Julia Kardon, PhD
Assistant Professor
Department of Biochemistry
Brandeis University
October 1, 2018
Control of the Mitochondrial Proteome: Activation, Repair, and Removal
Mitochondria are vitally important structures within a cell: they produce ATP, the currency of cellular energy, and are involved in cell metabolism. As cells age and change, mitochondria must adapt their proteins in order to continue to function properly. Not adapting can lead to disease. Dr. Karden and her lab examine how these proteins are remodeled or destroyed. They have determined that a specific protein, mtClpX, is necessary for the early stages of synthesis of heme (part of hemoglobin, which carries oxygen in the blood). She also determined that a mutation in mtClpX can lead to a build- up of heme, suggesting that mtClpX is also necessary for the breakdown of an enzyme used in the early stage of heme synthesis.
Mitochondria are centers of biosynthesis and respiration for the eukaryotic cell. To match the varied and dynamic metabolic needs of cells as they grow, develop, and age, mitochondria must maintain and control the activity and quality of their proteins. Failure to do so causes diverse neurodegenerative, metabolic, and developmental disorders and contribute to the distorted physiology of cancer cells. To accomplish this control, mitochondria have their own chaperones and proteases with which to remodel or demolish the structure of mitochondrial proteins. However, many of the proteins these machines act on, how their interaction is specified, and how the effect of their interaction is determined remains mysterious.
Mitochondrial chaperones and proteases are related to those of the mitochondrial ancestral bacterial endosymbiont; therefore, previous work in bacteria give clues to the types of functions these enzymes may enact in mitochondria. In bacteria, the protein unfoldase ClpX primarily acts on specific proteins for regulatory purposes, rather than acting to disassemble damaged proteins, suggesting that the mitochondrial ClpX homolog (mtClpX) also plays important regulatory roles. Large-scale genetic interaction maps in budding yeast indicated a connection between mtClpX function and heme biosynthesis. By metabolomic analysis, I determined that mtClpX stimulates the first step of heme biosynthesis, synthesis of 5-aminolevulinic acid (ALA). I found that mtClpX directly activates the ALA synthase (ALAS) by a novel, non-proteolytic activity: accelerating incorporation of its cofactor, pyridoxal phosphate (PLP).
mtClpX function in heme biosynthesis is broadly conserved; together with collaborators I found that mtClpX facilitates the heme-intensive process of red blood cell development (erythropoiesis) and that a familial mutation of mtClpX causes an erythropoietic disease. Surprisingly, this disease is characterized by toxic accumulation of heme caused by reduced mtClpX activity and overaccumulation of ALAS protein leading to precursors, suggesting that mtClpX conditionally directs degradation of ALAS as well. Work from other groups suggests that heme binding to ALAS may promote this degradation, providing an elegant feedback mechanism to coordinately promote and limit ALAS activity.
The discovery of mtClpX as an activator of ALAS posed an intriguing question: how does a protein unfoldase, whose basis mechanism is to pull apart the structure of other proteins, activate rather than inactivate an enzyme? I have determined that mtClpX pulls from the N-terminus of ALAS to unfold only a defined region that gates access to the active site, thus allowing cofactor entry and activation. My research group is currently working to determine how mtClpX unfolding of ALAS is limited, thus directing activation, and how these limiting signals may be bypassed to allow complete unfolding coupled to degradation of ALAS.