The objective of the proposed research is to understand why and how two isoenzymes of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) are localized and expressed in different cell lineages of the nematode, Caenorhabditis elegans. GAPDH-1 is enriched in non-muscle cells while GAPDH-2 is preferentially localized in the actin-containing zones of the body wall muscle lineages. Rabbit anti-nematode GAPDH antibody will be used to identify and hence purify the mRNAs coding for the two GAPDHs subsequent to cloning the homologous genomic and cDNA sequences. The GAPDH-1 and -2 genetic loci will then be identified by restriction site DNA polymorphism studies. The GAPDH DNA clones and the rabbit anti-nematode GAPDH antibody will be used as probes to monitor the temporal and spatial transcription/translation of GAPDH expression during the development of the wild-type nematode. The uncoordinated mutant, unc-52 II, decreases a specific body wall myosin heavy chain two-fold via a trans-acting mechanism. The concordant expression of GAPDH-2 with the myosins in wild-type will be analyzed in unc-52 to compare and test whether the trans-acting component also reduces body wall GAPDH expression. Twenty other uncoordinated mutants, defective in myofilament assembly, will be tested for deficiencies and electrophoretic variants in GAPDH and other actin-binding glycolytic enzymes. In this manner, the coordinate expression of these enzymes and/or their role in myofilament assembly will be defined.
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