M Oskarsson

Evidence for a role of Met-HGF/SF during Ras-mediated tumorigenesis/metastasis

Aberrations in Met-hepatocyte growth factor/scatter factor (HGF/SF) signaling have been implicated in the acquisition of tumorigenic and metastatic phenotypes. Here we show that murine NIH3T3 and C127 cells transformed by the Ras oncogene overexpress the Met receptor, resulting in enhanced HGF/SF-mediated responses in vitro including invasion through basement membrane. Accompanying the increase in Met in ras-transformed NIH3T3 cells, there is a decrease in endogenous HGF/SF expression as previously observed in cells exogenously overexpressing Met. However, subcutaneously grown tumors and experimental lung metastases derived from these cells express significantly higher levels of endogenous HGF/SF together with high levels of Met. These results suggest Met-HGF/SF signaling enhances tumor growth and metastasis of Ras-transformed NIH3T3 cells.

Chicken homolog of the mos proto-oncogene.

We compared the sequence and properties of the chicken mos homolog with the previously characterized mouse and human c-mos genes. Sequence analysis revealed one major open reading frame of 1,047 base pairs encoding a protein of 349 amino acids. Both the nucleotide sequence and the deduced amino acid sequence showed 62% overall homology to mouse and human c-mos, but regions of higher conservation (approximately 70%) occurred in the putative ATP-binding and kinase domains. We detected mos transcripts by Northern (RNA) analyses in RNA prepared from chicken and quail ovaries and testes. Evidence for low levels of mos RNA expression in adult chicken heart, kidney, and spleen and in the entire embryo was obtained by S1 nuclease protection experiments. In contrast to the low transforming efficiency of human c-mos when linked to a mouse retroviral long terminal repeat element, chicken c-mos transformed NIH 3T3 cells as efficiently as mouse c-mos did. We also show that chicken primary embryo fibroblasts were morphologically altered when infected with an avian retroviral vector containing the chicken c-mos coding region.

Cells transformed by certain strains of moloney sarcoma virus contain murine p60

It was previously demonstrated that the 60,000 dalton (p60) precursor-like polyprotein containing murine p30 was a constituent of the feline leukemia virus pseudotype of Moloney sarcoma virus [m1MSV(FeLV)]. It is now shown that p60 is detected in cells of five mammalian species transformed by m1MSV, indicating that p60 is specified by this genome. Moreover, little or no murine p30 is detected in the m1MSV-transformed cells, suggesting that the murine group p30 antigenic reactivity of S + L- cells is ude to p60. Pulse-chase studies in cells producing m1MSV(FeLV) show that p60 is the largest polypeptide detectable during the pulse, and that intracellular p60 is not cleaved into smaller (for example, p30) polypeptides during chase periods of up to 10 hr. The lack of cleavage of p60 is in contrast to the properties of p30 precursors detected in cells containing replicating avian or mammalian RNA tumor viruses. The inefficient cleavage of intracellular p60 and the kinetics of appearance of murine p30 in extracellular m1MSV(FeLV) suggest that p60 cleavage to p30 occurs in cells shortly before virus release. While only p60 was detected in the m1MSV-transformed cells, p60 and p70 were detected in m3MSV-transformed cells, and no immunoprecipitable polypeptides were detected in HT-1 MSV-transformed cells. The observed differences in the intracellular polypeptide expression by each of the strains of MSV suggests differences in genetic content.

A p60 polypeptide in the feline leukemia virus pseudotype of Moloney sarcoma virus with murine leukemia virus p30 antigenic determinants.

A 60,000-dalton polypeptide (p60) has been identified in the feline leukemia virus (FeLV) pseudotype of Moloney sarcoma virus [MSV(FeLV)]. This polypeptide is present in the purified virus complex in concentrations greater than either the murine p30 or the feline p27. Purified p60 crossreacts immunologically with murine p30 group antiserum and contains several interspecies determinants, whereas the group specific determinant of FeLV p27 is not detected. Comparison of peptide fingerprints of p60 and murine p30 show many peptides in common. Limited digestion of p60 with either trypsin or chymotrypsin produced p30-35 and p20 peptides which retain the MuLV p30 group and interspecies antigenic activities. The p30 produced by both enzymes comigrates in polyacrylamide gels with the murine p30 of MSV(FeLV), thus suggesting that p60 may be an uncleaved precursor to p30.

mos gene transforming efficiencies correlate with oocyte maturation and cytostatic factor activities.

The mos proto-oncogenes from different vertebrate species transform mouse NIH 3T3 cells with markedly different efficiencies. v-mos, mouse (c-mosmu), and chicken (c-mosch) mos transform NIH 3T3 cells 10- to 100-fold more efficiently than do human (c-moshu) and Xenopus (c-mosxc) mos. The mos genes with the highest transforming activity efficiently induce maturation in Xenopus oocytes and mimic cytostatic factor (CSF) by causing mitotic cleavage arrest in embryos. Chimeric v-mos/c-moshu proteins that had high transforming efficiencies in NIH 3T3 cells were also effective in the induction of oocyte maturation and CSF cleavage arrest. We measured the in vitro autophosphorylation activities of the different mos proteins and found that the levels of kinase activity of v-mos, c-mosmu, and c-mosch were much higher than that of c-mosxc. These data indicate that mos gene transforming efficiency and the ability to induce oocyte maturation or mimic CSF activity are correlated with in vitro autophosphorylation activity and suggest that the mos protein plays a similar role in transformed cells and normal oocytes.

Analysis of the transforming potential of the human homolog of mos

The human homolog, c-moshu, of the mouse cellular mos proto-oncogene (c-mosmu) transforms NIH 3T3 cells at low efficiency. Furthermore, the c-moshu-induced foci are less distinct, and transformed cells contain a high level of human mos protein. The transforming activity of hybrid mos genes derived from human and mouse sequences reveals three domains within the coding region, as well as a negative regulatory sequence upstream from the c-moshu ORF that reduces its transforming efficiency. The mos C-terminal region, however, which contains the src-kinase homology domain, appears to have the greatest influence on transforming efficiency. The low transforming efficiency of c-moshu may provide a selective advantage to the host, but it also may indicate a reduced or modified function of mos in humans.

The Long Terminal Repeat of Moloney Sarcoma Provirus Enhances Transformation

Transforming retroviruses are generally thought to arise from a recombination between nontransforming viruses and sequences of cellular origin. Until recently, it has not been possible to test whether the acquired cell sequences (onc)1 of transforming retroviruses act in concert with the conserved retrovirus sequences to establish transformation or whether the onc sequences alone are sufficient to cause transformation. For example, if retrovirus sequences are not essential, a finite number of naturally occurring malignancies would be expected to be caused by expression of C-onc sequences. Alternatively, if a region(s) of the retrovirus genome is (are) required for expression of the malignant phenotype, then the properties of this viral sequence may reveal how a normal cell sequence is modified to activate its transformation potential. We have cloned from normal mouse cell genomic DNA a sequence, C-mos that is homologous to the Moloney sarcoma virus (MSV) acquired V-mos sequence (Oskarsson et al. 1980). This cloned fragment and cloned integrated MSV proviral DNA have provided a direct means for addressing these questions and for testing the transforming activity of various combinations between the V-mos, C-mos, and MSV sequences derived from Moloney murine leukemia virus (M-MuLV).

TWO REGIONS OF THE MOLONEY LEUKEMIA VIRUS GENOME ARE REQUIRED FOR EFFICIENT TRANSFORMATION BY src/sarc

ABSTRACT Src -containing subgenomic DNA fragments of Moloney sarcoma provirus (MSV) with high transforming activity retain two regions of the parental Moloney leukemia virus (MuLV) genome. One MuLV region is the 600 base pair terminal repeat sequence of the provirus. The presence of this sequence either 5′ or 3′ to an internal MSV src sequence enhances transforming activity 1000-fold. The other MuLV region represents approximately 100 nucleotides preceeding the 5′ end of the src sequence. The requirement of this region for transformation is deduced by i) comparison of the endonuclease restriction maps of three MSV provirus isolates, ii) eliminating specific MuLV sequences in cloned subgenomic fragments without loss of biological activity, and iii) demonstrating that efficient transforming activity of the cell s arc sequence occurs only when MSV sequences have been substituted to replace the normal cell sequences at its 5′ end.