Animals, plants, fungi, and other eukaryotes assemble their respiratory apparatus from protein subunits made in the cytoplasm and protein subunits made inside mitochondria. We are trying to understand the basic mechanisms that allow this coordinated process to occur, since defects in this process affect the health and vitality of both plants and animals, including humans.

Oxygen is a very dangerous chemical. Cells that live in the presence of oxygen, and use it to burn their food must handle it very carefully. Interestingly, the apparatus that ultimately consumes oxygen in organelles called mitochondria is composed of proteins encoded by nuclear genes that are synthesized in the cytoplasm, and proteins encoded by genes in mitochondrial DNA that are synthesized by a separate genetic system inside the organelles. Defects in the complex interplay between these genetic systems in humans lead to disease and appear to play a significant role in the aging process, while variation in mitochondrial gene expression in maize and other plants influences male fertility and pathogen sensitivity.

We study the mechanisms employed to control mitochondrial protein synthesis in Baker's yeast (a model organism), and to target mitochondrially coded proteins to their proper destination within the organelle. During the past year our published work has focussed on nuclearly encoded components of mitochondrial ribosomes required for accurate initiation of protein synthesis. Mutations affecting the RNA sequence of the first ten codons of the mitochondrial gene COX2 strongly reduce translation of the mRNA. A dominant chromosomal mutation that suppresses these defects is an internal in-frame deletion of 67 codons from the gene YDR494w, encoding a 361 residue polypeptide with no similarity to proteins of known function. The product of this gene, now named RSM28, sedimented with the small subunit of mitochondrial ribosomes. Complete deletion of RSM28 caused only a modest decrease in respiratory growth, and enhanced the respiratory defect of the suppressible cox2 mutations. The rsm28 null mutation also reduced translation of an ARG8m reporter sequence inserted at the COX1, COX2, and COX3 mitochondrial loci. We tested the ability of RSM28-1 to suppress a variety of cox2 and cox3 mutations and found that initiation codon mutations in both genes were suppressed. Thus, Rsm28p is a dispensable small subunit mitochondrial ribosomal protein, previously undetected in systematic investigations of these ribosomes, with a positive role in translation of several mitochondrial mRNAs.

Our detailed examination of mitochondrial gene expression in yeast has revealed that organellar gene regulation is often at the level of protein synthesis instead of mRNA synthesis. And it has revealed that this regulatory mechanism plays an important role in targeting the newly synthesized mitochondrial gene products to sites where they can be efficiently assembled into respiratory complexes. These studies have taken advantage of the remarkably powerful genetic approaches that can be used to study yeast, principally replacement of genes in both nuclear and mitochondrial chromosomes, but not in more complex organisms such as plants and animals. Thus our exploratory work has provided a trailblazing look at the questions that should be studied in thes e more complex organisms, as well as helping to elucidate basic mechanisms and potential drug targets in fungi.

Basic biological research on cellular mechanisms

• Dr. Heather Fiumera
• Dr. Timothy Ellis
• Dr. Dieter Soll
• Dr. Nathalie Bonnefoy
• Dr. Thomas Mason (retired)