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 and others 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 peroxisomal membrane targeting. Whether PMPs are sorted to peroxisome membranes solely by their post- translation insertion into peroxisomes, or via the endoplasmic reticulum (ER), or possibly both, are ongoing debates. There is also considerable discussion regarding the relative contributions of the growth and division versus de novo biogenesis models for peroxisome biogenesis. Pex19 has been viewed as the mPTS receptor for the post-translational insertion of many PMPs into peroxisome membranes, but we found new roles for Pex3 and Pex19 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 from the ER investigating the roles of four proteins - Pex3, Pex16L, Pex19 and Pex25 in this process, including the assembly of two peroxisome membrane fission machineries that function in a spatiotemporally distinct manner to cause fission of tubular pre-peroxisomes at the ER and later to divide mature 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 budding of two classes of ppVs and in peroxisome inheritance, as well as the underlying mechanisms involved.
Aim2 tests specific hypotheses regarding the roles of Pex16-like (Pex16L) and Pex25 proteins in either ppV budding and/or peroxisome division, with attention on the components of the fission 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 also addresses 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.

Public Health Relevance

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 cells 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 in this field.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
2R01DK041737-25A1
Application #
8815471
Study Section
Membrane Biology and Protein Processing (MBPP)
Program Officer
Haft, Carol R
Project Start
1990-05-10
Project End
2018-08-31
Budget Start
2014-09-25
Budget End
2015-08-31
Support Year
25
Fiscal Year
2014
Total Cost
$491,353
Indirect Cost
$166,367
Name
University of California San Diego
Department
Biology
Type
Schools of Arts and Sciences
DUNS #
804355790
City
La Jolla
State
CA
Country
United States
Zip Code
92093
Katarzyna, Zientara-Rytter; Suresh, Subramani (2016) Autophagic degradation of peroxisomes in mammals. Biochem Soc Trans 44:431-40
Farré, Jean-Claude; Subramani, Suresh (2016) Mechanistic insights into selective autophagy pathways: lessons from yeast. Nat Rev Mol Cell Biol 17:537-52
Agrawal, Gaurav; Fassas, Scott N; Xia, Zhi-Jie et al. (2016) Distinct requirements for intra-ER sorting and budding of peroxisomal membrane proteins from the ER. J Cell Biol 212:335-48
Agrawal, Gaurav; Subramani, Suresh (2016) De novo peroxisome biogenesis: Evolving concepts and conundrums. Biochim Biophys Acta 1863:892-901
Subramani, Suresh (2015) A mammalian pexophagy target. Nat Cell Biol 17:1371-3
Hagstrom, Danielle; Ma, Changle (2014) Biochemically characterizing the subcellular localization of peroxisomal proteins by fractionation, protease protection, and carbonate extraction. Methods Mol Biol 1163:175-81
Hagstrom, Danielle; Ma, Changle; Guha-Polley, Soumi et al. (2014) The unique degradation pathway of the PTS2 receptor, Pex7, is dependent on the PTS receptor/coreceptor, Pex5 and Pex20. Mol Biol Cell 25:2634-43
Liu, Xueqian; Subramani, Suresh (2013) Unique requirements for mono- and polyubiquitination of the peroxisomal targeting signal co-receptor, Pex20. J Biol Chem 288:7230-40
Ma, Changle; Hagstrom, Danielle; Polley, Soumi Guha et al. (2013) Redox-regulated cargo binding and release by the peroxisomal targeting signal receptor, Pex5. J Biol Chem 288:27220-31
Liu, Xueqian; Ma, Changle; Subramani, Suresh (2012) Recent advances in peroxisomal matrix protein import. Curr Opin Cell Biol 24:484-9

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