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A Determination of the Outer Dimensions of Oncornaviruses by Several Electron Microscopic Procedures¹

Worcester Foundation for Experimental Biology, Shrewsbury, Massachusetts 01545 [R. B. L., P. N. M.], and Departments of Microbiology and Surgery, Duke University Medical Center, Durham, North Carolina 27710 [K. C., D. P. B.]

The effect of negative stain, freeze-fracture, and thin-sectioning procedures on the size of several oncornaviruses has been determined. After negative staining, the average outer dimensions measured over several experiments for the C-type avian viruses, avian myeloblastosis virus (AMV) and Rous sarcoma virus, Prague strain, were about 1370 and 1340 Å, respectively. These values were substantially smaller than the outer dimensions of the C-type murine viruses, Friend leukemia virus (FLV), namely, 1470 Å, and Moloney leukemia virus, namely, 1460 Å. The avian viruses were similar in size to the B-type murine virus, mouse mammary tumor virus, which measured about 1290 Å. All experimental values were obtained after glutaraldehyde (1.5%) fixation and uranyl acetate (2%, pH 4.2) staining. Other negative stains, such as sodium phosphotungstate (1%, pH 6.6), lead to disruption of the outer envelope and a concomitant diminution in size. Freeze-fractured preparations of AMV showed particles with or without a complete outer envelope. The former class ranged in size from 950 to 1450 Å; the latter ranged from 800 to 1150 Å. FLV, in contrast, exhibited only the smaller class of particles, suggesting that FLV is unable to sustain an internal fracture beyond its outer envelope. This apparently greater fragility of the FLV outer envelope is consistent with the more rapid loss of the envelope of FLV, compared to AMV, observed when virions were subjected to a brief 0.01% sodium dodecyl sulfate treatment. Utilizing catalase crystals as an internal calibration marker for the first time in thin sections, it was found that the diameter of both FLV and AMV were larger than previously published values. For FLV, the distribution of outer dimensions, when corrected for specimen shrinkage, ranged between 1100 and 1450 Å. The larger values of this distribution probably arose from sections near the equator of the particle and are more probably representative of the actual virion diameter. For AMV, the corrected range of values was between 1200 and 1500 Å, which was about 100 Å larger than expected. This result may be interpreted as indicating that the AMV outer envelope differs from that of FLV, in that it can swell under conditions of low osmolality, e.g., associated with OsO₄ fixation. Consistent with this interpretation is the observation that freeze-thawing of unprotected dilute AMV suspensions led to an increase of the outer diameter by as much as 200 Å and further that the swelling could be prevented by addition of bovine serum albumin (0.05%) to the viral preparation prior to freezing. No such change was found for FLV.

In summary, the estimated size ranges for three classes of oncornaviruses are: (a) 1250 to 1450 Å, for avian C-type viruses (AMV and Rous sarcoma virus, Prague strain); (b) 1350 to 1550 Å, for murine C-type viruses (FLV and Moloney leukemia virus); and (c) 1200 to 1400 Å, for B-type viruses (mouse mammary tumor virus).

Concomitant studies of oncornavirus cores indicated a clear difference between avian and murine type viruses. AMV cores measured about 730 Å in diameter and had a ribonucleoprotein component that was centrally collapsed FLV cores in contrast measured about 840 Å and appeared to have the ribonucleoprotein closely associated with the core shell. These results suggest that the 100-Å smaller value obtained for the outer dimension of avian viruses is due to the presence of a smaller core component.

¹ Supported by USPHS Grant CA-11976 from the National Cancer Institute and Contract NO-1-CP-33308. This research was initiated at Duke University.

² To whom request for reprints should be addressed.

Received 12/ 4/73. Accepted 4/ 5/74.