In a previous communication, we showed that the H2A.1/H2A.2 histone variant ratio decreases in a linear manner during the in vitro aging of human diploid fibroblasts. This ratio is known to decrease in the same manner in progressive stages of development and in the process of differentiation, and is thus considered to be a biochemical marker for differentiation. A detailed analysis of the synthesis of H2A and H3 histone variants as a function of cumulative population doublings in the same in vitro cell system is presented in this study. Quantitative analysis of these variants in the G0 phase, synchronized fibroblasts has shown that their relative amount in chromatin, as well as their biosynthesis rate, change during in vitro aging of human diploid fibroblasts, revealing both up-and down-regulation of certain variants as a function of cumulative population doublings. Furthermore, we show by morphometric studies employing the seven distinct fibroblast morphotypes, as described by the Bayreuther classification, that this regulation is attributable to the replicative sub-populations. These results reveal that histone variants of the H2A and H3 families are regulated during in vitro aging in the same manner as that during differentiation.
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