassell, 1993;Bassell et al, 1994). A linker is therefore required for the association of mRNAs to the cytoskeleton.…”

Section: Introductionmentioning

confidence: 99%

Interactions of Elongation Factor 1α with F-Actin and β-Actin mRNA: Implications for Anchoring mRNA in Cell Protrusions

Liu

Grant²,

Persky³

et al. 2002

MBoC

160

159

View full text Add to dashboard Cite

The targeting of mRNA and local protein synthesis is important for the generation and maintenance of cell polarity. As part of the translational machinery as well as an actin/ microtubule-binding protein, elongation factor 1␣ (EF1␣) is a candidate linker between the protein translation apparatus and the cytoskeleton. We demonstrate in this work that EF1␣ colocalizes with ␤-actin mRNA and F-actin in protrusions of chicken embryo fibroblasts and binds directly to F-actin and ␤-actin mRNA simultaneously in vitro in actin cosedimentation and enzyme-linked immunosorbent assays. To investigate the role of EF1␣ in mRNA targeting, we mapped the two actin-binding sites on EF1␣ at high resolution and defined one site at the N-terminal 49 residues of domain I and the other at the C-terminal 54 residues of domain III. In vitro actin-binding assays and localization in vivo of recombinant full-length EF1␣ and its various truncates demonstrated that the C terminus of domain III was the dominant actin-binding site both in vitro and in vivo. We propose that the EF1␣-F-actin complex is the scaffold that is important for ␤-actin mRNA anchoring. Disruption of this complex would lead to delocalization of the mRNA. This hypothesis was tested by using two dominant negative polypeptides: the actin-binding domain III of EF1␣ and the EF1␣-binding site of yeast Bni1p, a protein that inhibits EF1␣ binding to F-actin and also is required for yeast mRNA localization. We demonstrate that either domain III of EF1␣ or the EF1␣-binding site of Bni1p inhibits EF1␣ binding to ␤-actin mRNA in vitro and causes delocalization of ␤-actin mRNA in chicken embryo fibroblasts. Taken together, these results implicate EF1␣ in the anchoring of ␤-actin mRNA to the protrusion in crawling cells.

show abstract

Section: Introductionmentioning

confidence: 99%

Interactions of Elongation Factor 1α with F-Actin and β-Actin mRNA: Implications for Anchoring mRNA in Cell Protrusions

Liu

Grant²,

Persky³

et al. 2002

MBoC

160

159

View full text Add to dashboard Cite

show abstract

“…In contrast, poly(A) appeared to be preferentially conserved in the detergent-insoluble fraction when measured by hybridization with [aH]poly(U). Thus the CF of ascidian follicle cells, like that of a number of cell types (8,20,23), is enriched in poly(A). The CF of follicle cells resembles the cytoskeleton of vertebrate cells seen by high voltage electron microscopy (36) or revealed after extraction with nonionic detergents (3,20,27,29,30).…”

Section: In 5itu Hybridizationmentioning

confidence: 99%

mentioning

confidence: 99%

“…Gentle disruption of eukaryotic cells with nonionic detergents permits concomitant biochemical and morphological analyses of the CF after differential centrifugation is used to separate the cell lysate into detergent soluble and insoluble fractions. An important result of these biochemical studies is that most of the polyribosomal mRNA is found to be associated with the CF (8,20,33).The purpose of the present investigation was to test the possibility of mRNA interaction with the CF by cytological methods, in particular in situ hybridization. In this paper, in situ hybridization with poly(U) or a cloned actin DNA probe and radioactive isotope tracing methods have been used to demonstrate that a significant proportion of the mRNA of follicle ceils is associated with the CF.…”

mentioning

confidence: 99%

“…The CF includes microtubules, microfdaments, intermediate filaments, microtrabeculae (36), centrioles (20), a membrane protein lamina (2), and probably many other cellular components. It can be visualized by immunofluorescence microscopy of intact cells (19,24,25,35 ) or by electron microscopy of detergent-extracted cells (3,20,27,29,30). Gentle disruption of eukaryotic cells with nonionic detergents permits concomitant biochemical and morphological analyses of the CF after differential centrifugation is used to separate the cell lysate into detergent soluble and insoluble fractions.…”

mentioning

confidence: 99%

See 2 more Smart Citations

Messenger RNA in the cytoskeletal framework: analysis by in situ hybridization.

Jeffery¹

1982

The Journal of Cell Biology

View full text Add to dashboard Cite

The possibility of an association of mRNA with the cytoskeletal framework (CF) of ascidian (Styela plicata) follicle cells was examined in this study. The approach was to extract the follicle cells with Triton X-100 and determine whether mRNA persisted in the insoluble residue by two methods, in situ hybridization with poly(U) and actin DNA probes and the incorporation of radioactive isotopes into RNA. Triton X-100 extraction of follicle cells yielded a filamentous CF containing ~70% of the total poly (A) but only 9% of the total lipid, 23% of the total protein, and 28% of the total RNA. In situ hybridization with a poly (U) probe indicated that ~70% of the poly (A) was associated with the CF. In situ hybridization with a cloned actin DNA probe indicated that ~60% of the actin mRNA was associated with the CF. Autoradiography of detergent-extracted follicle cells, which had been labeled with [3H]uridine or [3H]adenosine, indicated that >90% of the newly synthesized poly (A)+RNA was preserved in the CF. Thus more newly synthesized mRNA than steady-state mRNA may be present in the Triton X-100 insoluble fraction. It is concluded that a significant proportion of the mRNA complement of ascidian follicle cells is associated with the CF.The cytoplasm of many eukaryotic cells contains an elaborate network of filaments and associated components which has been termed the cytoskeletal framework (CF) (21). The CF includes microtubules, microfdaments, intermediate filaments, microtrabeculae (36), centrioles (20), a membrane protein lamina (2), and probably many other cellular components. It can be visualized by immunofluorescence microscopy of intact cells (19,24,25,35 ) or by electron microscopy of detergent-extracted cells (3,20,27,29,30). Gentle disruption of eukaryotic cells with nonionic detergents permits concomitant biochemical and morphological analyses of the CF after differential centrifugation is used to separate the cell lysate into detergent soluble and insoluble fractions. An important result of these biochemical studies is that most of the polyribosomal mRNA is found to be associated with the CF (8,20,33).The purpose of the present investigation was to test the possibility of mRNA interaction with the CF by cytological methods, in particular in situ hybridization. In this paper, in situ hybridization with poly(U) or a cloned actin DNA probe and radioactive isotope tracing methods have been used to demonstrate that a significant proportion of the mRNA of follicle ceils is associated with the CF. The follicle cells of ascidian eggs were selected as a model system because they are known to contain a filamentous cytoskeleton (9, 10) and are extremely active in the synthesis and accumulation of THt IOURNA[ OE CELL 81OtOGY • VOLUME 95 OCTOBER 1982 1-7 © The Rockefeller University Press -0021-9525/82/10/1/7 $1.00 poly(A)+RNA during oogenesis and early development (15). The association of mRNA with the CF may be a mechanism for controlling the polarity of mRNA distribution in eukaryotic cells. MATERIALS AND M...

show abstract

Ribosomal Protein L9 is the Product ofGRC5, a Homolog of the Putative Tumor Suppressor QM inS. cerevisiae

Nika¹,

Erickson²,

Hannig³

1997

Yeast

View full text Add to dashboard Cite

Genes encoding members of the highly conserved QM family have been identified in eukaryotic organisms from yeast to man. Results of previous studies have suggested roles for QM in control of cell growth and proliferation, perhaps as a tumor suppressor, and in energy metabolism. We identified recessive lethal alleles of the Saccharomyces cerevisiae QM homolog GRC5 that increased GCN4 expression when present in multiple copies. These alleles encode truncated forms of the yeast QM protein Grc5p. Using a functional epitope‐tagged GRC5 allele, we localized Grc5p to a 60S fraction that contained the large ribosomal subunit. Two‐dimensional gel analysis of highly purified yeast ribosomes indicated that Grc5p corresponds to 60S ribosomal protein L9. This identification is consistent with the predicted physical characteristics of eukaryotic QM proteins, the highly biased codon usage of GRC5, and the presence of putative Rap1p‐binding sites in the 5′ sequences of the yeast GRC5 gene. © 1997 John Wiley & Sons, Ltd.

show abstract

A cytoskeletal structure with associated polyribosomes obtained from HeLa cells

Cited by 534 publications

References 26 publications

Interactions of Elongation Factor 1α with F-Actin and β-Actin mRNA: Implications for Anchoring mRNA in Cell Protrusions

Interactions of Elongation Factor 1α with F-Actin and β-Actin mRNA: Implications for Anchoring mRNA in Cell Protrusions

Messenger RNA in the cytoskeletal framework: analysis by in situ hybridization.

Ribosomal Protein L9 is the Product ofGRC5, a Homolog of the Putative Tumor Suppressor QM inS. cerevisiae

Contact Info

Product

Resources

About