Studies were conducted in the areas of tRNA transport and processing, the molecular biology of the Alu sequence, and the mechanism of mRNA transport. Based on an analysis of some 30 in vitro-generated point mutants in the human tRNA-met-i gene, we have shown that mutations which perturb nuclear tRNA transport are clustered in the T and D loops of the molecule, the most conserved portion of the tRNA molecule. All tRNA species defective in transport were also found to be inefficiently processed, suggesting functional linkage of the two systems. The 5' and 3' processing enzymes, the enzyme involved in tRNA maturation in eukaryotes were purified to homogeneity. The 3' processing enzyme is a single polypeptide which cuts the 3' trailer off the pre-tRNA species only after prior removal of the 5' leader. The 5' processing enzyme is a very complex species of about 400,000 in weight consisting of some 15 polypeptides. Recent electron microscopic analysis shows the enzyme to be a ring particle. Studies on the biology of the Alu sequence in mouse have yielded a new conversion pathway for a small cellular RNA of 135 nt. This B1 RNA lies complementary to an intron in the mouse alpha-fetoprotein gene. We have shown that this new RNA is transcribed, processed specifically, transported, and packaged in the cytoplasm of cells into a novel ribonucleoprotein particle. The levels of this species vary between different mouse tissues suggesting a role in tissue-specific gene expression. It may serve as the first example of the way a cell handles a natural anti-sense RNA. Studies have been initiated on defining the mechanism of mRNA transport. We have made the unexpected observation that mRNA transcription and transport can be uncoupled in the X. laevis oocyte, providing a fruitful system for the study of this critical process.
