Pex14p phosphorylation regulates peroxisome import

Peroxisomal matrix proteins are imported into peroxisomes via membrane-bound docking/translocation machinery. One central component of this machinery is Pex14p, a peroxisomal membrane protein involved in the docking of Pex5p, the receptor for peroxisome targeting signal type 1 (PTS1). Studies in several yeast species have shown that Pex14p is phosphorylated in vivo, whereas no function has been assigned to Pex14p phosphorylation in yeast and mammalian cells. Here, we investigated peroxisomal protein import and its dynamics in mitotic mammalian cells. In mitotically arrested cells, Pex14p is phosphorylated at Ser-232, resulting in a lower import efficiency of catalase, but not the majority of proteins including canonical PTS1 proteins. Conformational change induced by the mitotic phosphorylation of Pex14p more likely increases homomeric interacting affinity and suppresses topological change of its N-terminal part, thereby giving rise to the retardation of Pex5p export in mitotic cells. Taken together, these data show that mitotic phosphorylation of Pex14p and consequent suppression of catalase import are a mechanism of protecting DNA upon nuclear envelope breakdown at mitosis

Asymmetric Distribution of Plasmalogens and Their Roles—A Mini Review

Plasmalogens are a unique family of cellular glycerophospholipids that contain a vinyl-ether bond. The synthesis of plasmalogens is initiated in peroxisomes and completed in the endoplasmic reticulum. Plasmalogens are transported to the post-Golgi compartment, including endosomes and plasma membranes, in a manner dependent on ATP, but not vesicular transport. Plasmalogens are preferentially localized in the inner leaflet of the plasma membrane in a manner dependent on P4-type ATPase ATP8B2, that associates with the CDC50 subunit. Plasmalogen biosynthesis is spatiotemporally regulated by a feedback mechanism that senses the amount of plasmalogens in the inner leaflet of the plasma membrane and controls the stability of fatty acyl-CoA reductase 1 (FAR1), the rate-limiting enzyme for plasmalogen biosynthesis. The physiological consequences of such asymmetric localization and homeostasis of plasmalogens are discussed in this review

Dataset for: Plasmalogen homeostasis: regulation of plasmalogen biosynthesis and its physiological consequence in mammals

Plasmalogens, mostly ethanolamine-containing alkenyl ether phospholipids, are a major sub-class of glycerophospholipids. Plasmalogen synthesis is initiated in peroxisomes and completed in endoplasmic reticulum. The absence of plasmalogens in several organs of peroxisome biogenesis-defective patients suggests that<i> de novo</i> synthesis of plasmalogens plays a pivotal role in the plasmalogen homeostasis in tissues. Plasmalogen synthesis is regulated by modulating the stability of fatty acyl-CoA reductase 1 on peroxisomal membrane, a rate-limiting enzyme in plasmalogen synthesis, by sensing plasmalogens in the inner leaflet of plasma membranes. Dysregulation of plasmalogen homeostasis impairs cholesterol biosynthesis by altering the stability of squalene monooxygenase, a key enzyme in the cholesterol biosynthesis, implying physiological consequences of plasmalogen homeostasis in cholesterol metabolism in cells and organs such as liver

Pex11mediates peroxisomal proliferation by promoting deformation of the lipid membrane

Pex11p family proteins are key players in peroxisomal fission, but their molecular mechanisms remains mostly unknown. In the present study, overexpression of Pex11pβ caused substantial vesiculation of peroxisomes in mammalian cells. This vesicle formation was dependent on dynamin-like protein 1 (DLP1) and mitochondrial fission factor (Mff), as knockdown of these proteins diminished peroxisomal fission after Pex11pβ overexpression. The fission-deficient peroxisomes exhibited an elongated morphology, and peroxisomal marker proteins, such as Pex14p or matrix proteins harboring peroxisomal targeting signal 1, were discernible in a segmented staining pattern, like beads on a string. Endogenous Pex11pβ was also distributed a striped pattern, but which was not coincide with Pex14p and PTS1 matrix proteins. Altered morphology of the lipid membrane was observed when recombinant Pex11p proteins were introduced into proteo-liposomes. Constriction of proteo-liposomes was observed under confocal microscopy and electron microscopy, and the reconstituted Pex11pβ protein localized to the membrane constriction site. Introducing point mutations into the N-terminal amphiphathic helix of Pex11pβ strongly reduced peroxisomal fission, and decreased the oligomer formation. These results suggest that Pex11p contributes to the morphogenesis of the peroxisomal membrane, which is required for subsequent fission by DLP1

Mff functions with Pex11pβ and DLP1 in peroxisomal fission

Summary Peroxisomal division comprises three steps: elongation, constriction, and fission. Translocation of dynamin-like protein 1 (DLP1), a member of the large GTPase family, from the cytosol to peroxisomes is a prerequisite for membrane fission; however, the molecular machinery for peroxisomal targeting of DLP1 remains unclear. This study investigated whether mitochondrial fission factor (Mff), which targets DLP1 to mitochondria, may also recruit DLP1 to peroxisomes. Results show that endogenous Mff is localized to peroxisomes, especially at the membrane-constricted regions of elongated peroxisomes, in addition to mitochondria. Knockdown of MFF abrogates the fission stage of peroxisomal division and is associated with failure to recruit DLP1 to peroxisomes, while ectopic expression of MFF increases the peroxisomal targeting of DLP1. Co-expression of MFF and PEX11β, the latter being a key player in peroxisomal elongation, increases peroxisome abundance. Overexpression of MFF also increases the interaction between DLP1 and Pex11pβ, which knockdown of MFF, but not Fis1, abolishes. Moreover, results show that Pex11pβ interacts with Mff in a DLP1-dependent manner. In conclusion, Mff contributes to the peroxisomal targeting of DLP1 and plays a key role in the fission of the peroxisomal membrane by acting in concert with Pex11pβ and DLP1

Mammalian Homologue NME3 of DYNAMO1 Regulates Peroxisome Division

Peroxisomes proliferate by sequential processes comprising elongation, constriction, and scission of peroxisomal membrane. It is known that the constriction step is mediated by a GTPase named dynamin-like protein 1 (DLP1) upon efficient loading of GTP. However, mechanism of fuelling GTP to DLP1 remains unknown in mammals. We earlier show that nucleoside diphosphate (NDP) kinase-like protein, termed dynamin-based ring motive-force organizer 1 (DYNAMO1), generates GTP for DLP1 in a red alga, Cyanidioschyzon merolae. In the present study, we identified that nucleoside diphosphate kinase 3 (NME3), a mammalian homologue of DYNAMO1, localizes to peroxisomes. Elongated peroxisomes were observed in cells with suppressed expression of NME3 and fibroblasts from a patient lacking NME3 due to the homozygous mutation at the initiation codon of NME3. Peroxisomes proliferated by elevation of NME3 upon silencing the expression of ATPase family AAA domain containing 1, ATAD1. In the wild-type cells expressing catalytically-inactive NME3, peroxisomes were elongated. These results suggest that NME3 plays an important role in peroxisome division in a manner dependent on its NDP kinase activity. Moreover, the impairment of peroxisome division reduces the level of ether-linked glycerophospholipids, ethanolamine plasmalogens, implying the physiological importance of regulation of peroxisome morphology

Naïve-like conversion enhances the difference in innate in vitro differentiation capacity between rabbit ES cells and iPS cells

Quality evaluation of pluripotent stem cells using appropriate animal models needs to be improved for human regenerative medicine. Previously, we demonstrated that although the in vitro neural differentiating capacity of rabbit induced pluripotent stem cells (iPSCs) can be mitigated by improving their baseline level of pluripotency, i.e., by converting them into the so-called “naïve-like” state, the effect after such conversion of rabbit embryonic stem cells (ESCs) remains to be elucidated. Here we found that naïve-like conversion enhanced the differences in innate in vitro differentiation capacity between ESCs and iPSCs. Naïve-like rabbit ESCs exhibited several features indicating pluripotency, including the capacity for teratoma formation. They differentiated into mature oligodendrocytes much more effectively (3.3–7.2 times) than naïve-like iPSCs. This suggests an inherent variation in differentiation potential in vitro among PSC lines. When naïve-like ESCs were injected into preimplantation rabbit embryos, although they contributed efficiently to forming the inner cell mass of blastocysts, no chimeric pups were obtained. Thus, in vitro neural differentiation following naïve-like conversion is a promising option for determining the quality of PSCs without the need to demonstrate chimeric contribution. These results provide an opportunity to evaluate which pluripotent stem cells or treatments are best suited for therapeutic use