Examining membraneless compartmentalization in the nucleolus

Cells carry out hundreds of functions simultaneously, and compartmentalization allows these functions to occur efficiently. Some compartmentalization occurs via the use of membranes, and membrane-bound organelles include the mitochondria, nucleus, and endoplasmic reticulum. Other compartments form by phase separation, reminiscent of the separation of oil and water. A special region in the nucleus called the nucleolus is one such compartment. The nucleolus is the region of the cell that is dedicated to making ribosomes in a process called ribosome biogenesis. Ribosomes interpret genetic code to make all the proteins in the cell. Most of what we know about the phase separation of the nucleolus comes from studies involving only three proteins representing three distinct domains of the nucleolus, but hundreds of proteins are involved in ribosome biogenesis and localize to the nucleolus, many of which are suspected of phase separation.

In this thesis, I expanded support for a hypothesis that proposed that nucleolar proteins are localized into phase-separated compartments based on their order of utilization in ribosome biogenesis, and I added to the roster of proteins with preferred nucleolar domains. I showed that nucleolar morphology changes under conditions that inhibit ribosome biogenesis, and that compartmentalization depends on ribosome synthesis. I showed how nucleolar proteins find their preferred compartment is not identical to phase separation in other non-membrane bound organelles. I also showed that some proteins must bind to another protein already in a compartment to stay in that specific nucleolar compartment.

Nucleolar activity and structure both change in response to neurodegenerative diseases, cancers, viral infections, and changes in the cell cycle. More generally, aberrant-protein phase separation is the hallmark of neurodegenerative diseases like Alzheimer’s and Parkinson’s diseases. Understanding protein phase separation in the nucleolus and elsewhere in the cell is critical for disease management and the discovery of new treatments.