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Phosphorylation of Pex11p does not regulate peroxisomal fission in the yeast Hansenula polymorpha.

Thomas AS, Krikken AM, van der Klei IJ, Williams CP - Sci Rep (2015)

Bottom Line: Our data demonstrate that mutations to the phosphorylation site do not disturb the function of Pex11p in peroxisomal fission, nor do they alter the localization of Pex11p.Also, no effect on peroxisome inheritance was observed.Taken together, these data lead us to conclude that peroxisomal fission in H. polymorpha is not modulated by phosphorylation of Pex11p.

View Article: PubMed Central - PubMed

Affiliation: Molecular Cell Biology, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, the Netherlands.

ABSTRACT
Pex11p plays a crucial role in peroxisomal fission. Studies in Saccharomyces cerevisiae and Pichia pastoris indicated that Pex11p is activated by phosphorylation, which results in enhanced peroxisome proliferation. In S. cerevisiae but not in P. pastoris, Pex11p phosphorylation was shown to regulate the protein's trafficking to peroxisomes. However, phosphorylation of PpPex11p was proposed to influence its interaction with Fis1p, another component of the organellar fission machinery. Here, we have examined the role of Pex11p phosphorylation in the yeast Hansenula polymorpha. Employing mass spectrometry, we demonstrate that HpPex11p is also phosphorylated on a Serine residue present at a similar position to that of ScPex11p and PpPex11p. Furthermore, through the use of mutants designed to mimic both phosphorylated and unphosphorylated forms of HpPex11p, we have investigated the role of this post-translational modification. Our data demonstrate that mutations to the phosphorylation site do not disturb the function of Pex11p in peroxisomal fission, nor do they alter the localization of Pex11p. Also, no effect on peroxisome inheritance was observed. Taken together, these data lead us to conclude that peroxisomal fission in H. polymorpha is not modulated by phosphorylation of Pex11p.

No MeSH data available.


Phosphorylation of HpPex11p does not govern peroxisome inheritance.(A) Fluorescence microscopy images of glucose-grown pex11Δ cells using CLSM. Apart from the fluorescent peroxisomal matrix marker (GFP-SKL), cells produced WT or phosphorylation single mutant forms of Pex11p. All scale bars represent 1 μm. (B) Quantitative analysis of the images represented in (A) to show peroxisome distribution between mother and daughter cells. For all strains, at least 50 budding cells were counted. Error bars represent the standard deviation between two individual experiments.
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f5: Phosphorylation of HpPex11p does not govern peroxisome inheritance.(A) Fluorescence microscopy images of glucose-grown pex11Δ cells using CLSM. Apart from the fluorescent peroxisomal matrix marker (GFP-SKL), cells produced WT or phosphorylation single mutant forms of Pex11p. All scale bars represent 1 μm. (B) Quantitative analysis of the images represented in (A) to show peroxisome distribution between mother and daughter cells. For all strains, at least 50 budding cells were counted. Error bars represent the standard deviation between two individual experiments.

Mentions: It was shown previously that H. polymorpha pex11Δ cells display a peroxisome retention defect during growth on glucose13. The single peroxisome present in a cell migrates to the daughter cell upon budding, leaving the mother cell devoid of peroxisomes. To study if phosphorylation has a potential role in this process, budding cells of WT and Pex11p phosphorylation mutants producing GFP-SKL as peroxisomal marker were examined by fluorescence microscopy (Fig. 5A). We observed that peroxisome distribution over mother cells and buds is similar in WT and both phosphorylation mutants (Fig. 5B). This observation suggests that Pex11p phosphorylation does not affect peroxisome retention in H. polymorpha.


Phosphorylation of Pex11p does not regulate peroxisomal fission in the yeast Hansenula polymorpha.

Thomas AS, Krikken AM, van der Klei IJ, Williams CP - Sci Rep (2015)

Phosphorylation of HpPex11p does not govern peroxisome inheritance.(A) Fluorescence microscopy images of glucose-grown pex11Δ cells using CLSM. Apart from the fluorescent peroxisomal matrix marker (GFP-SKL), cells produced WT or phosphorylation single mutant forms of Pex11p. All scale bars represent 1 μm. (B) Quantitative analysis of the images represented in (A) to show peroxisome distribution between mother and daughter cells. For all strains, at least 50 budding cells were counted. Error bars represent the standard deviation between two individual experiments.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4477233&req=5

f5: Phosphorylation of HpPex11p does not govern peroxisome inheritance.(A) Fluorescence microscopy images of glucose-grown pex11Δ cells using CLSM. Apart from the fluorescent peroxisomal matrix marker (GFP-SKL), cells produced WT or phosphorylation single mutant forms of Pex11p. All scale bars represent 1 μm. (B) Quantitative analysis of the images represented in (A) to show peroxisome distribution between mother and daughter cells. For all strains, at least 50 budding cells were counted. Error bars represent the standard deviation between two individual experiments.
Mentions: It was shown previously that H. polymorpha pex11Δ cells display a peroxisome retention defect during growth on glucose13. The single peroxisome present in a cell migrates to the daughter cell upon budding, leaving the mother cell devoid of peroxisomes. To study if phosphorylation has a potential role in this process, budding cells of WT and Pex11p phosphorylation mutants producing GFP-SKL as peroxisomal marker were examined by fluorescence microscopy (Fig. 5A). We observed that peroxisome distribution over mother cells and buds is similar in WT and both phosphorylation mutants (Fig. 5B). This observation suggests that Pex11p phosphorylation does not affect peroxisome retention in H. polymorpha.

Bottom Line: Our data demonstrate that mutations to the phosphorylation site do not disturb the function of Pex11p in peroxisomal fission, nor do they alter the localization of Pex11p.Also, no effect on peroxisome inheritance was observed.Taken together, these data lead us to conclude that peroxisomal fission in H. polymorpha is not modulated by phosphorylation of Pex11p.

View Article: PubMed Central - PubMed

Affiliation: Molecular Cell Biology, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, the Netherlands.

ABSTRACT
Pex11p plays a crucial role in peroxisomal fission. Studies in Saccharomyces cerevisiae and Pichia pastoris indicated that Pex11p is activated by phosphorylation, which results in enhanced peroxisome proliferation. In S. cerevisiae but not in P. pastoris, Pex11p phosphorylation was shown to regulate the protein's trafficking to peroxisomes. However, phosphorylation of PpPex11p was proposed to influence its interaction with Fis1p, another component of the organellar fission machinery. Here, we have examined the role of Pex11p phosphorylation in the yeast Hansenula polymorpha. Employing mass spectrometry, we demonstrate that HpPex11p is also phosphorylated on a Serine residue present at a similar position to that of ScPex11p and PpPex11p. Furthermore, through the use of mutants designed to mimic both phosphorylated and unphosphorylated forms of HpPex11p, we have investigated the role of this post-translational modification. Our data demonstrate that mutations to the phosphorylation site do not disturb the function of Pex11p in peroxisomal fission, nor do they alter the localization of Pex11p. Also, no effect on peroxisome inheritance was observed. Taken together, these data lead us to conclude that peroxisomal fission in H. polymorpha is not modulated by phosphorylation of Pex11p.

No MeSH data available.