The experiments described in this proposal are designed to analyze changes which occur in the chromatin surrounding the contractile protein genes during embryonic muscle development. We have previously shown that the contractile protein genes are not expressed in dividing myoblasts, and are coordinately activated when myoblasts begin to fuse to form multinucleate myofibers. Recently changes in methylation of DNA bases and changes in the sensitivity of regions of the DNA to low levels of nuclease have been coorelated with the activation of specific genes during development. We intend to use contractile protein genes which we have cloned as probes to look for changes in the pattern of methylation and nuclease sensitivity of chromatin surrounding the contractile protein genes at various times during myoblast differentiaton. We might expect that a group of coordinately activated genes would show similar changes in methylation or nuclease sensitivity if these parameters are indeed associated with activation of genes during development. By using agents which alter the methylation of DNA and other agents which interfere with myogenesis we hope to be able to determine whether changes in methylation or nuclease sensitivity are actual causal events in the activation of the contractile protein genes during myoblast differentiation.
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