We have performed detailed studies of the spreading of X inactivation in five unbalanced human X;autosome translocations. Using allele-specific RT-PCR we observed long-range silencing of autosomal genes located up to 45 Mb from the translocation breakpoint, directly demonstrating the ability of X inactivation to spread in cis through autosomal DNA. Spreading of gene silencing occurred in either a continuous or discontinuous fashion in different cases, suggesting that some autosomal DNA is resistant to the X inactivation signal. This spread of inactivation was accompanied by, but not dependent upon, CpG island methylation. Observations of late-replication, histone acetylation and histone methylation show that X inactivation can spread in the absence of cytogenetic features normally associated with the inactive X. However, the distribution of histone modifications which distinguish the inactive X are more accurate cytogenetic measures of the spread of X inactivation than late-replication. Overall, despite remarkable variation in the spread of X inactivation among the five cases there was good correlation between the pattern of gene silencing and the attenuation of clinical phenotype associated with each partial autosomal trisomy. We discuss our observations in the context of hypotheses which address the spread of X inactivation.