ANTISENSE INHIBITION OF LOW-AFFINITY NERVE GROWTH-FACTOR RECEPTOR IN KIDNEY CULTURES - POWER AND PITFALLS

1. Antisense inhibition of gene expression implies that the expression of the target protein is selectively inhibited at either the translat ional or the transcriptional level by complementary DNA or RNA constru cts that are antiparallel to the target sequence. The antisense inhibi tion strategy provides means to study the roles of individual proteins and has, in spite of its limitations, gained a wide range of both the rapeutic and experimental applications, 2. In developmental biology, p rotein expression has been selectively inhibited by the use of antisen se gene transfection and by antisense deoxyoligonucleotides, The trans fectability of embryonic tissues is variable, but in general fetal and embryonic cells take up foreign DNA relatively efficiently, in partic ular, short deoxyoligonucleotides that penetrate mesenchymal cells wit hin a few hours without any manipulation. 3. We have now evaluated the advantages and pitfalls of antisense inhibition by deoxyoligonucleoti des in organ culture and describe our experience from the inhibition o f low-affinity nerve growth factor receptor expression in embryonic mo use and rat kidneys. 4. The expression of nerve growth factor receptor can be specifically inhibited by deoxyoligonucleotides, but the targe t sequence-dependent window of, in particular, phosphorothioate-modifi ed oligonucleotides is quite narrow. The culture conditions affect the response to the oligonucleotides and their cellular incorporation is variable with respect to the cell type and stage of differentiation.