scholarly journals Peroxisomal Peripheral Membrane Protein YlInp1p Is Required for Peroxisome Inheritance and Influences the Dimorphic Transition in the Yeast Yarrowia lipolytica

2007 ◽  
Vol 6 (9) ◽  
pp. 1528-1537 ◽  
Author(s):  
Jinlan Chang ◽  
Andrei Fagarasanu ◽  
Richard A. Rachubinski
Keyword(s):  
Membrane Protein ◽  
Yarrowia Lipolytica ◽  
Mother Cell ◽  
Molecular Mechanisms ◽  
Peroxisomal Membrane ◽  
Mycelial Form ◽  
Dimorphic Transition ◽  
Dimorphic Yeast ◽  
In Cells

ABSTRACT Eukaryotic cells have evolved molecular mechanisms to ensure the faithful inheritance of organelles by daughter cells in order to maintain the benefits afforded by the compartmentalization of biochemical functions. Little is known about the inheritance of peroxisomes, organelles of lipid metabolism. We have analyzed peroxisome dynamics and inheritance in the dimorphic yeast Yarrowia lipolytica. Most peroxisomes are anchored at the periphery of cells of Y. lipolytica. In vivo video microscopy showed that at cell division, approximately half of the anchored peroxisomes in the mother cell are dislodged individually from their static positions and transported to the bud. Peroxisome motility is dependent on the actin cytoskeleton. YlInp1p is a peripheral peroxisomal membrane protein that affects the partitioning of peroxisomes between mother cell and bud in Y. lipolytica. In cells lacking YlInp1p, most peroxisomes were transferred to the bud, with only a few remaining in the mother cell, while in cells overexpressing YlInp1p, peroxisomes were preferentially retained in the mother cell, resulting in buds nearly devoid of peroxisomes. Our results are consistent with a role for YlInp1p in anchoring peroxisomes in cells. YlInp1p has a role in the dimorphic transition in Y. lipolytica, as cells lacking the YlINP1 gene more readily convert from the yeast to the mycelial form in oleic acid-containing medium, the metabolism of which requires peroxisomal activity, than does the wild-type strain. This study reports the first analysis of organelle inheritance in a true dimorphic yeast and identifies the first protein required for peroxisome inheritance in Y. lipolytica.

scholarly journals Inp1p is a peroxisomal membrane protein required for peroxisome inheritance in Saccharomyces cerevisiae

2005 ◽  
Vol 169 (5) ◽  
pp. 765-775 ◽  
Author(s):  
Monica Fagarasanu ◽  
Andrei Fagarasanu ◽  
Yuen Yi C. Tam ◽  
John D. Aitchison ◽  
Richard A. Rachubinski
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Division ◽  
Membrane Protein ◽  
Molecular Mechanisms ◽  
Cell Cortex ◽  
Cell Periphery ◽  
Peroxisomal Membrane ◽  
Mother Cells ◽  
Peroxisome Division

Cells have evolved molecular mechanisms for the efficient transmission of organelles during cell division. Little is known about how peroxisomes are inherited. Inp1p is a peripheral membrane protein of peroxisomes of Saccharomyces cerevisiae that affects both the morphology of peroxisomes and their partitioning during cell division. In vivo 4-dimensional video microscopy showed an inability of mother cells to retain a subset of peroxisomes in dividing cells lacking the INP1 gene, whereas cells overexpressing INP1 exhibited immobilized peroxisomes that failed to be partitioned to the bud. Overproduced Inp1p localized to both peroxisomes and the cell cortex, supporting an interaction of Inp1p with specific structures lining the cell periphery. The levels of Inp1p vary with the cell cycle. Inp1p binds Pex25p, Pex30p, and Vps1p, which have been implicated in controlling peroxisome division. Our findings are consistent with Inp1p acting as a factor that retains peroxisomes in cells and controls peroxisome division. Inp1p is the first peroxisomal protein directly implicated in peroxisome inheritance.

scholarly journals miR-182-5p and miR-378a-3p regulate ferroptosis in I/R-induced renal injury

2020 ◽  
Vol 11 (10) ◽  
Author(s):  
Chenguang Ding ◽  
Xiaoming Ding ◽  
Jin Zheng ◽  
Bo Wang ◽  
Yang Li ◽  
...  
Keyword(s):  
Cell Death ◽  
Renal Injury ◽  
Molecular Mechanisms ◽  
Ischemia Reperfusion ◽  
Kidney Injury ◽  
Luciferase Reporter ◽  
Renal Tubular Cell ◽  
In Cells

Abstract Renal tubular cell death is the key factor of the pathogenesis of ischemia/reperfusion (I/R) kidney injury. Ferroptosis is a type of regulated cell death (RCD) found in various diseases. However, the underlying molecular mechanisms related to ferroptosis in renal I/R injury remain unclear. In the present study, we investigated the regulatory role of microRNAs on ferroptosis in I/R-induced renal injury. We established the I/R-induced renal injury model in rats, and H/R induced HK-2 cells injury in vitro. CCK-8 was used to measure cell viability. Fe2+ and ROS levels were assayed to evaluate the activation of ferroptosis. We performed RNA sequencing to profile the miRNAs expression in H/R-induced injury and ferroptosis. Western blot analysis was used to detect the protein expression. qRT-PCR was used to detect the mRNA and miRNA levels in cells and tissues. We further used luciferase reporter assay to verify the direct targeting effect of miRNA. We found that ischemia/reperfusion-induced ferroptosis in rat’s kidney. We identified that miR-182-5p and miR-378a-3p were upregulated in the ferroptosis and H/R-induced injury, and correlates reversely with glutathione peroxidases 4 (GPX4) and solute carrier family 7 member 11 (SLC7A11) expression in renal I/R injury tissues, respectively. In vitro studies showed that miR-182-5p and miR-378a-3p induced ferroptosis in cells. We further found that miR-182-5p and miR-378a-3p regulated the expression of GPX4 and SLC7A11 negatively by directly binding to the 3′UTR of GPX4 and SLC7A11 mRNA. In vivo study showed that silencing miR-182-5p and miR-378a-3p alleviated the I/R-induced renal injury in rats. In conclusion, we demonstrated that I/R induced upregulation of miR-182-5p and miR-378a-3p, leading to activation of ferroptosis in renal injury through downregulation of GPX4 and SLC7A11.

scholarly journals α5β1 integrin recycling promotes Arp2/3-independent cancer cell invasion via the formin FHOD3

2015 ◽  
Vol 210 (6) ◽  
pp. 1013-1031 ◽  
Author(s):  
Nikki R. Paul ◽  
Jennifer L. Allen ◽  
Anna Chapman ◽  
Maria Morlan-Mairal ◽  
Egor Zindy ◽  
...  
Keyword(s):  
Cancer Cells ◽  
Cell Invasion ◽  
Molecular Mechanisms ◽  
Cancer Metastasis ◽  
Cancer Cell Invasion ◽  
Coupling Protein ◽  
Α5β1 Integrin ◽  
In Cells

Invasive migration in 3D extracellular matrix (ECM) is crucial to cancer metastasis, yet little is known of the molecular mechanisms that drive reorganization of the cytoskeleton as cancer cells disseminate in vivo. 2D Rac-driven lamellipodial migration is well understood, but how these features apply to 3D migration is not clear. We find that lamellipodia-like protrusions and retrograde actin flow are indeed observed in cells moving in 3D ECM. However, Rab-coupling protein (RCP)-driven endocytic recycling of α5β1 integrin enhances invasive migration of cancer cells into fibronectin-rich 3D ECM, driven by RhoA and filopodial spike-based protrusions, not lamellipodia. Furthermore, we show that actin spike protrusions are Arp2/3-independent. Dynamic actin spike assembly in cells invading in vitro and in vivo is regulated by Formin homology-2 domain containing 3 (FHOD3), which is activated by RhoA/ROCK, establishing a novel mechanism through which the RCP–α5β1 pathway reprograms the actin cytoskeleton to promote invasive migration and local invasion in vivo.

scholarly journals Regulating peroxisome–ER contacts via the ACBD5-VAPB tether by FFAT motif phosphorylation and GSK3β

2022 ◽  
Vol 221 (3) ◽  
Author(s):  
Suzan Kors ◽  
Christian Hacker ◽  
Chloe Bolton ◽  
Renate Maier ◽  
Lena Reimann ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Mammalian Cells ◽  
Molecular Mechanisms ◽  
Glycogen Synthase ◽  
Current Model ◽  
Protein Domain ◽  
Major Sperm Protein ◽  
Peroxisomal Membrane ◽  
Membrane Contact Sites ◽  
Contact Sites

Peroxisomes and the endoplasmic reticulum (ER) cooperate in cellular lipid metabolism. They form membrane contacts through interaction of the peroxisomal membrane protein ACBD5 (acyl-coenzyme A–binding domain protein 5) and the ER-resident protein VAPB (vesicle-associated membrane protein–associated protein B). ACBD5 binds to the major sperm protein domain of VAPB via its FFAT-like (two phenylalanines [FF] in an acidic tract) motif. However, molecular mechanisms, which regulate formation of these membrane contact sites, are unknown. Here, we reveal that peroxisome–ER associations via the ACBD5-VAPB tether are regulated by phosphorylation. We show that ACBD5-VAPB binding is phosphatase-sensitive and identify phosphorylation sites in the flanking regions and core of the FFAT-like motif, which alter interaction with VAPB—and thus peroxisome–ER contact sites—differently. Moreover, we demonstrate that GSK3β (glycogen synthase kinase-3 β) regulates this interaction. Our findings reveal for the first time a molecular mechanism for the regulation of peroxisome–ER contacts in mammalian cells and expand the current model of FFAT motifs and VAP interaction.

scholarly journals Formation of the peroxisome lumen is abolished by loss of Pichia pastoris Pas7p, a zinc-binding integral membrane protein of the peroxisome.

1995 ◽  
Vol 15 (11) ◽  
pp. 6406-6419 ◽  
Author(s):  
J E Kalish ◽  
C Theda ◽  
J C Morrell ◽  
J M Berg ◽  
S J Gould
Keyword(s):  
Pichia Pastoris ◽  
Membrane Protein ◽  
Protein Translocation ◽  
Point Mutations ◽  
Integral Membrane Protein ◽  
Zinc Binding ◽  
Peroxisomal Membrane ◽  
Peroxisomal Targeting ◽  
Targeting Signal

We have cloned and sequenced PAS7, a gene required for peroxisome assembly in the yeast Pichia pastoris. The product of this gene, Pas7p, is a member of the C3HC4 superfamily of zinc-binding proteins. Point mutations that alter conserved residues of the C3HC4 motif abolish PAS7 activity and reduce zinc binding, suggesting that Pas7p binds zinc in vivo and that zinc binding is essential for PAS7 function. As with most pas mutants, pas7 cells exhibit a pronounced deficiency in import of peroxisomal matrix proteins that contain either the type 1 peroxisomal targeting signal (PTS1) or the type 2 PTS (PTS2). However, while other yeast and mammalian pas mutants accumulate ovoid, vesicular peroxisomal intermediates, loss of Pas7p leads to accumulation of membrane sheets and vesicles which lack a recognizable lumen. Thus, Pas7p appears to be essential for protein translocation into peroxisomes as well as formation of the lumen of the organelle. Consistent with these data, we find that Pas7p is an integral peroxisomal membrane protein which is entirely resistant to exogenous protease and thus appears to reside completely within the peroxisome. Our observations suggest that the function of Pas7p defines a previously unrecognized step in peroxisome assembly: formation of the peroxisome lumen. Furthermore, because the peroxisomal intermediates in the pas7 delta mutant proliferate in response to peroxisome-inducing environmental conditions, we conclude that Pas7p is not required for peroxisome proliferation.

scholarly journals Saccharomyces cerevisiae PTS1 Receptor Pex5p Interacts with the SH3 Domain of the Peroxisomal Membrane Protein Pex13p in an Unconventional, Non-PXXP–related Manner

2000 ◽  
Vol 11 (11) ◽  
pp. 3963-3976 ◽  
Author(s):  
Gina Bottger ◽  
Phil Barnett ◽  
AndréT. J. Klein ◽  
Astrid Kragt ◽  
Henk F. Tabak ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Membrane Protein ◽  
Protein Import ◽  
Sh3 Domain ◽  
Site Directed Mutagenesis ◽  
Binding Motif ◽  
Binding Studies ◽  
Peroxisomal Membrane ◽  
Yeast Saccharomyces Cerevisiae

A number of peroxisome-associated proteins have been described that are involved in the import of proteins into peroxisomes, among which is the receptor for peroxisomal targeting signal 1 (PTS1) proteins Pex5p, the integral membrane protein Pex13p, which contains an Src homology 3 (SH3) domain, and the peripheral membrane protein Pex14p. In the yeast Saccharomyces cerevisiae, both Pex5p and Pex14p are able to bind Pex13p via its SH3 domain. Pex14p contains the classical SH3 binding motif PXXP, whereas this sequence is absent in Pex5p. Mutation of the conserved tryptophan in the PXXP binding pocket of Pex13-SH3 abolished interaction with Pex14p, but did not affect interaction with Pex5p, suggesting that Pex14p is the classical SH3 domain ligand and that Pex5p binds the SH3 domain in an alternative way. To identify the SH3 binding site in Pex5p, we screened a randomly mutagenized PEX5 library for loss of interaction with Pex13-SH3. Such mutations were all located in a small region in the N-terminal half of Pex5p. One of the altered residues (F208) was part of the sequence W204XXQF208, that is conserved between Pex5 proteins of different species. Site-directed mutagenesis of Trp204 confirmed the essential role of this motif in recognition of the SH3 domain. The Pex5p mutants could only partially restore PTS1-protein import in pex5Δ cells in vivo. In vitro binding studies showed that these Pex5p mutants failed to interact with Pex13-SH3 in the absence of Pex14p, but regained their ability to bind in the presence of Pex14p, suggesting the formation of a heterotrimeric complex consisting of Pex5p, Pex14p, and Pex13-SH3. In vivo, these Pex5p mutants, like wild-type Pex5p, were still found to be associated with peroxisomes. Taken together, this indicates that in the absence of Pex13-SH3 interaction, other protein(s) is able to bind Pex5p at the peroxisome; Pex14p is a likely candidate for this function.

scholarly journals Regulating peroxisome-ER contacts via the ACBD5-VAPB tether by FFAT motif phosphorylation and GSK3β

2021 ◽  
Author(s):  
Suzan Kors ◽  
Christian Hacker ◽  
Chloe Bolton ◽  
Renate Maier ◽  
Lena Reimann ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Mammalian Cells ◽  
Molecular Mechanisms ◽  
Glycogen Synthase ◽  
Current Model ◽  
Protein Domain ◽  
Major Sperm Protein ◽  
Peroxisomal Membrane ◽  
Membrane Contact Sites ◽  
Contact Sites

Peroxisomes and the endoplasmic reticulum (ER) cooperate in cellular lipid metabolism. They form membrane contacts through interaction of the peroxisomal membrane protein ACBD5 [acyl-coenzyme A-binding domain protein 5] and the ER-resident protein VAPB [vesicle-associated membrane protein-associated protein B]. ACBD5 binds to the major sperm protein domain of VAPB via its FFAT-like [two phenylalanines (FF) in an acidic tract] motif. However, molecular mechanisms, which regulate formation of these membrane contact sites, are unknown. Here, we reveal that peroxisome-ER associations via the ACBD5-VAPB tether are regulated by phosphorylation. We show that ACBD5-VAPB binding is phosphatase-sensitive and identify phosphorylation sites in the flanking regions and core of the FFAT-like motif, which alter interaction with VAPB and thus, peroxisome-ER contact sites differently. Moreover, we demonstrate that GSK3β [glycogen synthase kinase-3 beta] regulates this interaction. Our findings reveal for the first time a molecular mechanism for the regulation of peroxisome-ER contacts in mammalian cells and expand the current model of FFAT motifs and VAP interaction.

scholarly journals A phosphatidylinositol/phosphatidylcholine transfer protein is required for differentiation of the dimorphic yeast Yarrowia lipolytica from the yeast to the mycelial form.

1994 ◽  
Vol 125 (1) ◽  
pp. 113-127 ◽  
Author(s):  
M C Lopez ◽  
J M Nicaud ◽  
H B Skinner ◽  
C Vergnolle ◽  
J C Kader ◽  
...  
Keyword(s):  
Yarrowia Lipolytica ◽  
Secretory Pathway ◽  
Stationary Growth Phase ◽  
Mycelial Form ◽  
Amino Terminal ◽  
Transfer Protein ◽  
Dimorphic Yeast ◽  
Phosphatidylcholine Transfer Protein ◽  
Yeast Yarrowia Lipolytica

The SEC14SC gene encodes the phosphatidylinositol/phosphatidylcholine transfer protein (PI/PC-TP) of Saccharomyces cerevisiae. The SEC14SC gene product (SEC14pSC) is associated with the Golgi complex as a peripheral membrane protein and plays an essential role in stimulating Golgi secretory function. We report the characterization of SEC14YL, the structural gene for the PI/PC-TP of the dimorphic yeast Yarrowia lipolytica. SEC14YL encodes a primary translation product (SEC14YL) that is predicted to be a 497-residue polypeptide of which the amino-terminal 300 residues are highly homologous to the entire SEC14pSC, and the carboxyl-terminal 197 residues define a dispensible domain that is not homologous to any known protein. In a manner analogous to the case for SEC14pSC, SEC14pYL localizes to punctate cytoplasmic structures in Y. lipolytica that likely represent Golgi bodies. However, SEC14pYL is neither required for the viability of Y. lipolytica nor is it required for secretory pathway function in this organism. This nonessentiality of SEC14pYL for growth and secretion is probably not the consequence of a second PI/PC-TP activity in Y. lipolytica as cell-free lysates prepared from delta sec14YL strains are devoid of measurable PI/PC-TP activity in vitro. Phenotypic analyses demonstrate that SEC14pYL dysfunction results in the inability of Y. lipolytica to undergo the characteristic dimorphic transition from the yeast to the mycelial form that typifies this species. Rather, delta sec14YL mutants form aberrant pseudomycelial structures as cells enter stationary growth phase. The collective data indicate a role for SEC14pYL in promoting the differentiation of Y. lipolytica cells from yeast to mycelia, and demonstrate that PI/PC-TP function is utilized in diverse ways by different organisms.

scholarly journals Peroxisome assembly: matrix and membrane protein biogenesis

2011 ◽  
Vol 193 (1) ◽  
pp. 7-16 ◽  
Author(s):  
Changle Ma ◽  
Gaurav Agrawal ◽  
Suresh Subramani
Keyword(s):  
Membrane Protein ◽  
Matrix Protein ◽  
Molecular Mechanisms ◽  
Protein Import ◽  
Protein Translocation ◽  
Peroxisomal Membrane ◽  
Peroxisome Biogenesis Disorders ◽  
Molecular Events ◽  
Protein Biogenesis ◽  
Shed Light

The biogenesis of peroxisomal matrix and membrane proteins is substantially different from the biogenesis of proteins of other subcellular compartments, such as mitochondria and chloroplasts, that are of endosymbiotic origin. Proteins are targeted to the peroxisome matrix through interactions between specific targeting sequences and receptor proteins, followed by protein translocation across the peroxisomal membrane. Recent advances have shed light on the nature of the peroxisomal translocon in matrix protein import and the molecular mechanisms of receptor recycling. Furthermore, the endoplasmic reticulum has been shown to play an important role in peroxisomal membrane protein biogenesis. Defining the molecular events in peroxisome assembly may enhance our understanding of the etiology of human peroxisome biogenesis disorders.

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