Delta-crystallin gene expression and patterns of hypomethylation demonstrate two levels of regulation for the delta-crystallin genes in embryonic chick tissues.
In this study we address two questions regarding the control of delta-crystallin gene expression in chick embryos. First we have determined whether delta-crystallin mRNA is found outside of the developing lens, in which it is the predominant mRNA. We find that this mRNA can be detected, although at relatively low levels, in all embryonic tissues we have examined (from the definitive streak stage onward). This low level of transcription may be related to a second function for one or both of the delta-crystallin genes: both genes have a high degree of sequence identity to the enzyme argininosuccinate lyase. This result led us to a second set of experiments in which we reevaluated the possible role of hypomethylation in the expression of the delta-crystallin genes. Previous work showed that particular HhaI and HpaII sites in the crystallin genes undergo hypomethylation early in the process of lens differentiation when there is a burst of delta-crystallin mRNA accumulation. We not find that these sites remain methylated in nonlens tissues, implying that they cannot be required for the delta-crystallin gene activity found in these tissues. Other sites are constitutively hypomethylated, however, and may be functionally linked to this low level of gene activity. From an analysis of the kinetics of the developmentally regulated hypomethylation of HhaI and HpaII sites we also find that complete hypomethylation of these sites is not required for activating high levels of delta-crystallin transcription during lens differentiation. We do find, however, that these sites approach a fully hypomethylated state later in the lens differentiation process. Our analyses of mRNA levels and hypomethylation together lead us to propose that the delta-crystallin genes are regulated by two different mechanisms, one that leads to high levels of expression in the lens and the other which is responsible for low level expression in all other tissues in the chick embryo.