Our long term goal is to understand both the biogenesis and turnover mechanisms involved in the homeostasis of peroxisomes, an essential subcellular organelle that is intimately involved in many metabolic pathways, particularly lipid metabolism. Impairment of peroxisome biogenesis is the underlying cause of many fatal and debilitating human peroxisome biogenesis disorders (PBDs). Peroxisomal matrix proteins with the peroxisomal targeting signals, PTS1 or PTS2, are recognized by receptors, Pex5 and Pex7, respectively. The PTS2 pathway also uses co-receptors. In previous work, we demonstrated the dynamic behavior of some PTS receptors/co-receptors during the matrix protein import cycle, showing that they shuttle to and from peroxisomes. Receptor/co-receptor recycling and degradation are dependent on mono- and poly-ubiquitination steps, with the latter process also engaging the ubiquitin proteasome system (UPS). Peroxisomal membrane protein (PMP) biogenesis is poorly understood, with mPTSs serving as signals for membrane targeting. Whether proteins are sorted to peroxisome membranes solely by their post-translation insertion into peroxisomes, or via the ER or possibly both, are hotly debated. There is also an ongoing debate regarding whether peroxisomes arise by growth and division or by de novo biogenesis. Pex19 is viewed as the mPTS receptor for the post-translational insertion of many PMPs into peroxisome membranes, but we found a role in the budding of pre-peroxisomal vesicles (ppVs) from the ER. Other proteins, such as Pex3, Pex16L and Pex25 also play a role in de novo peroxisome biogenesis, division and inheritance.
Aims1 and 2 of this proposal focus on the de novo biogenesis of peroxisomes investigating the roles of four proteins - Pex3, Pex16L, Pex19 and Pex25 in this process, including the assembly of redundant peroxisome division machineries that function in a spatiotemporally distinct manner to cause fission of tubular pre- peroxisomes at the ER or at pre-existing peroxisomes.
Aim1 asks how Pex19 and Pex3 perform a role in budding of ppVs from the ER, focusing on Pex19 domains involved in ppV budding, testing specific hypotheses regarding Pex3 and Pex19 functions in ppV budding, peroxisome division and inheritance and asking what other proteins are involved.
Aim2 centers on how Pex16-like (Pex16L) and Pex25 proteins play dual roles in biogenesis and division, with attention on the components of the redundant division machineries and their temporal action.
Aim3 addresses PTS receptor dynamics and its regulation by metabolic cues, focusing on Pex7, whose dynamics has not been studied to date.
Aim3 addresses the mechanism of Pex7 dynamics, asking whether Pex7 and cargo turnover are regulated by the UPS and metabolic cues. Answers to these questions will not only provide a mechanistic understanding of these processes but also shed light on human disease states because all the proteins we focus on are conserved in humans and mutated in PBDs.
Peroxisomes are essential subcellular compartments whose biogenesis is critical for human health and whose impairment is the cause of many fatal, costly and debilitating human diseases, collectively called peroxisome biogenesis disorders (PBDs). Understanding how these organelles are assembled intracellularly, how they import proteins across their membranes, how they divide and how they are inherited from mother to daughter cell is critical for a mechanistic understanding of a fundamentally important organelle but also provides information on how human mutations in conserved proteins cause PBDs. The research addresses some of the most challenging problems on this field.
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