Joris Winderickx | KU Leuven (original) (raw)
Papers by Joris Winderickx
Biochemistry, 2005
Hyperphosphorylation and aggregation of protein tau are typical for neurodegenerative tauopathies... more Hyperphosphorylation and aggregation of protein tau are typical for neurodegenerative tauopathies, including Alzheimer's disease (AD). We demonstrate here that human tau expressed in yeast acquired pathological phosphoepitopes, assumed a pathological conformation, and formed aggregates. These processes were modulated by yeast kinases Mds1 and Pho85, orthologues of GSK-3 and cdk5, respectively. Surprisingly, inactivation of Pho85 increased phosphorylation of tau-4R, concomitant with increased conformational change defined by antibody MC1 and a 40-fold increase in aggregation. Soluble protein tau, purified from yeast lacking PHO85, spontaneously and rapidly formed tau filaments in vitro. Further fractionation of tau by anion-exchange chromatography yielded a hyperphosphorylated monomeric subfraction, termed hP-tau/MC1, with slow electrophoretic mobility and enriched with all major epitopes, including MC1. Isolated hP-tau/MC1 vastly accelerated in vitro aggregation of wild-type tau-4R, demonstrating its functional capacity to initiate aggregation, as well as its structural stability. Combined, this novel yeast model recapitulates hyperphosphorylation, conformation, and aggregation of protein tau, provides insight in molecular changes crucial in tauopathies, offers a source for isolation of modified protein tau, and has potential for identification of modulating compounds and genes. † This investigation was supported by the Fonds voor Wetenschappelijk Onderzoek-Vlaanderen (FWO-Vlaanderen), KULeuven Special Research Fund (KULeuven-BOF), Instituut voor Wetenschappelijk en Technisch Onderzoek (IWT), KULeuven R&D, and the Roomsfund.
Molecular Microbiology, 1999
FEMS Yeast Research, 2010
Unraveling the biochemical and genetic alterations that control the aggregation of protein tau is... more Unraveling the biochemical and genetic alterations that control the aggregation of protein tau is crucial to understand the etiology of tau-related neurodegenerative disorders. We expressed wild type and six clinical frontotemporal dementia with parkinsonism (FTDP) mutants of human protein tau in wild-type yeast cells and cells lacking Mds1 or Pho85, the respective orthologues of the tau kinases GSK3β and cdk5. We compared tau phosphorylation with the levels of sarkosyl-insoluble tau (SinT), as a measure for tau aggregation. The deficiency of Pho85 enhanced significantly the phosphorylation of serine-409 (S409) in all tau mutants, which coincided with marked increases in SinT levels. FTDP mutants tau-P301L and tau-R406W were least phosphorylated at S409 and produced the lowest levels of SinT, indicating that S409 phosphorylation is a direct determinant for tau aggregation. This finding was substantiated by the synthetic tau-S409A mutant that failed to produce significant amounts of SinT, while its pseudophosphorylated counterpart tau-S409E yielded SinT levels higher than or comparable to wild-type tau. Furthermore, S409 phosphorylation reduced the binding of protein tau to preformed microtubules. The highest SinT levels were found in yeast cells subjected to oxidative stress and with mitochondrial dysfunction. Under these conditions, the aggregation of tau was enhanced although the protein is less phosphorylated, suggesting that additional mechanisms are involved. Our results validate yeast as a prime model to identify the genetic and biochemical factors that contribute to the pathophysiology of human tau.
Molecular Microbiology, 2000
Yeast, 1999
When glucose is added to Saccharomyces cerevisiae cells grown into stationary phase or on non-fer... more When glucose is added to Saccharomyces cerevisiae cells grown into stationary phase or on non-fermentable carbon sources a rapid loss of heat stress resistance occurs. Mutants that retain high stress resistance after addition of glucose are called 'fil', for deficient in fermentation induced loss of stress resistance. Transformation of the fil1 mutant, which harbours a point mutation in adenylate cyclase, with a yeast gene library on a single copy plasmid resulted in transformants that were again stress-sensitive. One of the genes isolated in this way was a gene of previously unknown function. We have called it SFI1, for suppressor of fil1. SFI1 is an essential gene. Combination of Sfi1 and cAMP pathway mutations indicates that Sfi1 itself is not involved in the cAMP pathway. Conditional sfi1 mutants did not show enhanced heat resistance under the restrictive condition, whereas overexpression of SFI1 rendered cells heat-sensitive. Sfi1 may be a downstream target of the protein kinase A pathway, but its precise relationship with heat resistance remains unclear. Further analysis showed that Sfi1 is required for cell cycle progression, more specifically for progression through G 2 -M transition. Cells expressing SFI1 under the control of a galactose-inducible promoter arrest after addition of glucose as doublets of undivided mother and daughter cells. These doublets contain a single nucleus and lack mitotic spindles. Sfi1 shares homology with Xenopus laevis XCAP-C, a protein required for chromosome assembly. The conserved residues between these two proteins show a strong bias for charged amino acids. Hence, Sfi1 might be required for correct mitotic spindle assembly and its precise role might be in chromosome condensation. In conclusion, we have identified an essential function in the G 2 -M transition of the cell cycle for a yeast gene of previously unknown function. The EMBL Accession No. of the SFI1 nucleotide sequence is X95569.
Journal of the American Society of Brewing Chemists, 2001
ABSTRACT A strategy to alter the flocculation properties of nonflocculent yeast in such a way tha... more ABSTRACT A strategy to alter the flocculation properties of nonflocculent yeast in such a way that flocculation occurred toward the end of fermentation was developed. In a double cross-over event, the wild-type FLO1 promoter of the haploid, nonflocculent S. cerevisiae FY23 strain was replaced by a construct consisting of the SMRI-410 marker gene and the HSP30 promoter. In this way, the genomic copy of the wild-type FLO1 open reading frame was brought under transcriptional control of the HSP30 promoter. The transformants showed strong flocculation toward the end of fermentation, resulting in a distinctly clearer beer than the beer obtained with wild-type cells. The other properties of the wild-type strain were conserved. Moreover, it was shown that the transformants were extremely stable and that flocculation could be induced earlier during fermentation by a heat-shock treatment or the addition of ethanol to the medium. These results suggest that the flocculation properties of weakly flocculent brewer's yeast strains can be improved using controlled expression of the FLO1 gene.
Technical Quarterly Master Brewers Association of the Americas, 2003
The study of signal transduction in microorganisms has become a major research topic in molecular... more The study of signal transduction in microorganisms has become a major research topic in molecular and cellular biology. In this era, thorough knowledge of microbial physiology is no longer the sole and exclusive interest of academic research. It is now being acknowledged as a major importance for food, feed, and nutritional R&D. Detailed investigation of the mechanisms by which cells
CMU controls growth, however, is still an issue of conjecture, and further elucidation of this pr... more CMU controls growth, however, is still an issue of conjecture, and further elucidation of this process will certainly de-University of Geneva 1211 Geneva 4 pend on the identification of downstream effectors of PKA. One such effector, the protein kinase Rim15, is Switzerland 2 Functional Biology required for proper establishment of the G 0 program and is inhibited by PKA-mediated phosphorylation under Katholieke Universiteit Leuven Leuven-Heverlee conditions of nutrient abundance (Reinders et al., 1998). Interestingly, downregulation of PKA following nutrient B-3001 Flanders Belgium limitation liberates Rim15 from PKA-inhibition and results in activation of G 0 -related changes, which are strikingly similar to the changes observed following rapamycin treatment. These findings suggest a model in Summary which the TOR pathway may converge on a component and/or target of the PKA pathway to control entry into G 0 .
Documenta Ophthalmologica Proceedings Series, 1995
Topics in Current Genetics, 2002
Page 1. 7 From feast to famine; adaptation to nutrient availability in yeast Joris Winderickx1, I... more Page 1. 7 From feast to famine; adaptation to nutrient availability in yeast Joris Winderickx1, Inge Holsbeeks1, Ole Lagatie1, Frank Giots1, Johan Thevelein1 and Han de Winde2,3 1Laboratorium voor Moleculaire Celbiologie ...
Nature, Jan 2, 1992
Genetic variation of human senses within the normal range probably exists but usually cannot be i... more Genetic variation of human senses within the normal range probably exists but usually cannot be investigated in detail for lack of appropriate methods. The study of subtle perceptual differences in red-green colour vision is feasible since both photopigment genotypes and psychophysical phenotypes can be assessed by sophisticated techniques. Red-green colour vision in humans is mediated by two different visual pigments: red (long-wavelength sensitive) and green (middle-wavelength sensitive). The apoproteins of these highly homologous photopigments are encoded by genes on the X chromosome. Colour matches of males with normal colour vision fall into two main groups that appear to be transmitted by X-linked inheritance. This difference in colour matching is likely to reflect small variations in the absorption maxima of visual pigments, suggesting the presence of two common variants of the red and/or green visual pigments that differ in spectral positioning. We report that a common singl...
BMC Meeting Abstracts, 2001
Oxidative Medicine and Cellular Longevity, 2013
Over the past decade, the baker's yeast Saccharomyces cerevisiae has proven to be a useful model ... more Over the past decade, the baker's yeast Saccharomyces cerevisiae has proven to be a useful model system to investigate fundamental questions concerning the pathogenic role of human proteins in neurodegenerative diseases such as Parkinson's disease (PD). These so-called humanized yeast models for PD initially focused on -synuclein, which plays a key role in the etiology of PD. Upon expression of this human protein in the baker's yeast Saccharomyces cerevisiae, the events leading to aggregation and the molecular mechanisms that result in cellular toxicity are faithfully reproduced. More recently, a similar model to study the presumed pathobiology of the -synuclein interaction partner synphilin-1 has been established. In this review we will discuss recent advances using these humanized yeast models, pointing to new roles for cell wall integrity signaling, Ca 2+ homeostasis, mitophagy, and the cytoskeleton.
Documenta Ophthalmologica Proceedings Series, 1993
Yeast, 1999
RNA was extracted from yeast cells as described previously . For quantitation of transcript level... more RNA was extracted from yeast cells as described previously . For quantitation of transcript levels, 10 g of each RNA preparation were electrophoresed under 330 . . .
Biochemistry, 2005
Hyperphosphorylation and aggregation of protein tau are typical for neurodegenerative tauopathies... more Hyperphosphorylation and aggregation of protein tau are typical for neurodegenerative tauopathies, including Alzheimer's disease (AD). We demonstrate here that human tau expressed in yeast acquired pathological phosphoepitopes, assumed a pathological conformation, and formed aggregates. These processes were modulated by yeast kinases Mds1 and Pho85, orthologues of GSK-3 and cdk5, respectively. Surprisingly, inactivation of Pho85 increased phosphorylation of tau-4R, concomitant with increased conformational change defined by antibody MC1 and a 40-fold increase in aggregation. Soluble protein tau, purified from yeast lacking PHO85, spontaneously and rapidly formed tau filaments in vitro. Further fractionation of tau by anion-exchange chromatography yielded a hyperphosphorylated monomeric subfraction, termed hP-tau/MC1, with slow electrophoretic mobility and enriched with all major epitopes, including MC1. Isolated hP-tau/MC1 vastly accelerated in vitro aggregation of wild-type tau-4R, demonstrating its functional capacity to initiate aggregation, as well as its structural stability. Combined, this novel yeast model recapitulates hyperphosphorylation, conformation, and aggregation of protein tau, provides insight in molecular changes crucial in tauopathies, offers a source for isolation of modified protein tau, and has potential for identification of modulating compounds and genes. † This investigation was supported by the Fonds voor Wetenschappelijk Onderzoek-Vlaanderen (FWO-Vlaanderen), KULeuven Special Research Fund (KULeuven-BOF), Instituut voor Wetenschappelijk en Technisch Onderzoek (IWT), KULeuven R&D, and the Roomsfund.
Molecular Microbiology, 1999
FEMS Yeast Research, 2010
Unraveling the biochemical and genetic alterations that control the aggregation of protein tau is... more Unraveling the biochemical and genetic alterations that control the aggregation of protein tau is crucial to understand the etiology of tau-related neurodegenerative disorders. We expressed wild type and six clinical frontotemporal dementia with parkinsonism (FTDP) mutants of human protein tau in wild-type yeast cells and cells lacking Mds1 or Pho85, the respective orthologues of the tau kinases GSK3β and cdk5. We compared tau phosphorylation with the levels of sarkosyl-insoluble tau (SinT), as a measure for tau aggregation. The deficiency of Pho85 enhanced significantly the phosphorylation of serine-409 (S409) in all tau mutants, which coincided with marked increases in SinT levels. FTDP mutants tau-P301L and tau-R406W were least phosphorylated at S409 and produced the lowest levels of SinT, indicating that S409 phosphorylation is a direct determinant for tau aggregation. This finding was substantiated by the synthetic tau-S409A mutant that failed to produce significant amounts of SinT, while its pseudophosphorylated counterpart tau-S409E yielded SinT levels higher than or comparable to wild-type tau. Furthermore, S409 phosphorylation reduced the binding of protein tau to preformed microtubules. The highest SinT levels were found in yeast cells subjected to oxidative stress and with mitochondrial dysfunction. Under these conditions, the aggregation of tau was enhanced although the protein is less phosphorylated, suggesting that additional mechanisms are involved. Our results validate yeast as a prime model to identify the genetic and biochemical factors that contribute to the pathophysiology of human tau.
Molecular Microbiology, 2000
Yeast, 1999
When glucose is added to Saccharomyces cerevisiae cells grown into stationary phase or on non-fer... more When glucose is added to Saccharomyces cerevisiae cells grown into stationary phase or on non-fermentable carbon sources a rapid loss of heat stress resistance occurs. Mutants that retain high stress resistance after addition of glucose are called 'fil', for deficient in fermentation induced loss of stress resistance. Transformation of the fil1 mutant, which harbours a point mutation in adenylate cyclase, with a yeast gene library on a single copy plasmid resulted in transformants that were again stress-sensitive. One of the genes isolated in this way was a gene of previously unknown function. We have called it SFI1, for suppressor of fil1. SFI1 is an essential gene. Combination of Sfi1 and cAMP pathway mutations indicates that Sfi1 itself is not involved in the cAMP pathway. Conditional sfi1 mutants did not show enhanced heat resistance under the restrictive condition, whereas overexpression of SFI1 rendered cells heat-sensitive. Sfi1 may be a downstream target of the protein kinase A pathway, but its precise relationship with heat resistance remains unclear. Further analysis showed that Sfi1 is required for cell cycle progression, more specifically for progression through G 2 -M transition. Cells expressing SFI1 under the control of a galactose-inducible promoter arrest after addition of glucose as doublets of undivided mother and daughter cells. These doublets contain a single nucleus and lack mitotic spindles. Sfi1 shares homology with Xenopus laevis XCAP-C, a protein required for chromosome assembly. The conserved residues between these two proteins show a strong bias for charged amino acids. Hence, Sfi1 might be required for correct mitotic spindle assembly and its precise role might be in chromosome condensation. In conclusion, we have identified an essential function in the G 2 -M transition of the cell cycle for a yeast gene of previously unknown function. The EMBL Accession No. of the SFI1 nucleotide sequence is X95569.
Journal of the American Society of Brewing Chemists, 2001
ABSTRACT A strategy to alter the flocculation properties of nonflocculent yeast in such a way tha... more ABSTRACT A strategy to alter the flocculation properties of nonflocculent yeast in such a way that flocculation occurred toward the end of fermentation was developed. In a double cross-over event, the wild-type FLO1 promoter of the haploid, nonflocculent S. cerevisiae FY23 strain was replaced by a construct consisting of the SMRI-410 marker gene and the HSP30 promoter. In this way, the genomic copy of the wild-type FLO1 open reading frame was brought under transcriptional control of the HSP30 promoter. The transformants showed strong flocculation toward the end of fermentation, resulting in a distinctly clearer beer than the beer obtained with wild-type cells. The other properties of the wild-type strain were conserved. Moreover, it was shown that the transformants were extremely stable and that flocculation could be induced earlier during fermentation by a heat-shock treatment or the addition of ethanol to the medium. These results suggest that the flocculation properties of weakly flocculent brewer's yeast strains can be improved using controlled expression of the FLO1 gene.
Technical Quarterly Master Brewers Association of the Americas, 2003
The study of signal transduction in microorganisms has become a major research topic in molecular... more The study of signal transduction in microorganisms has become a major research topic in molecular and cellular biology. In this era, thorough knowledge of microbial physiology is no longer the sole and exclusive interest of academic research. It is now being acknowledged as a major importance for food, feed, and nutritional R&D. Detailed investigation of the mechanisms by which cells
CMU controls growth, however, is still an issue of conjecture, and further elucidation of this pr... more CMU controls growth, however, is still an issue of conjecture, and further elucidation of this process will certainly de-University of Geneva 1211 Geneva 4 pend on the identification of downstream effectors of PKA. One such effector, the protein kinase Rim15, is Switzerland 2 Functional Biology required for proper establishment of the G 0 program and is inhibited by PKA-mediated phosphorylation under Katholieke Universiteit Leuven Leuven-Heverlee conditions of nutrient abundance (Reinders et al., 1998). Interestingly, downregulation of PKA following nutrient B-3001 Flanders Belgium limitation liberates Rim15 from PKA-inhibition and results in activation of G 0 -related changes, which are strikingly similar to the changes observed following rapamycin treatment. These findings suggest a model in Summary which the TOR pathway may converge on a component and/or target of the PKA pathway to control entry into G 0 .
Documenta Ophthalmologica Proceedings Series, 1995
Topics in Current Genetics, 2002
Page 1. 7 From feast to famine; adaptation to nutrient availability in yeast Joris Winderickx1, I... more Page 1. 7 From feast to famine; adaptation to nutrient availability in yeast Joris Winderickx1, Inge Holsbeeks1, Ole Lagatie1, Frank Giots1, Johan Thevelein1 and Han de Winde2,3 1Laboratorium voor Moleculaire Celbiologie ...
Nature, Jan 2, 1992
Genetic variation of human senses within the normal range probably exists but usually cannot be i... more Genetic variation of human senses within the normal range probably exists but usually cannot be investigated in detail for lack of appropriate methods. The study of subtle perceptual differences in red-green colour vision is feasible since both photopigment genotypes and psychophysical phenotypes can be assessed by sophisticated techniques. Red-green colour vision in humans is mediated by two different visual pigments: red (long-wavelength sensitive) and green (middle-wavelength sensitive). The apoproteins of these highly homologous photopigments are encoded by genes on the X chromosome. Colour matches of males with normal colour vision fall into two main groups that appear to be transmitted by X-linked inheritance. This difference in colour matching is likely to reflect small variations in the absorption maxima of visual pigments, suggesting the presence of two common variants of the red and/or green visual pigments that differ in spectral positioning. We report that a common singl...
BMC Meeting Abstracts, 2001
Oxidative Medicine and Cellular Longevity, 2013
Over the past decade, the baker's yeast Saccharomyces cerevisiae has proven to be a useful model ... more Over the past decade, the baker's yeast Saccharomyces cerevisiae has proven to be a useful model system to investigate fundamental questions concerning the pathogenic role of human proteins in neurodegenerative diseases such as Parkinson's disease (PD). These so-called humanized yeast models for PD initially focused on -synuclein, which plays a key role in the etiology of PD. Upon expression of this human protein in the baker's yeast Saccharomyces cerevisiae, the events leading to aggregation and the molecular mechanisms that result in cellular toxicity are faithfully reproduced. More recently, a similar model to study the presumed pathobiology of the -synuclein interaction partner synphilin-1 has been established. In this review we will discuss recent advances using these humanized yeast models, pointing to new roles for cell wall integrity signaling, Ca 2+ homeostasis, mitophagy, and the cytoskeleton.
Documenta Ophthalmologica Proceedings Series, 1993
Yeast, 1999
RNA was extracted from yeast cells as described previously . For quantitation of transcript level... more RNA was extracted from yeast cells as described previously . For quantitation of transcript levels, 10 g of each RNA preparation were electrophoresed under 330 . . .