Cellular Control of Endoplasmic Reticulum Biogenesis

Eukaryotic cells are composed of a basic set of organelle 'building blocks' that include (among others) the nucleus, endoplasmic reticulum (ER), Golgi apparatus, lysosomes, mitochondria, peroxisomes and, in photosynthetic organisms, chloroplasts. This relatively small number of separate cellular components belies the amazing diversity of cell structure and function that exists in eukaryotic organisms. In each case, the specialization of cell structure and function is mirrored by changes in organelle composition. Thus, a key feature of cell biology is the regulation of subcellular organization, that is, the regulation of which organelles are present and their size, composition, location, number, and life spans. Surprisingly, given the importance of this regulation, there is not yet a single example in which the molecular nature of this regulation is well understood.

The experiments that will be performed explore this question by focusing on the regulation of endoplasmic reticulum structure and function in the yeast, Saccharomyces cerevisiae. As in all other cell-types examined, the organization of the ER in yeast is sensitive to the levels of a subset of ER proteins. One of these proteins, HMG-CoA reductase, catalyzes the first committed step in sterol and isoprene biosynthesis. In yeast, expression of increased levels of HMG-CoA reductase induces assembly of specialized regions of ER termed karmellae, consisting of stacks of paired smooth membranes that are closely associated with the nucleus. The ability to control karmella assembly by merely changing the levels of a single protein provides a unique opportunity to explore the molecular mechanisms by which cells increase biogenesis of a particular ER domain when dictated by physiological demands.

To uncover these mechanisms, Dr. Wright will use a genetic approach that takes advantage of new resources available as a result of completion of the Yeast Genome Project. Specifically, she will use a population genetic approach to identify deletion mutants that display defects in growth rate when they assemble karmellae. Coupled with information from a Two-Hybrid approach to identify gene products that interact with HMG-CoA reductase, this approach should reveal genes that have important roles in assembly of karmellae. The analysis of karmella assembly mutants will be guided by in vivo time-lapse light microscopy and electron microscopic analysis of karmella assembly. Completion of these experiments should uncover basic features of the communication network that reports and regulates ER function coordinately with changing physiological demands. Such knowledge will have specific application to understanding the cellular regulation of ER structure and function, but should also provide general insights concerning the cellular regulation of organelle biogenesis.