Ida Van Der Klei - Academia.edu (original) (raw)

Papers by Ida Van Der Klei

Research paper thumbnail of Growth of catalase A and catalase T deficient mutant strains of Saccharomyces cerevisiae on ethanol and oleic acid. Growth profiles and catalase activities in relation to microbody proliferation

The parental strain (A+T +) of Saccharomyces cerevisiae and mutants, deficient in catalase T (A+T... more The parental strain (A+T +) of Saccharomyces cerevisiae and mutants, deficient in catalase T (A+T-), catalase A (A-T +) or both catalases (A-T-), grew on ethanol and oleic acid with comparable doubling times. Specific activities of catalase were low in glucose-and ethanol-grown cells. In the two oleic acid-grown A +strains (A+T + and A+T -) high catalase activities were found; catalase activity invariably remained low in the A -T + strain and was never detected in the A -T -strain. The levels of t-oxidation enzymes in oleic acid-grown cells of the parental and all mutant strains were not significantly different. However, cytochrome C peroxidase activity had increased 8-fold in oleic acid grown Astrains (A-T + and A -T -) compared to parental strain cells. The degree of peroxisomal proliferation was comparable among the different strains. Catalase A was shown to be located in peroxisomes. Catalase T is most probably cytosolic in nature and/or present in the periplasmic space.

Research paper thumbnail of Protein complexes in the peroxisomal membrane visualized by blue native gel electrophoresis

Research paper thumbnail of Fluorescence analysis of the Hansenula polymorpha peroxisomal targeting signal-1 receptor, Pex5p

European Journal of Biochemistry, 2003

Correct sorting of newly synthesized peroxisomal matrix proteins is dependent on a peroxisomal ta... more Correct sorting of newly synthesized peroxisomal matrix proteins is dependent on a peroxisomal targeting signal (PTS). So far two PTSs are known. PTS1 consists of a tripeptide that is located at the extreme C terminus of matrix proteins and is specifically recognized by the PTS1-receptor Pex5p. We studied Hansenula polymorpha Pex5p (HpPex5p) using fluorescence spectroscopy. The intensity of Trp fluorescence

Research paper thumbnail of Hansenula polymorpha: A Versatile Model Organism in Peroxisome Research

Research paper thumbnail of Hansenula polymorpha Vps1p is required for fission of uninduced constitutive peroxisomes but not of substrate-induced organelles

Research paper thumbnail of A crucial role of the mitochondrial protein import receptor MOM19 for the biogenesis of mitochondria

The Journal of Cell Biology

The novel genetic method of "sheltered RIP" (repeat induced point mutation) was used to generate ... more The novel genetic method of "sheltered RIP" (repeat induced point mutation) was used to generate a Neurospora crassa mutant in which MOM19, a component of the protein import machinery of the mitochondrial outer membrane, can be depleted. Deficiency in MOM19 resulted in a severe growth defect, but the cells remained viable. The number of mitochondrial profiles was not grossly changed, but mutant mitochondria were highly deficient in cristae membranes, cytochromes, and protein synthesis activity. Protein import into isolated mutant mitochondria was decreased by factors of 6 to 30 for most proteins from all suborganellar compartments. Proteins like the ADP/ATP carrier, MOM19, and cytochrome c, whose import into wild-type mito-

Research paper thumbnail of A stretch of positively charged amino acids at the N terminus of Hansenula polymorpha Pex3p is involved in incorporation of the protein into the peroxisomal membrane

Journal of Biological Chemistry

Pex3p is a peroxisomal membrane protein that is essential for peroxisome biogenesis. Here, we sho... more Pex3p is a peroxisomal membrane protein that is essential for peroxisome biogenesis. Here, we show that a conserved stretch of positively charged amino acids (Arg 11 -X-Lys-Lys-Lys 15 ) in the N terminus of Hansenula polymorpha Pex3p is involved in incorporation of the protein into its target membrane. Despite the strong conservation, this sequence shows a high degree of redundancy. Substitution of either Arg 11 , Lys 13 , Lys 14 , or Lys 15 with uncharged or negatively charged amino acids did not interfere with Pex3p location and function. However, a mutant Pex3p, carrying negatively charged amino acids at position 13 and 15 (K13E/K15E), caused moderate but significant defects in peroxisome assembly and matrix protein import. Additional changes in the N terminus of Pex3p, e.g. replacing three or four of the positively charged amino acids with negatively charged ones, led to a typical pex3 phenotype, i.e. accumulation of peroxisomal matrix proteins in the cytosol and absence of peroxisomal remnants. Also, in these cases, the mutant Pex3p levels were reduced. Remarkably, mutant Pex3p proteins were mislocalized to mitochondria or the cytosol, depending on the nature of the mutation. Furthermore, in case of reduced amounts of Pex3p, the levels of other peroxisomal membrane proteins, e.g. Pex10p and Pex14p, were also diminished, suggesting that Pex3p maybe involved in the recruitment or stabilization of these proteins (in the membrane). The abbreviations used are: AMO, amine oxidase; AO, alcohol oxidase; GFP, green fluorescent protein; eGFP, enhanced GFP; kb, kilobase pair(s); mPTS, membrane peroxisomal targeting signal; PMP, peroxisomal membrane protein; P AMO , amine oxidase promoter; P AOX , alcohol oxidase promoter; WT, wild type; Pex3p N50 , N-terminal 50 amino acids of Pex3p; Pex3p N224 , N-terminal 224 amino acids of Pex3p.

Research paper thumbnail of IMPROVED PENICILLIN PRODUCTION

The invention relates to improved means and methods for producing a compound of interest in funga... more The invention relates to improved means and methods for producing a compound of interest in fungal host cells. Provided is a method for producing at least one secondary metabolite in a fungal host cell, comprising culturing a fungal host cell capable of producing said metabolite under conditions allowing for production of said metabolite, wherein the host cell has been modified to display elevated levels of glyoxalase I and glyoxalase II activity.

Research paper thumbnail of Cellular responses of

FEMS Yeast Research

Extremely low specific growth rates (below 0.01 h−1) represent a largely unexplored area of micro... more Extremely low specific growth rates (below 0.01 h−1) represent a largely unexplored area of microbial physiology. In this study, anaerobic, glucose‐limited retentostats were used to analyse physiological and genome‐wide transcriptional responses of Saccharomyces cerevisiae to cultivation at near‐zero specific growth rates. While quiescence is typically investigated as a result of carbon starvation, cells in retentostat are fed by small, but continuous carbon and energy supply. Yeast cells cultivated near‐zero specific growth rates, while metabolically active, exhibited characteristics previously associated with quiescence, including accumulation of storage polymers and an increased expression of genes involved in exit from the cell cycle into G0. Unexpectedly, analysis of transcriptome data from retentostat and chemostat cultures showed, as specific growth rate was decreased, that quiescence‐related transcriptional responses were already set in at specific growth rates above 0.025 h...

Research paper thumbnail of Central Role of Mic10 in the Mitochondrial Contact Site and Cristae Organizing System

Cell Metabolism, 2015

Graphical Abstract Highlights d MICOS core subunit Mic10 is targeted into mitochondria by a charg... more Graphical Abstract Highlights d MICOS core subunit Mic10 is targeted into mitochondria by a charged internal loop d Two conserved glycine motifs are required to form large Mic10 oligomers d Overexpression of Mic10 strongly alters inner membrane extent and architecture d Mic10 oligomers play a central role in the formation of crista junctions

Research paper thumbnail of Structure of Pex5p and Pex5-20 complexes in the yeast Hansenula polymorpha. Pex20p causes a conformational change upon binding to Pex5p tetramers involved in peroxisomal protein transport

EMC 2008 14th European Microscopy Congress 1–5 September 2008, Aachen, Germany, 2008

ABSTRACT

Research paper thumbnail of Hansenula polymorpha pex11 cells are affected in peroxisome retention

The FEBS journal, 2009

We have cloned and characterized the Hansenula polymorpha PEX11 gene. Our morphological data are ... more We have cloned and characterized the Hansenula polymorpha PEX11 gene. Our morphological data are consistent with previous observations that peroxisome proliferation can be regulated by modulating Pex11p levels. Surprisingly, pex11 cells also showed a defect in peroxisome retention in mother cells during vegetative cell reproduction. Until now, Saccharomyces cerevisiae Inp1p has been the only peroxisomal protein that has been shown to play a role in the organelle retention process. H. polymorpha inp1 cells are also affected in peroxisome retention, like pex11 cells. We show by time-lapse imaging that Inp1-green fluorescent protein localization varies during the cell cycle and that the protein is normally recruited to peroxisomes in pex11 cells. Taken together, our data show that H. polymorpha Pex11p is not only important for peroxisome proliferation but is also required for proper peroxisome segregation during cell division.

Research paper thumbnail of How shall I eat thee?

Autophagy

autophagy, mitophagy, pexophagy, protein degradation, stress, xenophagy "All the cell's a stage, ... more autophagy, mitophagy, pexophagy, protein degradation, stress, xenophagy "All the cell's a stage, and all the cytoplasm in it merely a substrate for autophagy." From As You Eat It in Shakespeare for the Cell Biologist If you work in the field of autophagy we do not really need to tell you that this research area has grown tremendously. Along with that growth has developed a need for some unification of the nomenclature. In 2003, researchers working with the yeast model system proposed the use of the acronym ATG to denote AuTophaGy-related genes, 1 and this designation has also been adopted for most of the genes involved in autophagy in higher eukaryotes. Similarly, a common nomenclature for isoforms of lysosome associated protein type 2 (LAMP-2) was recently proposed, hopefully reducing some of the confusion resulting from the use of multiple names. 2 At this time we thought it worthwhile to consider the terms being used to describe different types of lysosomal or vacuolar degradative pathways. Many names are being introduced, and this is reasonable to the extent that these various processes have distinct features; each unique process needs a specific name to avoid confusion, and to eliminate the need for a lengthy description. It would be helpful, however, if the community agreed on their use. Finally, the addition or use of a name that implies a unique process must be backed up by data that justify the nomenclature. Thus, researchers should verify that a process is specific before using a name that implies specificity. For example, to demonstrate selectivity in organelle degradation it is incumbent upon the researcher to show that the organelle in question, and not other organelles, is sequestered and/or degraded with kinetics that distinguish it from a bulk, nonspecific process.

Research paper thumbnail of A peroxisomal lon protease and peroxisome degradation by autophagy play key roles in vitality of Hansenula polymorpha cells

Autophagy

In eukaryote cells various mechanisms exist that are responsible for the removal of non-functiona... more In eukaryote cells various mechanisms exist that are responsible for the removal of non-functional proteins. Here we show that in the yeast Hansenula polymorpha (H. polymorpha) a peroxisomal Lon protease, Pln, plays a role in degradation of unfolded and non-assembled peroxisomal matrix proteins. In addition, we demonstrate that whole peroxisomes are constitutively degraded by autophagy during normal vegetative growth of WT cells. Deletion of both H. polymorpha PLN and ATG1, required for autophagy, resulted in a significant increase in peroxisome numbers, paralleled by a decrease in cell viability relative to WT cells. Also, in these cells and in cells of PLN and ATG1 single deletion strains, the intracellular levels of reactive oxygen species had increased relative to WT controls. The enhanced generation of reactive oxygen species may be related to an uneven distribution of peroxisomal catalase activities in the mutant cells, as demonstrated by cytochemistry. We speculate that in th...

Research paper thumbnail of ATG Genes Involved in Non-Selective Autophagy are Conserved from Yeast to Man, but the Selective Cvt and Pexophagy Pathways also Require Organism-Specific Genes

Research paper thumbnail of How Shall I Eat Thee?

Autophagy, 2007

autophagy, mitophagy, pexophagy, protein degradation, stress, xenophagy "All the cell's a stage, ... more autophagy, mitophagy, pexophagy, protein degradation, stress, xenophagy "All the cell's a stage, and all the cytoplasm in it merely a substrate for autophagy." From As You Eat It in Shakespeare for the Cell Biologist If you work in the field of autophagy we do not really need to tell you that this research area has grown tremendously. Along with that growth has developed a need for some unification of the nomenclature. In 2003, researchers working with the yeast model system proposed the use of the acronym ATG to denote AuTophaGy-related genes, 1 and this designation has also been adopted for most of the genes involved in autophagy in higher eukaryotes. Similarly, a common nomenclature for isoforms of lysosome associated protein type 2 (LAMP-2) was recently proposed, hopefully reducing some of the confusion resulting from the use of multiple names. 2 At this time we thought it worthwhile to consider the terms being used to describe different types of lysosomal or vacuolar degradative pathways. Many names are being introduced, and this is reasonable to the extent that these various processes have distinct features; each unique process needs a specific name to avoid confusion, and to eliminate the need for a lengthy description. It would be helpful, however, if the community agreed on their use. Finally, the addition or use of a name that implies a unique process must be backed up by data that justify the nomenclature. Thus, researchers should verify that a process is specific before using a name that implies specificity. For example, to demonstrate selectivity in organelle degradation it is incumbent upon the researcher to show that the organelle in question, and not other organelles, is sequestered and/or degraded with kinetics that distinguish it from a bulk, nonspecific process.

Research paper thumbnail of Pex14 is the sole component of the peroxisomal translocon that is required for pexophagy

Autophagy, 2008

Pex14 was initially identified as a peroxisomal membrane protein that is involved in docking of t... more Pex14 was initially identified as a peroxisomal membrane protein that is involved in docking of the soluble receptor proteins Pex5 and Pex7, which are required for import of PTS1-or PTS2-containing peroxisomal matrix proteins. However, Hansenula polymorpha Pex14 is also required for selective degradation of peroxisomes (pexophagy). Previously we showed that Pex1, Pex4, Pex6 and Pex8 are not required for this process. Here we show that also in the absence of various other peroxins, namely Pex2, Pex10, Pex12, Pex13 and Pex17, pexophagy can normally occur. These peroxins are, like Pex14, components of the peroxisomal translocon. Our data confirm that Pex14 is the sole peroxin that has a unique dual function in two apparent opposite processes, namely peroxisome formation and selective degradation.

Research paper thumbnail of The requirement of matrix ATP for the import of precursor proteins into the mitochondrial matrix and intermembrane space

European Journal of Biochemistry, 1994

The role of ATP in the matrix for the import of precursor proteins into the various mitochondrial... more The role of ATP in the matrix for the import of precursor proteins into the various mitochondrial subcompartments was investigated by studying protein translocation at experimentally defined ATP levels. Proteins targeted to the matrix were neither imported or processed when matrix ATP was depleted. Import and processing of precytochrome b2 (pb2), a precursor carrying a bipartite presequence, into the intermembrane

Research paper thumbnail of Pex14p is Not Required for Microautophagy and in Catalytic Amounts for Macropexophagy in Hansenula polymorpha

Research paper thumbnail of Cytoplasmic recycling of 60S preribosomal factors depends on the AAA protein Drg1

Allelic forms of DRG1/AFG2 confer resistance to the drug diazaborine, an inhibitor of ribosome bi... more Allelic forms of DRG1/AFG2 confer resistance to the drug diazaborine, an inhibitor of ribosome biogenesis in Saccharomyces cerevisiae. Our results show that the AAA-ATPase Drg1 is essential for 60S maturation and associates with 60S precursor particles in the cytoplasm. Functional inactivation of Drg1 leads to an increased cytoplasmic localization of shuttling pre-60S maturation factors like Rlp24, Arx1, and Tif6. Surprisingly, Nog1, a nuclear pre-60S factor, was also relocalized to the cytoplasm under these conditions, suggesting that it is a previously unsuspected shuttling preribosomal factor that is exported with the precursor particles and very rapidly reimported. Proteins that became cytoplasmic under drg1 mutant conditions were blocked on pre-60S particles at a step that precedes the association of Rei1, a later-acting preribosomal factor. A similar cytoplasmic accumulation of Nog1 and Rlp24 in pre-60S-bound form could be seen after overexpression of a dominant-negative Drg1 variant mutated in the D2 ATPase domain. We conclude that the ATPase activity of Drg1 is required for the release of shuttling proteins from the pre-60S particles shortly after their nuclear export. This early cytoplasmic release reaction defines a novel step in eukaryotic ribosome maturation.

Research paper thumbnail of Growth of catalase A and catalase T deficient mutant strains of Saccharomyces cerevisiae on ethanol and oleic acid. Growth profiles and catalase activities in relation to microbody proliferation

The parental strain (A+T +) of Saccharomyces cerevisiae and mutants, deficient in catalase T (A+T... more The parental strain (A+T +) of Saccharomyces cerevisiae and mutants, deficient in catalase T (A+T-), catalase A (A-T +) or both catalases (A-T-), grew on ethanol and oleic acid with comparable doubling times. Specific activities of catalase were low in glucose-and ethanol-grown cells. In the two oleic acid-grown A +strains (A+T + and A+T -) high catalase activities were found; catalase activity invariably remained low in the A -T + strain and was never detected in the A -T -strain. The levels of t-oxidation enzymes in oleic acid-grown cells of the parental and all mutant strains were not significantly different. However, cytochrome C peroxidase activity had increased 8-fold in oleic acid grown Astrains (A-T + and A -T -) compared to parental strain cells. The degree of peroxisomal proliferation was comparable among the different strains. Catalase A was shown to be located in peroxisomes. Catalase T is most probably cytosolic in nature and/or present in the periplasmic space.

Research paper thumbnail of Protein complexes in the peroxisomal membrane visualized by blue native gel electrophoresis

Research paper thumbnail of Fluorescence analysis of the Hansenula polymorpha peroxisomal targeting signal-1 receptor, Pex5p

European Journal of Biochemistry, 2003

Correct sorting of newly synthesized peroxisomal matrix proteins is dependent on a peroxisomal ta... more Correct sorting of newly synthesized peroxisomal matrix proteins is dependent on a peroxisomal targeting signal (PTS). So far two PTSs are known. PTS1 consists of a tripeptide that is located at the extreme C terminus of matrix proteins and is specifically recognized by the PTS1-receptor Pex5p. We studied Hansenula polymorpha Pex5p (HpPex5p) using fluorescence spectroscopy. The intensity of Trp fluorescence

Research paper thumbnail of Hansenula polymorpha: A Versatile Model Organism in Peroxisome Research

Research paper thumbnail of Hansenula polymorpha Vps1p is required for fission of uninduced constitutive peroxisomes but not of substrate-induced organelles

Research paper thumbnail of A crucial role of the mitochondrial protein import receptor MOM19 for the biogenesis of mitochondria

The Journal of Cell Biology

The novel genetic method of "sheltered RIP" (repeat induced point mutation) was used to generate ... more The novel genetic method of "sheltered RIP" (repeat induced point mutation) was used to generate a Neurospora crassa mutant in which MOM19, a component of the protein import machinery of the mitochondrial outer membrane, can be depleted. Deficiency in MOM19 resulted in a severe growth defect, but the cells remained viable. The number of mitochondrial profiles was not grossly changed, but mutant mitochondria were highly deficient in cristae membranes, cytochromes, and protein synthesis activity. Protein import into isolated mutant mitochondria was decreased by factors of 6 to 30 for most proteins from all suborganellar compartments. Proteins like the ADP/ATP carrier, MOM19, and cytochrome c, whose import into wild-type mito-

Research paper thumbnail of A stretch of positively charged amino acids at the N terminus of Hansenula polymorpha Pex3p is involved in incorporation of the protein into the peroxisomal membrane

Journal of Biological Chemistry

Pex3p is a peroxisomal membrane protein that is essential for peroxisome biogenesis. Here, we sho... more Pex3p is a peroxisomal membrane protein that is essential for peroxisome biogenesis. Here, we show that a conserved stretch of positively charged amino acids (Arg 11 -X-Lys-Lys-Lys 15 ) in the N terminus of Hansenula polymorpha Pex3p is involved in incorporation of the protein into its target membrane. Despite the strong conservation, this sequence shows a high degree of redundancy. Substitution of either Arg 11 , Lys 13 , Lys 14 , or Lys 15 with uncharged or negatively charged amino acids did not interfere with Pex3p location and function. However, a mutant Pex3p, carrying negatively charged amino acids at position 13 and 15 (K13E/K15E), caused moderate but significant defects in peroxisome assembly and matrix protein import. Additional changes in the N terminus of Pex3p, e.g. replacing three or four of the positively charged amino acids with negatively charged ones, led to a typical pex3 phenotype, i.e. accumulation of peroxisomal matrix proteins in the cytosol and absence of peroxisomal remnants. Also, in these cases, the mutant Pex3p levels were reduced. Remarkably, mutant Pex3p proteins were mislocalized to mitochondria or the cytosol, depending on the nature of the mutation. Furthermore, in case of reduced amounts of Pex3p, the levels of other peroxisomal membrane proteins, e.g. Pex10p and Pex14p, were also diminished, suggesting that Pex3p maybe involved in the recruitment or stabilization of these proteins (in the membrane). The abbreviations used are: AMO, amine oxidase; AO, alcohol oxidase; GFP, green fluorescent protein; eGFP, enhanced GFP; kb, kilobase pair(s); mPTS, membrane peroxisomal targeting signal; PMP, peroxisomal membrane protein; P AMO , amine oxidase promoter; P AOX , alcohol oxidase promoter; WT, wild type; Pex3p N50 , N-terminal 50 amino acids of Pex3p; Pex3p N224 , N-terminal 224 amino acids of Pex3p.

Research paper thumbnail of IMPROVED PENICILLIN PRODUCTION

The invention relates to improved means and methods for producing a compound of interest in funga... more The invention relates to improved means and methods for producing a compound of interest in fungal host cells. Provided is a method for producing at least one secondary metabolite in a fungal host cell, comprising culturing a fungal host cell capable of producing said metabolite under conditions allowing for production of said metabolite, wherein the host cell has been modified to display elevated levels of glyoxalase I and glyoxalase II activity.

Research paper thumbnail of Cellular responses of

FEMS Yeast Research

Extremely low specific growth rates (below 0.01 h−1) represent a largely unexplored area of micro... more Extremely low specific growth rates (below 0.01 h−1) represent a largely unexplored area of microbial physiology. In this study, anaerobic, glucose‐limited retentostats were used to analyse physiological and genome‐wide transcriptional responses of Saccharomyces cerevisiae to cultivation at near‐zero specific growth rates. While quiescence is typically investigated as a result of carbon starvation, cells in retentostat are fed by small, but continuous carbon and energy supply. Yeast cells cultivated near‐zero specific growth rates, while metabolically active, exhibited characteristics previously associated with quiescence, including accumulation of storage polymers and an increased expression of genes involved in exit from the cell cycle into G0. Unexpectedly, analysis of transcriptome data from retentostat and chemostat cultures showed, as specific growth rate was decreased, that quiescence‐related transcriptional responses were already set in at specific growth rates above 0.025 h...

Research paper thumbnail of Central Role of Mic10 in the Mitochondrial Contact Site and Cristae Organizing System

Cell Metabolism, 2015

Graphical Abstract Highlights d MICOS core subunit Mic10 is targeted into mitochondria by a charg... more Graphical Abstract Highlights d MICOS core subunit Mic10 is targeted into mitochondria by a charged internal loop d Two conserved glycine motifs are required to form large Mic10 oligomers d Overexpression of Mic10 strongly alters inner membrane extent and architecture d Mic10 oligomers play a central role in the formation of crista junctions

Research paper thumbnail of Structure of Pex5p and Pex5-20 complexes in the yeast Hansenula polymorpha. Pex20p causes a conformational change upon binding to Pex5p tetramers involved in peroxisomal protein transport

EMC 2008 14th European Microscopy Congress 1–5 September 2008, Aachen, Germany, 2008

ABSTRACT

Research paper thumbnail of Hansenula polymorpha pex11 cells are affected in peroxisome retention

The FEBS journal, 2009

We have cloned and characterized the Hansenula polymorpha PEX11 gene. Our morphological data are ... more We have cloned and characterized the Hansenula polymorpha PEX11 gene. Our morphological data are consistent with previous observations that peroxisome proliferation can be regulated by modulating Pex11p levels. Surprisingly, pex11 cells also showed a defect in peroxisome retention in mother cells during vegetative cell reproduction. Until now, Saccharomyces cerevisiae Inp1p has been the only peroxisomal protein that has been shown to play a role in the organelle retention process. H. polymorpha inp1 cells are also affected in peroxisome retention, like pex11 cells. We show by time-lapse imaging that Inp1-green fluorescent protein localization varies during the cell cycle and that the protein is normally recruited to peroxisomes in pex11 cells. Taken together, our data show that H. polymorpha Pex11p is not only important for peroxisome proliferation but is also required for proper peroxisome segregation during cell division.

Research paper thumbnail of How shall I eat thee?

Autophagy

autophagy, mitophagy, pexophagy, protein degradation, stress, xenophagy "All the cell's a stage, ... more autophagy, mitophagy, pexophagy, protein degradation, stress, xenophagy "All the cell's a stage, and all the cytoplasm in it merely a substrate for autophagy." From As You Eat It in Shakespeare for the Cell Biologist If you work in the field of autophagy we do not really need to tell you that this research area has grown tremendously. Along with that growth has developed a need for some unification of the nomenclature. In 2003, researchers working with the yeast model system proposed the use of the acronym ATG to denote AuTophaGy-related genes, 1 and this designation has also been adopted for most of the genes involved in autophagy in higher eukaryotes. Similarly, a common nomenclature for isoforms of lysosome associated protein type 2 (LAMP-2) was recently proposed, hopefully reducing some of the confusion resulting from the use of multiple names. 2 At this time we thought it worthwhile to consider the terms being used to describe different types of lysosomal or vacuolar degradative pathways. Many names are being introduced, and this is reasonable to the extent that these various processes have distinct features; each unique process needs a specific name to avoid confusion, and to eliminate the need for a lengthy description. It would be helpful, however, if the community agreed on their use. Finally, the addition or use of a name that implies a unique process must be backed up by data that justify the nomenclature. Thus, researchers should verify that a process is specific before using a name that implies specificity. For example, to demonstrate selectivity in organelle degradation it is incumbent upon the researcher to show that the organelle in question, and not other organelles, is sequestered and/or degraded with kinetics that distinguish it from a bulk, nonspecific process.

Research paper thumbnail of A peroxisomal lon protease and peroxisome degradation by autophagy play key roles in vitality of Hansenula polymorpha cells

Autophagy

In eukaryote cells various mechanisms exist that are responsible for the removal of non-functiona... more In eukaryote cells various mechanisms exist that are responsible for the removal of non-functional proteins. Here we show that in the yeast Hansenula polymorpha (H. polymorpha) a peroxisomal Lon protease, Pln, plays a role in degradation of unfolded and non-assembled peroxisomal matrix proteins. In addition, we demonstrate that whole peroxisomes are constitutively degraded by autophagy during normal vegetative growth of WT cells. Deletion of both H. polymorpha PLN and ATG1, required for autophagy, resulted in a significant increase in peroxisome numbers, paralleled by a decrease in cell viability relative to WT cells. Also, in these cells and in cells of PLN and ATG1 single deletion strains, the intracellular levels of reactive oxygen species had increased relative to WT controls. The enhanced generation of reactive oxygen species may be related to an uneven distribution of peroxisomal catalase activities in the mutant cells, as demonstrated by cytochemistry. We speculate that in th...

Research paper thumbnail of ATG Genes Involved in Non-Selective Autophagy are Conserved from Yeast to Man, but the Selective Cvt and Pexophagy Pathways also Require Organism-Specific Genes

Research paper thumbnail of How Shall I Eat Thee?

Autophagy, 2007

autophagy, mitophagy, pexophagy, protein degradation, stress, xenophagy "All the cell's a stage, ... more autophagy, mitophagy, pexophagy, protein degradation, stress, xenophagy "All the cell's a stage, and all the cytoplasm in it merely a substrate for autophagy." From As You Eat It in Shakespeare for the Cell Biologist If you work in the field of autophagy we do not really need to tell you that this research area has grown tremendously. Along with that growth has developed a need for some unification of the nomenclature. In 2003, researchers working with the yeast model system proposed the use of the acronym ATG to denote AuTophaGy-related genes, 1 and this designation has also been adopted for most of the genes involved in autophagy in higher eukaryotes. Similarly, a common nomenclature for isoforms of lysosome associated protein type 2 (LAMP-2) was recently proposed, hopefully reducing some of the confusion resulting from the use of multiple names. 2 At this time we thought it worthwhile to consider the terms being used to describe different types of lysosomal or vacuolar degradative pathways. Many names are being introduced, and this is reasonable to the extent that these various processes have distinct features; each unique process needs a specific name to avoid confusion, and to eliminate the need for a lengthy description. It would be helpful, however, if the community agreed on their use. Finally, the addition or use of a name that implies a unique process must be backed up by data that justify the nomenclature. Thus, researchers should verify that a process is specific before using a name that implies specificity. For example, to demonstrate selectivity in organelle degradation it is incumbent upon the researcher to show that the organelle in question, and not other organelles, is sequestered and/or degraded with kinetics that distinguish it from a bulk, nonspecific process.

Research paper thumbnail of Pex14 is the sole component of the peroxisomal translocon that is required for pexophagy

Autophagy, 2008

Pex14 was initially identified as a peroxisomal membrane protein that is involved in docking of t... more Pex14 was initially identified as a peroxisomal membrane protein that is involved in docking of the soluble receptor proteins Pex5 and Pex7, which are required for import of PTS1-or PTS2-containing peroxisomal matrix proteins. However, Hansenula polymorpha Pex14 is also required for selective degradation of peroxisomes (pexophagy). Previously we showed that Pex1, Pex4, Pex6 and Pex8 are not required for this process. Here we show that also in the absence of various other peroxins, namely Pex2, Pex10, Pex12, Pex13 and Pex17, pexophagy can normally occur. These peroxins are, like Pex14, components of the peroxisomal translocon. Our data confirm that Pex14 is the sole peroxin that has a unique dual function in two apparent opposite processes, namely peroxisome formation and selective degradation.

Research paper thumbnail of The requirement of matrix ATP for the import of precursor proteins into the mitochondrial matrix and intermembrane space

European Journal of Biochemistry, 1994

The role of ATP in the matrix for the import of precursor proteins into the various mitochondrial... more The role of ATP in the matrix for the import of precursor proteins into the various mitochondrial subcompartments was investigated by studying protein translocation at experimentally defined ATP levels. Proteins targeted to the matrix were neither imported or processed when matrix ATP was depleted. Import and processing of precytochrome b2 (pb2), a precursor carrying a bipartite presequence, into the intermembrane

Research paper thumbnail of Pex14p is Not Required for Microautophagy and in Catalytic Amounts for Macropexophagy in Hansenula polymorpha

Research paper thumbnail of Cytoplasmic recycling of 60S preribosomal factors depends on the AAA protein Drg1

Allelic forms of DRG1/AFG2 confer resistance to the drug diazaborine, an inhibitor of ribosome bi... more Allelic forms of DRG1/AFG2 confer resistance to the drug diazaborine, an inhibitor of ribosome biogenesis in Saccharomyces cerevisiae. Our results show that the AAA-ATPase Drg1 is essential for 60S maturation and associates with 60S precursor particles in the cytoplasm. Functional inactivation of Drg1 leads to an increased cytoplasmic localization of shuttling pre-60S maturation factors like Rlp24, Arx1, and Tif6. Surprisingly, Nog1, a nuclear pre-60S factor, was also relocalized to the cytoplasm under these conditions, suggesting that it is a previously unsuspected shuttling preribosomal factor that is exported with the precursor particles and very rapidly reimported. Proteins that became cytoplasmic under drg1 mutant conditions were blocked on pre-60S particles at a step that precedes the association of Rei1, a later-acting preribosomal factor. A similar cytoplasmic accumulation of Nog1 and Rlp24 in pre-60S-bound form could be seen after overexpression of a dominant-negative Drg1 variant mutated in the D2 ATPase domain. We conclude that the ATPase activity of Drg1 is required for the release of shuttling proteins from the pre-60S particles shortly after their nuclear export. This early cytoplasmic release reaction defines a novel step in eukaryotic ribosome maturation.