Complexing Rauscher leukemia virus reverse transcriptase with human plasma ribonuclease from Hodgkin's disease patients.

Human ribonucleases were purified from the sera of Hodgkin's disease patients by sequential column chromatography. The purified enzyme interacted with reverse transcriptase of Rauscher leukemia virus and formed an additive complex of Mr = 130,000. RNase and oligo(dG)-directed reverse transcriptase activities were diminished in the complex. The complex could be dissociated with the subsequent restoration of both activities in the presence of spermidine. The molecular weight of the complex suggest that the 2 RNase molecules bind to a single reverse transcriptase molecule.


Human ribonucleases
were purified from the sera of Hodgkin's disease patients by sequential column chromatography. The purified enzyme interacted with reverse transcriptase of Rauscher leukemia virus and formed an additive complex of II+', = 130,000. RNase and oligo(dG)-directed reverse transcriptase activities were diminished in the complex. The complex could be dissociated with the subsequent restoration of both activities in the presence of spermidine. The molecular weight of the complex suggest that the 2 RNase molecules bind to a single reverse transcriptase molecule.
It is well established that reverse transcriptase is a marker of RNA tumor viruses (1,2). These viruses, commonly found in the lower animals, have been demonstrated to be major etiologic agents in the development of cancer within the species studied (3)(4)(5)(6). Studies in man have been performed to determine whether such viruses play an analogous etiologic role in the development of human neoplasia but with little success.
The presence of reverse transcriptase and tumor virus-specific RNA in human cancer has been described (7, 81,  A-A, RNase activity.

Specificity
of Interaction  between  Reverse  Transcriptase  and Serum  RNase-The  binding  between  serum  RNase  and  Rauscher  leukemia  virus  reverse  transcriptase  appeared  to  be limited to RNase obtained from sera of Hodgkin's disease patients (Table  II). Although several attempts were made to bind serum RNase obtained from normal volunteers to reverse transcriptase, no evidence for complexing of any kind could be seen. To examine if the binding between the two enzymes was nonspecific, an attempt was made to bind the serum RNase from Hodgkin's or from normal sera to a variety of other proteins.
Of interest is that the other major viral specific protein, ~30, did not bind to RNase obtained from either source (Table II), nor was there any immunological or physical evidence for the existence of a complex (e.g. the migration of a complex in the heavier region of the gradient) between p30 and human serum RNase (Fig. 4). Similarly, nonviral proteins also failed to show any binding to the serum RNase (Table  II)  RNase Reverse Transcriptase Interaction for a more definitive study because of the known elevation of nucleolytic activity found in these patients (73%) (25). Four patients selected in this study have about a 2-fold higher serum RNase activity than the two other normal individuals. Purification and partial characterization of serum RNase from two normal individuals and the four Hodgkin's disease patients demonstrated no significant difference between RNase with respect to molecular weight, template specificity, and their elution profile when subjected to column chromatography. Serum RNase from Hodgkin's disease patients appear, however, to complex specifically with Rauscher leukemia virus reverse transcriptase when a variety of other viral and nonviral proteins do not. Formation of the RNase.reverse transcriptase complex can be studied by the addition of Mn*+ but not by a number of other divalent cations.
Although the exact molar ratio of RNase and reverse transcriptase in the complex is not known, it is suggested because of a molecular weight of about 130,000, that 2 RNase molecules bind to a single reverse transcriptase molecule. This assumes a molecular weight of 30,000 for the RNase and one of 70,000 for reverse transcriptase (13,15). The small percentage (3 to 10%) of binding between Rauscher leukemia virus reverse transcriptase and Hodgkin's disease serum RNase may be due to heterologous disparity. Similar disparity was also observed by Kufe et al. in their molecular hybridization studies between proviral DNA of murine leukemia virus and polysomal RNA of human sarcoma (26).
The present observation demonstrating the binding of Hodgkin's disease serum RNase with Rauscher leukemia virus reverse transcriptase leading to the specific inhibition of dGMP incorporation suggests a potential controlling role of the RNase in the synthesis of viral DNA. Further work utilizing a variety of other type C virus reverse transcriptases and RNases obtained from Hodgkin's disease patients as well as other patients with neoplasia is now underway and may add to our understanding of these observations. Acknowledgments -Thanks are due to Drs. Stanford Moore and Dalton Wang (The Rockefeller University) for their evaluation of this manuscript.