Protein and DNA synthesis demonstrated in cell-free Ehrlichia chaffeensis organisms in axenic medium

ABSTRACT Rickettsiae belong to the Anaplasmataceae family, which includes mostly tick-transmitted pathogens causing human, canine, and ruminant diseases. Biochemical characterization of the pathogens remains a major challenge because of their obligate parasitism. We investigated the use of an axenic medium for growth of two important pathogens—Anaplasma phagocytophilum and Ehrlichia chaffeensis—in host cell-free phagosomes. We recently reported that the axenic medium promotes protein and DNA biosynthesis in host cell-free replicating form of E. chaffeensis, although the bacterial replication is limited. We now tested the hypothesis that growth on axenic medium can be improved if host cell-free rickettsia-containing phagosomes are used. Purification of phagosomes from A. phagocytophilum- and E. chaffeensis-infected host cells was accomplished by density gradient centrifugation combined with magnet-assisted cell sorting. Protein and DNA synthesis was observed for both organisms in cell-free phagosomes with glucose-6-phosphate and/or ATP. The levels of protein and DNA synthesis were the highest for a medium pH of 7. The data demonstrate bacterial DNA and protein synthesis for the first time in host cell-free phagosomes for two rickettsial pathogens. The host cell support-free axenic growth of obligate pathogenic rickettsiae will be critical in advancing research goals in many important tick-borne diseases impacting human and animal health.

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Use of Antibody to aid Visualization of Protein at the Termini of Bacteriophage Ø29 DNA

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s1431927600002385 ◽

1980 ◽

Vol 38 ◽

pp. 540-541

Author(s):

Dwight Anderson ◽

Charlene Peterson ◽

Gursaran Notani ◽

Bernard Reilly

Keyword(s):

Electron Microscopy ◽

Bacillus Subtilis ◽

Dna Synthesis ◽

Restriction Endonuclease ◽

Protein Complex ◽

Protein Product ◽

Viral Dna ◽

Small Proteins ◽

Antibody Labeling ◽

Subtilis Bacteriophage

The protein product of cistron 3 of Bacillus subtilis bacteriophage Ø29 is essential for viral DNA synthesis and is covalently bound to the 5’-termini of the Ø29 DNA. When the DNA-protein complex is cleaved with a restriction endonuclease, the protein is bound to the two terminal fragments. The 28,000 dalton protein can be visualized by electron microscopy as a small dot and often is seen only when two ends are in apposition as in multimers or in glutaraldehyde-fixed aggregates. We sought to improve the visibility of these small proteins by use of antibody labeling.

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Early effects of ultraviolet light on DNA synthesis in human skin in vivo

Archives of Dermatology ◽

10.1001/archderm.100.1.84 ◽

1969 ◽

Vol 100 (1) ◽

pp. 84-89 ◽

Cited By ~ 35

Author(s):

W. L. Epstein

Keyword(s):

Dna Synthesis ◽

Human Skin ◽

Ultraviolet Light ◽

Early Effects

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Protein Synthesis DNA Synthesis and Repair RNA Synthesis Energy-Linked ATPases Synthetases

10.1016/s1874-6047(08)x6008-1 ◽

1974 ◽

Keyword(s):

Protein Synthesis ◽

Dna Synthesis ◽

Rna Synthesis

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The uvsI gene of Aspergillus nidulans required for UV-mutagenesis encodes a homolog to REV3, a subunit of the DNA polymerase ζ of yeast involved in translesion DNA synthesis

FEMS Microbiology Letters ◽

10.1016/s0378-1097(98)00185-2 ◽

1998 ◽

Vol 164 (1) ◽

pp. 13-19 ◽

Cited By ~ 1

Author(s):

K Han

Keyword(s):

Dna Synthesis ◽

Aspergillus Nidulans ◽

Dna Polymerase ◽

Uv Mutagenesis ◽

Translesion Dna Synthesis ◽

A Subunit ◽

Translesion Dna

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Classification of small type B/C follicles as primordial follicles in mature rats

Reproduction ◽

10.1530/reprod/119.1.43 ◽

2000 ◽

pp. 43-48 ◽

Cited By ~ 30

Author(s):

S Meredith ◽

G Dudenhoeffer ◽

K Jackson

Keyword(s):

Dna Synthesis ◽

Granulosa Cells ◽

Single Layer ◽

Reliable Indicator ◽

Type B ◽

Primordial Follicles ◽

Type C

In the present study, follicles were classified according to the morphology of their granulosa cells. Type B follicles contained only flattened granulosa cells; type B/C follicles had a mixture of flattened and cuboidal granulosa cells in a single layer, and type C follicles had a single layer of cuboidal granulosa cells. The primary objectives of the study were to determine whether 5-bromo-2-deoxyuridine incorporation into type B/C follicles was a marker for initiation of growth and how long type B/C follicles could remain at the same stage before transformation to type C follicles. Female Holtzman rats received bromo-deoxyuridine for 7 days. After the infusion (day minipumps were removed = day 0), rats were ovariectomized on days 0 (n = 9), 30 (n = 8), 90 (n = 8) and 150 (n = 9). The numbers of type B, B/C and C follicles within one ovary were determined using modified fractionator counting. Analysis over all times demonstrated that there were more (P < 0.0001) type B/C (941 +/- 61 per ovary) than type C (140 +/- 18 per ovary) or type B (159 +/- 19 per ovary) follicles. The numbers of type B and type C follicles did not differ from each other at any time. Only one of 34 rats evaluated had bromo-deoxyuridine-labelled type B follicles. On day 150, 57% of the bromo-deoxyuridine-labelled type B/C follicles remained from day 0. It is concluded that (1) DNA synthesis in granulosa cells of type B/C follicles is not a reliable indicator of impending growth; and (2) type B and type B/C follicles are both components of the pool of primordial follicles.

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