The broad long-term objectives of the proposed research are to understand in molecular detain the general mechanisms regulating tissue- specific gene expression and to obtain a complete molecular understanding of type III glycogen storage disease. This will be accomplished through the study of the human gene encoding glycogen debranching enzyme and its various subtypes of enzyme deficiency. In type III glycogen storage disease there are patients with deficient debranching enzyme activity in both liver and muscle, and patients with deficiency confined to the liver. In addition, there are patients with selective loss of one of the two enzyme activities associated with the debranching enzyme. The findings of multiple tissue-specific debrancher isoform mRNAs generated by differential transcription and splicing from a single debranching enzyme gene and identification of a striking and specific association of exon 3 mutations in glycogen storage disease type III patients having debranching enzyme deficiency only in the liver, make this disease a particularly informative model to examine the molecular mechanisms underlying the complexity of the control of tissue- specific gene expression.
The first aim i s to further characterize the promoters and regulatory region of the debranching enzyme gene and to investigate whether myogenic regulatory factors are involved in the expression of muscle-specific debrancher isoforms.
The second aim i s to elucidate molecular mechanisms underlying exon 3 mutations in conferring differential expression of the debranching enzyme in liver and muscle by analysis of mRNA isoforms and protein expressed in the muscle.
The third aim i s to identify and characterize mutations in different subtypes of type III glycogen storage disease by biochemical subtyping of the disease, SCCP analysis, DNA sequencing and expression studies. Information gained by analysis of the debrancher gene and molecular dissection of different subtypes of type III glycogen storage disease will provide insight into the molecular basis of the disease, functional domain for this multifunctional enzyme, and general mechanisms controlling tissue-specific gene expression.
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