A mutant v-rel with increased ability to transform B lymphocytes.
We observed that two strains of REV-T differ in the ability to transform bursal cells in vitro. REV-TW, with v-rel derived from a well-characterized clone and considered the prototype of the wild type, fails to generate colonies in soft agar. In contrast, REV-S2A3, derived from the S2A3 cell line, readily transforms bursal cells. With PCR, a 1,591-bp fragment containing v-rel from the REV-S2A3 provirus was cloned into plasmid pREV-0. Except for the absence of v-rel, pREV-0 is identical to pREV-TW. Five clones of pREV-PCR, each produced by an independent amplification, were obtained. The REV-PCR viruses displayed the strong transforming phenotype of REV-S2A3. Two mutations were identified in the 5' region of v-rel from REV-PCR1 to REV-PCR5: a silent mutation and a G-to-T transversion, changing the alanine at position 40 to serine. To confirm the relevance of this amino acid substitution, a 478-bp fragment containing the mutations was exchanged between REV-TW and REV-PCR1. Only the mutant viruses were able to form large colonies of bursal cells in liquid culture and to generate bursal cell colonies in soft agar. When tested on splenocytes, the wild-type viruses induced predominantly non-B-cell colonies while the mutant viruses gave origin mainly to B-cell colonies. The above results indicate that the substitution of serine for alanine at position 40 of v-Rel enhances the ability of REV-T to transform B lymphocytes in vitro. This mutation is close to the DNA-binding region, and the variant v-Rel oncoprotein shows increased kappa B-binding activity, thus confirming the relevance of this property for transformation.
Romero
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Mary Babb Randolph Cancer Center, West Virginia University, Morgantown 26506-9162.
Humphries
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D000409:Alanine / Q000235:genetics; D000595:Amino Acid Sequence; D000818:Animals; D001402:B-Lymphocytes / Q000473:pathology*; D001483:Base Sequence; D002460:Cell Line; D002471:Cell Transformation, Neoplastic / Q000235:genetics*; D002645:Chickens; D017931:DNA Primers; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D009154:Mutation; D020572:Oncogene Proteins v-rel; D015682:Retroviridae Proteins, Oncogenic / Q000235:genetics* / Q000378:metabolism; D012694:Serine / Q000235:genetics
D017931:DNA Primers; D004268:DNA-Binding Proteins; D020572:Oncogene Proteins v-rel; D015682:Retroviridae Proteins, Oncogenic; D012694:Serine; D000409:Alanine