Kirsten Murine Leukemia Virus Research Articles

Transformation/neodifferentiation of human skin fibroblasts by acute oncornaviruses and dexamethasone.

We demonstrated that the Kirsten murine sarcoma virus (KiMSV) and the Harvey murine sarcoma virus (HaMSV) converted human skin fibroblasts (HSF) into adipocytes. Adipocytic conversion of HSF by KiMSV and HaMSV was dependent on the presence of glucocorticosteroids. The Kirsten murine leukemia virus, the Harvey murine sarcoma [corrected] virus and the amphotropic helper virus (AP292) were ineffective by themselves. Balb murine sarcoma virus and Moloney murine sarcoma virus were, to a lesser degree, able to effect adipocytic conversion of HSF. In contrast, the feline sarcoma virus and the simian sarcoma virus did not cause this conversion. Together, the results suggest a role for certain oncogenes and glucocorticosteroids in the transformation/neodifferentiation of human cells.

Virus-like 30S RNA is selectively packaged by Kirsten leukemia virus in virions of smaller size class. Brief report.

Virus particles containing Kirsten murine leukemia virus 38 S RNA and a pseudotyped virus-like cellular 30 S RNA, were partially separable. This provides a means for enriching a population of virions for those containing 30 S RNA.

Characterization and cloning of the Kirsten murine leukemia virus genome

We have characterized the intracellular and circular unintegrated proviral DNA species of Kirsten murine leukemia virus by restriction mapping using the Southern blotting technique. These studies show the 8.5 kilobase pair genome to possess long terminal repeats (0.5 kilobase pairs in length) which are indistinguishable from those of the derivative Kirsten murine sarcoma virus. In addition, we have identified a 3′-located region in Kirsten murine leukemia virus which is very similar to the putative leukemogenic region of Gross murine leukemia virus. We also report the cloning of the leukemia virus genome using DNA obtained from the endogenous reverse transcriptase reaction of detergent disrupted virions.

Dissociation between transformed and differentiated phenotype in rat thyroid epithelial cells after transformation with a temperature-sensitive mutant of the Kirsten murine sarcoma virus.

Differentiated rat thyroid epithelial cells, infected in vitro with a temperature-sensitive mutant of the Kirsten murine sarcoma virus, expressed at the permissive temperature (33 degrees C) some phenotypic properties typical of transformed cells, including morphological features, colony formation in agar, and induction of tumors in newborn animals. Specific functional markers of these differentiated cells, i.e., synthesis/secretion of thyroglobulin, synthesis of thyroglobulin mRNA and iodide uptake, were blocked during growth at 33 degrees C. Normal morphology, failure to grow in agar, and the requirement of hormones for optimal growth were all restored after shifting to the temperature nonpermissive for transformation (39 degrees C), though the typical differentiated functions remained blocked. Infection with a leukemia helper virus clone (Moloney or Kirsten murine leukemia virus) did not lead to the loss of the differentiated phenotype of rat epithelial thyroid cells, thus demonstrating that the loss of the differentiated phenotype is caused by the sarcoma virus component. These results indicate that the expression of some of the phenotypic properties of transformed differentiated rat thyroid epithelial cells is under the direct control of the p21 thermosensitive activity, whereas the block in the expression of two typical differentiation markers of thyroid epithelial cells is irreversible and probably controlled by different mechanisms.

Sequence relationships between kirsten retrovirus genomes and the genomes of other murine retroviruses

RNA sequence relationships between the genomes of the Kirsten murine sarcoma virus (MSV-K) complex, the Kirsten murine leukemia virus (MuLV-K) complex, the Gross murine leukemia virus (MuLV-G), and the Moloney murine leukemia virus (MuLV-M) were investigated. Sedimentation analyses revealed the expected 30 and 34 S RNA subunits in the MSV-K complex and a previously undetected 30 S RNA subunit accompanying the 34 S RNA subunit in the MuLV-K complex. Nucleic acid hybridization data indicated that each Kirsten virus 30 S RNA subunit had about 40% sequence homology with the RNA genome of MuLV-G, although these sequences were only partially homologous between the two 30 S subunits. In contrast, the MuLV-K 34 S RNA subunit had 96% sequence homology with the MuLV-G genome, whereas the MSV-K 34 S RNA subunit displayed only 71% sequence homology with the MuLV-G genome. Similar relationships were indicated by oligonucleotide fingerprinting. The oligonucleotide data, taken with published sequence data on the MuLV-G and MuLV-M genomes, enabled us to construct partial sequence maps of the MuLV-K 34 S RNA subunit and the MSV-K 34 and 30 S RNA subunits. The sequence arrangements indicated that (1) the MuLV-K 34 S RNA subunit is a variant of the MuLV-G genome; (2) the MSV-K 34 S RNA subunit is a recombinant molecule, which maintains the lenght of its leukemia virus parent; and (3) the MSV-K 30 S RNA subunit may have been generated from the MuLV-K 34 S genome by a two-stage process, culminating in the retention of parental sequences only within the U5 and U3 noncoding segments and within several amino-terminal coding segments. Further examination of published retrovirus genome sequences revealed several strategically situated sets of potential recognition signals for transcription and translation and suggested a model for genetic recombination based on mRNA splicing signals and areas of limited sequence homology. This model may explain how foreign gene elements can be inserted into retrovirus genomes to generate either functional or defective recombinant retroviruses.

Genetic organization and cloning of kirsten murine sarcoma virus DNA

Summary The Kirsten murine sarcoma virus genome is a recombinant molecule derived from the Kirsten murine leukemia virus genome and sequences transduced from a normal rat cell. We show here that leukemia virus derived sequences comprise the 5′ terminal 0.5 kilobasepairs (kbp) and the 3′ terminal 1.2 kbp of the linear 6.5 kbp sarcoma virus DNA. The parental leukemia virus thus contributes the long terminal repeat sequences (0.5 kbp) and some additional 3′ sequences to the sarcoma virus genome. Using DNA synthesised by detergent-disrupted virions a 4.6 kbp SmaI fragment containing the rat-specific sequences of the sarcoma virus was cloned into the plasmid pAT153. This clone was then used as a sarcoma virus specific probe to clone full sized circular viral DNA from infected NIH 3T3 cells.

Isolation of a Kirsten murine leukemia virus mutant temperature sensitive for particle production and env gene minus

An NIH/3T3 cell, nonproductively infected with Kirsten murine leukemia virus (Ki-MLV), was isolated shortly after a low multiplicity of infection. The clone, K23, harbors a Ki-MLV genome with two defects: virus particle production is temperature sensitive and no envelope gene products can be detected immunologically. Intracellular Ki-MLV RNA consists of a single species which is of genome size. No Ki-MLV RNA is detected which corresponds to the env gene mRNA. This defect in RNA processing accounts for the lack of gp70 and p15E. The temperature-sensitive lesion may be in processing of Pr65 gag or Pr65 gag/pol precursor proteins. At 39 or 33° the cells harbor Pr65 gag and Pr180 gag/pol . Intracellular p30, however, is only detected in cells at 33°. At 33°, the permissive temperature, virus particles are released which have a density of 1.13 g/ml, contain p30, p12, p10, reverse transcriptase, and RNA, but lack gp70 and p15E. At 39° no particles are detected. Thus, K23 appears to contain a viral genome with mutations which affect processing of both viral precursor proteins and mRNAs. The nature of these defects is discussed.

Physical map of the Kirsten sarcoma virus genome as determined by fingerprinting RNase T1-resistant oligonucleotides

From analysis of the large RNase T1-resistant oligonucleotides of Kirsten sarcoma virus (Ki-SV), a physical map of the virus genome was deduced. Kirsten murine leukemia virus (Ki-MuLV) sequences were detected in T1 oligonucleotides closest to the 3' end of the viral RNA and extended approximately 1,000 nucleotides into the genome. The rat genetic sequences started at this point and extended all the way to the very 5' end of the RNA molecules, where a small stretch of Ki-MuLV sequence was detected. By comparison of the fingerprints of Ki-SV RNA and the RNA of the endogenous rat src genetic sequences, it was found that more than 50% of the T1 oligonucleotides were similar between Ki-SV and the endogenous rat src RNA, suggesting an identical primary nucleotide sequence in over 50% of the viral genomes. The results indicate that Ki-SV arose by recombination between the 5' and 3' ends of Ki-MuLV and a large portion of the homologous sequences of the endogenous rat src RNA.

Murine leukemia viruses containing an ∼30 S RNA subunit of unknown biological activity, in addition to the 38 S subunit of the viral genome

The 50–70 S RNA complex of a clone of Kirsten murine leukemia virus (Ki-MLV), propagated in NIH 3T3 cells, as well as that of a Moloney (Mo)-MLV, propagated in a mouse thymus-bone marrow cell line, contained an ∼30 S RNA of unknown biological activity in addition to 38 S genomic RNA. The ∼30 S RNA shares with 38 S virion RNA the ability to form 50–70 S RNA complexes and to be packaged by virus particles. The morphology of the cell lines propagating these viruses was indistinguishable from that of uninfected controls.

Studies on the type C viral replication defect in MSV-transformed rat nonproducer variant clones

Two clonal derivatives of an MSV-transformed nonproducer rat cell line, K-NRK C1 32, have been previously found to be both relatively contact inhibited and nonpermissive for type C viral replication, unlike the parental cell. In the present report, the infectivity of vesicular stomatitis virus (VSV) and its Kirsten murine leukemia virus pseudotype [VSV(KiMuLV)] have been compared on the two variants and the parental clone. All three cell lines are equally permissive for VSV, but compared to K-NRK C1 32, both variants are nonpermissive for VSV(KiMuLV). These data strongly suggest that both subclones are defective in the processes of type C virus adsorption, penetration, and/or uncoating.

An attenuated mouse leukemia virus. I. Origin and immunization.

An attenuated Kirsten murine leukemia virus was isolated from the culture fluids of a chronically infected C3Hf/Gs mouse embryo cell line. The attenuated virus failed to induce erythroblastosis or lymphomas in mice after 16–18 months. Inoculation of 3-week-old C3Hf/Gs mice with attenuated leukemia virus prevented induction of erythroblastosis or lymphomas by virulent virus given 1 month later. A similar protective effect was observed when 3-week-old Wistar/Furth rats were inoculated with attenuated virus and challenged 1 month later with a lethal dose of rat-adapted leukemia virus.

Ribonucleic acid components of murine sarcoma and leukemia viruses.

Defective Kirsten murine sarcoma virus was present as leukemia virus pseudotype [Ki-MSV(MLV)] in a 10- to 100-fold excess over its helper, Kirsten murine leukemia virus (Ki-MLV), when the two viruses were propagated in an NRK rat cell line. The s(omega,20) of the fastsedimenting RNA complex of Ki-MLV and of Ki-MSV-(MLV) was 62 S and 55 S, respectively. Gel electrophoresis in buffered aqueous or formamide solution of the dissociated 62S RNA complex of Ki-MLV showed a single major peak of molecular weight about 2.5 x 10(6). Dissociated 55S RNA of Ki-MSV(MLV) was resolved into a major component with a higher electrophoretic mobility than that of Ki-MLV RNA and molecular weight about 2.3 x 10(6). Occasionally, a minor component with the same electrophoretic mobility as Ki-MLV RNA was observed in Ki-MSV(MLV) RNA; it is thought to be the RNA of Ki-MLV present as helper virus in our stocks of Ki-MSV(MLV). The RNA of an endogenous rat C-type RNA virus was electrophoretically different from both Ki-MLV RNA and Ki-MSV(MLV) RNAs. Oligonucleotide fingerprinting of the RNAs digested with RNase T1 indicated that the RNAs of Ki-MSV(MLV) and Ki-MLV are different. However, the extent of the difference between the two RNAs could not be estimated by this method. The heat-dissociated 50-70S RNAs of two other defective murine sarcoma-leukemia viruses; Harvey-MSV(MLV) and Moloney-MSV(MLV) and of defective spleen-focus-forming Friend virus were resolved electrophoretically into two components. The larger components had the same electrophoretic mobility as the RNA of Ki-MLV or Moloney MLV. The smaller were not present in leukemia virus. It is suggested that the small RNA components of the two murine viruses and of Friend virus represent specific genetic information of these replication-defective transforming viruses. Possible relationships between the RNAs of murine leukemia viruses and replication-defective murine sarcoma and Friend viruses are discussed.