We propose to continue our search for ways of understanding normal and abnormal gene expression in man by studying human and other mammalian somatic cells in culture.
The specific aims of this investigation include: further characterization of the apoenzymes and coenzymes involved in propionate and methylmalonate metabolism; definition and regulation of the cellular pathway of cobalamin (vitamin B12) utilization; elucidation of the bases for biochemical heterogeneity and pyridoxine-responsiveness in cystathionine synthase deficiency; and extension of current approaches employed for prenatal detection of inborn errors. Toward these aims, skin fibroblasts and heterokaryons therefrom will be propagated. We will also explore the properties of rat hepatocytes in short-term culture as they relate to the scientific areas mentioned above. To accomplish this work, the following kinds of experiments will be employed: binding of vitamins to specific receptor sites on cell surfaces; regulation of inter-organellar movement of vitamins; kinetics of apoenzyme-coenzyme interaction; immunochemical characterization of enzymatic deficiency; regulation of apoenzyme turnover by coenzyme; and complementation analyses in intraspecific heterokaryons or hybrids. It is anticipated that these studies will define normal biochemical mechanisms as they further our understanding of inherited human disease states.
