Differentiation-associated changes in in vitro mRNA translation in the HL60 cell line

Abstract

In order to characterize certain aspects of gene expression during the granulocytic differentiation of the HL60 cell line, we have analysed changes in the population of mRNA available for translation in vitro. RNA extracts of DMSO-induced and control cells were translated in vitro in a wheat germ cell-free protein synthesizing system. Translation products were analysed by two-dimensional electrophoresis followed by autoradiography. Autoradiograms were analysed by a computer-assisted method utilizing a drum-scanning microdensitometer. Spots were identified by their relative positions on the films and their relative intensity was estimated. One hundred and eighty-one peptides were identified in both the DMSO-induced and untreated control HL60 cells, 31 of which showed differentiation-associated changes in synthesis in vitro. The 11 peptides which decreased in synthesis did so early in the differentiation process, whereas most of the 20 peptides which increased did so at a later time. Three peptides were shown to increase more than 8-fold by day 4 of induction. A comparison with normal granule proteins from human leukocytes suggests that at least two of these may correspond to functional granule proteins. The changes in peptide patterns which we describe demonstrates that the programm of gene expression during HL60 differentiation includes changes in the relative abundance of specific mRNA transcripts. The data described here also provides a standard for comparison of other proteins, such as oncogene products, as they are identified.