Movatterモバイル変換

[0]ホーム
URL:

Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
Thehttps:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

NIH NLM Logo
Log inShow account info
Access keysNCBI HomepageMyNCBI HomepageMain ContentMain Navigation
pubmed logo
Advanced Clipboard
User Guide

Full text links

Atypon full text link Atypon Free PMC article
Full text links

Actions

Share

.2010 Jul 20;107(29):12883-8.
doi: 10.1073/pnas.1001085107. Epub 2010 Jun 21.

Endocytosis-like protein uptake in the bacterium Gemmata obscuriglobus

Affiliations

Endocytosis-like protein uptake in the bacterium Gemmata obscuriglobus

Thierry G A Lonhienne et al. Proc Natl Acad Sci U S A..

Abstract

Endocytosis is a process by which extracellular material such as macromolecules can be incorporated into cells via a membrane-trafficking system. Although universal among eukaryotes, endocytosis has not been identified in Bacteria or Archaea. However, intracellular membranes are known to compartmentalize cells of bacteria in the phylum Planctomycetes, suggesting the potential for endocytosis and membrane trafficking in members of this phylum. Here we show that cells of the planctomycete Gemmata obscuriglobus have the ability to uptake proteins present in the external milieu in an energy-dependent process analogous to eukaryotic endocytosis, and that internalized proteins are associated with vesicle membranes. Occurrence of such ability in a bacterium is consistent with autogenous evolution of endocytosis and the endomembrane system in an ancestral noneukaryote cell.

PubMed Disclaimer

Conflict of interest statement

The authors declare no conflict of interest.

Figures

Fig. 1.
Fig. 1.
G. obscuriglobus cells uptake proteins but not DNA in an energy-dependent process. (A) Bright field microscopy and CLSM images ofG. obscuriglobus cells incubated with GFP (10 μg/mL) for 60 min.Left andRight microscopy images correspond to bright field and GFP fluorescence, respectively. Because of optical sectioning effects, only some of the cells in the clump positive for GFP signal can be visualized. (Scale bar, 4 μm.) (B) Time-course experiment with GFP alone (filled circles), with GFP and 1 mM sodium azide (open circles), or with GFP and both 1 mM sodium azide and 5 mM ATP (filled triangles). After washing ofG. obscuriglobus cells, GFP fluorescence was quantitatively analyzed by microplate reader assay. Error bars represent the SD from three experimental replicates. (C) Western blot analysis showing uptake of full-length GFP (≈27 kDa) byG. obscuriglobus. Cells were incubated with GFP for 1 h. After washing, cell lysates were electrophoretically separated on an SDS gel (Left), then blotted and probed with an anti-GFP antibody (Right). Lane 1,E. coli DH5α incubated with GFP; lane 2,G. obscuriglobus incubated without GFP; lane 3,G. obscuriglobus incubated with GFP. (D) Competition uptake experiment with GFP, together with either BSA or DNA.G. obscuriglobus cells were incubated for 1 h with 10 μg/mL of GFP alone, with GFP and BSA (100 μg/mL), or with GFP and plasmid DNA (100 μg/mL). Cells were then washed and analyzed for fluorescence emission by microplate reader assay. Error bars represent the SD from three experimental replicates.
Fig. 2.
Fig. 2.
Internalized proteins inG. obscuriglobus cells are localized to paryphoplasm. (A)G. obscuriglobus cells were incubated with GFP and then stained with DAPI and SynaptoRed. A GFP-containing region is seen in the cytoplasm bounded by the cytoplasmic membrane as defined by the SynaptoRed staining and is separated from the nuclear body (DAPI staining). (B) TEM image of a section of high-pressure frozen cryosubstituted cells ofG. obscuriglobus, immunogold-labeled to detect GFP via anti-GFP antibody and secondary antibody conjugated with 10 nm colloidal gold. Gold particles (short arrows) labeling internalized GFP are only associated with paryphoplasm (P). Gold particles were excluded from both the double membrane-bounded nucleoid and the riboplasm. The riboplasm (R), fibrillar nucleoid (N), and intracytoplasmic membrane (ICM) are indicated. (Scale bar, 500 nm.) (C) Diagram representing the functional compartmentalization ofG. obscuriglobus. N, nucleoid; NE, nuclear envelope; ICM, intracytoplasmic membrane; R, riboplasm; CM, cytoplasmic membrane; CW, cell wall.
Fig. 3.
Fig. 3.
TEM of high-pressure frozen cryosubstitutedG. obscuriglobus cells showing association of GFP and MC protein with membranes and vesicles. (A) Cells were immunogold-labeled to detect GFP, which is seen only in the paryphoplasm.A1 and the enlargements of boxed areas 2–4 show association of several gold particles with the same membrane. Arrowheads, membranes; V, vesicles. (Scale bars, 1 μm inA1, 50 nm inA2–A4.) (B) A section showing infolding of cytoplasmic membrane (arrowheads) associated with GFP (gold particles). (Scale bar, 50 nm.) (C) Cells were incubated with HRP-conjugated rabbit anti-mouse antibody. Peroxidase activity was detected via cytochemistry using diaminobenzidine substrate, yielding an electron-dense product. An example is shown of a region where the cytoplasmic membrane (CM) seems to have invaginated (arrowheads) together with HRP-conjugated antibody, as indicated by the dense reaction product. (Scale bar, 50 nm.) (D)G. obscuriglobus cells preincubated with GFP were immunogold-labeled to detect MC protein.D1 and enlargements of boxed areas 2 and 3 show association of several gold particles with membrane vesicle. Arrowheads, membranes; V, vesicles. (Scale bars, 500 nm inD1, 200 nm inD2, 50 nm inD3.)
Fig. 4.
Fig. 4.
Colocalization of GFP and MC proteins to the same density gradient membrane fractions of lysedG. obscurigobus cells. These cells had been incubated with GFP. (A andB) Immunogold labeling of the material obtained from fraction 9 of the 40–70% sucrose density gradient centrifugation (Fig. S9) showing cofractionation of GFP and MC protein and association of both proteins with membranes. (A) Fraction 9 was immunolabeled with anti-GFP antibody and secondary anti-mouse-10 nm gold antibody to visualize GFP. Gold particles (g) indicate that GFP is associated with broken membranes (Upper) and the vesicle membrane (Lower) rather than the vesicle interior. Arrowheads, membranes; V, vesicle. (Scale bar, 200 nm.) (B) Fraction 9 was immunolabeled with anti-gp4978 antibody and protein A-15 nm gold to visualize MC-like gp4978 protein. Arrowheads (membranes) and gold particles (g) indicate that MC protein is associated with broken membranes. (Scale bar, 200 nm.)
See this image and copyright information in PMC

Comment in

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. de Duve C. The origin of eukaryotes: A reappraisal. Nat Rev Genet. 2007;8:395–403. - PubMed
    1. Doherty GJ, McMahon HT. Mechanisms of endocytosis. Annu Rev Biochem. 2009;78:857–902. - PubMed
    1. Dacks JB, Poon PP, Field MC. Phylogeny of endocytic components yields insight into the process of nonendosymbiotic organelle evolution. Proc Natl Acad Sci USA. 2008;105:588–593. - PMC - PubMed
    1. Dacks JB, Field MC. Evolution of the eukaryotic membrane-trafficking system: Origin, tempo and mode. J Cell Sci. 2007;120:2977–2985. - PubMed
    1. Lindsay MR, et al. Cell compartmentalisation in planctomycetes: Novel types of structural organisation for the bacterial cell. Arch Microbiol. 2001;175:413–429. - PubMed

Publication types

MeSH terms

Substances

Grants and funding

LinkOut - more resources

Full text links
Atypon full text link Atypon Free PMC article
Cite
Send To

NCBI Literature Resources

MeSHPMCBookshelfDisclaimer

The PubMed wordmark and PubMed logo are registered trademarks of the U.S. Department of Health and Human Services (HHS). Unauthorized use of these marks is strictly prohibited.

[8]ページ先頭
©2009-2025 Movatter.jp