THE ACTIVATED MLVI-4 LOCUS IN MOLONEY MURINE LEUKEMIA VIRUS-INDUCED RAT T-CELL LYMPHOMAS ENCODES AN ENV MLVI-4 FUSION PROTEIN/

A genomic DNA probe derived from the region immediately 3' of the clus ters of integrated proviruses in the Mlvi-4 locus detects a 5.5-kb mRN A transcript which is specifically expressed in normal rat thymus and spleen. The same probe detects two tumor-specific mRNA transcripts 2.5 and 10 kb long, both of which are expressed only in tumors carrying a provirus in the Mlvi-4 locus. Sequence analysis of two cDNA clones (L E3a and B1.1) of the 2.5-kb tumor-specific mRNA, obtained from two ind ependent tumors (6889 and B1), revealed that they are both derived fro m hybrid env/Mlvi-4 mRNA transcripts. The splicing of env to Mlvi-4 se quences linked a cryptic splice donor site at nucleotide position 6397 of the viral genome with a splice acceptor site in the region immedia tely 3' of the integrated provirus. The mRNA that gives rise to cDNA c lone B1.1 terminates 1,005 bases 3' of the splice acceptor site withou t additional splicing. The mRNA that gives rise to cDNA clone LE3a ter minates in the same site but undergoes differential splicing of an 81- base-long intron. The resulting mRNAs contain 247-amino-acid (clone B1 .1) or 226-amino-acid (clone LE3a) open reading frames sharing 221 N-t erminal amino acids, of which 207 are derived from the viral env gene and 14 are derived from Mlvi-4. RNase protection assays using 6889 tum or cell RNA and a probe derived from the cDNA clone LE3a detected both mRNA transcripts. More abundant of the two, however, was the one enco ding the putative 247-amino-acid protein. Transient transfections of a construct expressing the RNA transcript defined by clone B1.1 into D1 7 cells led to the expression of an Env/Mlvi-4 fusion protein with an apparent molecular mass of 33 kDa. Given that cells with provirus inse rtions in the Mlvi-4 locus are selected and that retroviral env gene p roducts may have profound effects in the biology of hematopoietic cell s, we suggest that the detected fusion proteins may contribute to the growth of T-cell lymphomas.