Stem cell gene expression changes induced specifically by mutated K-ras.

摘要

K-Ras proteins transduce signals from membrane-bound receptors via multiple downstream effector pathways and thereby regulate fundamental stem cell processes that affect neoplasia, including proliferation, apoptosis, and differentiation, but their contribution to tumourigenesis is unclear. Because cancers develop from stem cells, we set out to determine the characteristic changes in gene expression brought about by mutated K-ras (without interference from normal K-ras) in otherwise normal stem cells. cDNA microarrays were used to analyze gene expression profiles comparing wild-type murine embryonic stem (ES) cells with K-ras(Val12) expressing ES cells (previously made null for both endogenous K-ras alleles and transfected with K-ras(Val12), with valine for glycine at codon 12). K-ras(Val12) was expressed at 1.2-fold normal K-ras levels and produced transcripts for both activated K-Ras4A and 4B isoforms. The array expression data were confirmed by real-time quantitative PCR analysis of selected genes expressed both in the K-ras(Val12) expressing ES cells (R = 0.91 with array data) and in the normal intestinal tissues of K-ras(Val12) transgenic mice (R = 0.91 with array data). Changes in gene expression were correlated with the effects of K-ras(Val12) expression on ES cells of enhancing self-renewal in an undifferentiated state, increasing susceptibility to DNA damage-induced apoptosis, and increased proliferation. These expression data may explain, at least in part, some neoplasia-related aspects of the phenotypic changes brought about in this ES cell line by mutated K-ras, in that upregulation of cell growth-related proteins and DNA-associated proteins is consistent with increased proliferation; upregulation of certain apoptosis-related proteins is consistent with a greater susceptibility to DNA damage-induced apoptosis; and downregulation of structural proteins, extracellular matrix components, secretory proteins and receptors is consistent with a less differentiated phenotype.