MODULATION OF RAS TRANSFORMATION AFFECTING CHROMATIN SUPRAORGANIZATION AS ASSESSED BY IMAGE-ANALYSIS

Changes in chromatin supraorganization defined in terms of patterns of chromatin texture were studied by video image analysis in Feulgen-sta ined revertants of LTR-ras-transformed NIH 3T3 cells and in cell lines obtained by transfection of these revertants with sense and antisense constructs of the lysyl oxidase gene (also named Lox or ''ras recisio n gene''). The objective was to determine whether changes in expressio n of the Lox gene, which have been assumed to modulate cell transforma tion by ras, could also affect the chromatin supraorganization changes known to be elicited in NM 3T3 cells by ras transformation. The image analysis results revealed that, although a nuclear phenotype visually similar to the most frequent one (III) in ras-transformed NM 3T3 cell s also appeared in the revertant, it contained a remarkably less tight chromatin packing state. This situation was also found in the reverta nt transfected with the sense construct of the Lox gene, but in the re vertant transfected with the Lox antisense constructs the chromatin te xture of the III phenotype was equal to or close to that of the ras-tr ansformed cells. With regard to the nuclear phenotype characterized by abundant loosely packed chromatin and less represented in the transfo rmed cell lines (I'), changes in the various cell lines, although dete ctable, were not as drastic as those reported for the III phenotype, T he enhancement in chromatin condensation of the type III nuclei, which affects euchromatin, is probably associated with a limited transcript ion of the genome. Although the image analysis results are mostly in a greement with previously published data on the molecular biology and t umorigenicity of the same cell lines, it appears that the phenomenon o f chromatin condensation once established in NM 3T3 cells by LTR-ras t ransformation could not be totally reverted by simply affecting Lox ex pression. (C) 1995 Academic Press, Inc.