RELAXATION OF INSULIN-LIKE GROWTH-FACTOR-II IMPRINTING IN RAT CULTURED-CELLS

The parental-specific expression of the insulin-like growth factor-2 ( Igf-2) and H19 genes was studied in rat fibroblast cells derived from a 3 day-old first-generation hybrid animal obtained by crossing Fisher and Wistar strains (F x W cells). Results showed that the reciprocal imprinting of the Igf-2 and H19 penes was conserved in the rat tissues and in the derived F x W cells when cultured with frequent transfer. Igf-2 and H19 gene expression was coordinately up-regulated upon reach ing confluence, but Igf-2 RNA levels were further increased in a time- dependent manner and the repressed state of the maternal Igf-2 allele was progressively relaxed in cultures held in the confluent state and in the presence of low serum for more than 3 days. The active expressi on and relaxed imprinting status of the Igf-2 gene persisted over cell generations when the growth-constraining conditions were released by trypsinization and dilution. On the contrary, the imprinting of the H1 9 gene appeared to be unaffected by changes in growth conditions and i ts expression was down-regulated when the confluent cells were passage d. Methylation of the H19 promoter and Igf-2 coding regions was increa sed in the F x W cells extensively held under confluence and in the de rived 'post-confluent' cultures. The heritable changes in the expressi on, and imprinting status of the Igf-2 and H19 genes observed in the F x W cells closely resembles events described in human embryonal cance rs and cancer-predisposing syndromes. The occurrence of imprinting rel axation under strong growth-inhibitory conditions supports the hypothe sis that it is an epigenetic change. (C) 1997 Elsevier Science Ireland Ltd.