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Fatty acid acylation regulates trafficking of the unusual Plasmodium falciparum calpain to the nucleolus.

Russo I, Oksman A, Goldberg DE - Mol. Microbiol. (2009)

Bottom Line: Palmitoylation status has an important role in dictating P. falciparum calpain localization.The targeting signals function in mammalian cells as well as in the parasite.P. falciparum calpain is a unique nucleolar protein with an interesting mechanism of targeting.

View Article: PubMed Central - PubMed

Affiliation: Howard Hughes Medical Institute, Washington University School of Medicine, Department of Molecular Microbiology, St Louis, Missouri 63110, USA.

ABSTRACT
The Plasmodium falciparum genome encodes a single calpain. By generating P. falciparum clones expressing C-terminally tagged calpain, we localized this protein to the nucleolus. Pf_calpain possesses an unusual and long N-terminal domain in which we identified three subregions that are highly conserved among Plasmodium species. Two have putative targeting signals: a myristoylation motif and a nuclear localization sequence. We assessed their functionality. Our data show that the nuclear localization sequence is an active nuclear import motif that contains an embedded signal conferring nucleolar localization on various chimeras. The N-terminus is myristoylated at Gly2 and palmitoylated at Cys3 and Cys22. Palmitoylation status has an important role in dictating P. falciparum calpain localization. The targeting signals function in mammalian cells as well as in the parasite. P. falciparum calpain is a unique nucleolar protein with an interesting mechanism of targeting.

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Related in: MedlinePlus

Features and conservation of Plasmodium calpains. A. Calpain conservation in Plasmodium species and Pf_calpain AT content. In the graph, the x-axis shows amino acid position along the alignment of Fig. S2. The black line is the percentage of similarity among calpains encoded by P. falciparum, reichenowi, gallinaceum, yoelii yoelii, berghei, chabaudi, knowlesi and vivax (Table S1). The degree of conservation was calculated in a 20-amino-acid sliding window throughout the aligned calpain sequences. AT percentage of each P. falciparum subdomain is graphically represented by the green shading. The relative positions of nine low-complexity regions (LCX 1–9) detected in 3D7, are indicated as dark green bars, the height of which indicates AT% content of the corresponding DNA fragment. LCXs 5, 7 and 9 correspond to DNA repeats. Six highly conserved (HC 1–6) and five poorly conserved (PC 1–5) regions are depicted in the lower green bar, in scale to the graph. B. Diagram of Pf_calpain domains and subdomains. Regions are in scale to the graph in (A). The highly conserved regions in the N-terminal domain are, in order, a conserved myristoylation consensus sequence (Myr seq), a subdomain common to some nucleic acid-interacting proteins (NA interac) and a putative nuclear localization signal (NLS). The central domain has homology (E-value 9.59 e−34) to calpain catalytic domains (domain II) containing the catalytic triad C, H and N (black arrows indicate their relative positions). The C-terminus shows homology (E-value 1.84 e−3) to calpain domain III. Positions of repeats (shaded boxes) and peptides used to generate rabbit antisera #19 and #35 (bars) are shown.
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fig04: Features and conservation of Plasmodium calpains. A. Calpain conservation in Plasmodium species and Pf_calpain AT content. In the graph, the x-axis shows amino acid position along the alignment of Fig. S2. The black line is the percentage of similarity among calpains encoded by P. falciparum, reichenowi, gallinaceum, yoelii yoelii, berghei, chabaudi, knowlesi and vivax (Table S1). The degree of conservation was calculated in a 20-amino-acid sliding window throughout the aligned calpain sequences. AT percentage of each P. falciparum subdomain is graphically represented by the green shading. The relative positions of nine low-complexity regions (LCX 1–9) detected in 3D7, are indicated as dark green bars, the height of which indicates AT% content of the corresponding DNA fragment. LCXs 5, 7 and 9 correspond to DNA repeats. Six highly conserved (HC 1–6) and five poorly conserved (PC 1–5) regions are depicted in the lower green bar, in scale to the graph. B. Diagram of Pf_calpain domains and subdomains. Regions are in scale to the graph in (A). The highly conserved regions in the N-terminal domain are, in order, a conserved myristoylation consensus sequence (Myr seq), a subdomain common to some nucleic acid-interacting proteins (NA interac) and a putative nuclear localization signal (NLS). The central domain has homology (E-value 9.59 e−34) to calpain catalytic domains (domain II) containing the catalytic triad C, H and N (black arrows indicate their relative positions). The C-terminus shows homology (E-value 1.84 e−3) to calpain domain III. Positions of repeats (shaded boxes) and peptides used to generate rabbit antisera #19 and #35 (bars) are shown.

Mentions: The averaged overall homology among Plasmodium calpain amino acid sequences is 67% and the identity 55%. However, the homology is not evenly distributed along the protein. Therefore, in order to analyse the conservation distribution, we designed a PERL program that calculates homology in a sliding 20-amino-acid window. The result shows that Pf_calpain has six highly conserved regions (HC) interspersed with five poorly conserved ones (PC) (Fig. 4A).


Fatty acid acylation regulates trafficking of the unusual Plasmodium falciparum calpain to the nucleolus.

Russo I, Oksman A, Goldberg DE - Mol. Microbiol. (2009)

Features and conservation of Plasmodium calpains. A. Calpain conservation in Plasmodium species and Pf_calpain AT content. In the graph, the x-axis shows amino acid position along the alignment of Fig. S2. The black line is the percentage of similarity among calpains encoded by P. falciparum, reichenowi, gallinaceum, yoelii yoelii, berghei, chabaudi, knowlesi and vivax (Table S1). The degree of conservation was calculated in a 20-amino-acid sliding window throughout the aligned calpain sequences. AT percentage of each P. falciparum subdomain is graphically represented by the green shading. The relative positions of nine low-complexity regions (LCX 1–9) detected in 3D7, are indicated as dark green bars, the height of which indicates AT% content of the corresponding DNA fragment. LCXs 5, 7 and 9 correspond to DNA repeats. Six highly conserved (HC 1–6) and five poorly conserved (PC 1–5) regions are depicted in the lower green bar, in scale to the graph. B. Diagram of Pf_calpain domains and subdomains. Regions are in scale to the graph in (A). The highly conserved regions in the N-terminal domain are, in order, a conserved myristoylation consensus sequence (Myr seq), a subdomain common to some nucleic acid-interacting proteins (NA interac) and a putative nuclear localization signal (NLS). The central domain has homology (E-value 9.59 e−34) to calpain catalytic domains (domain II) containing the catalytic triad C, H and N (black arrows indicate their relative positions). The C-terminus shows homology (E-value 1.84 e−3) to calpain domain III. Positions of repeats (shaded boxes) and peptides used to generate rabbit antisera #19 and #35 (bars) are shown.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2746569&req=5

fig04: Features and conservation of Plasmodium calpains. A. Calpain conservation in Plasmodium species and Pf_calpain AT content. In the graph, the x-axis shows amino acid position along the alignment of Fig. S2. The black line is the percentage of similarity among calpains encoded by P. falciparum, reichenowi, gallinaceum, yoelii yoelii, berghei, chabaudi, knowlesi and vivax (Table S1). The degree of conservation was calculated in a 20-amino-acid sliding window throughout the aligned calpain sequences. AT percentage of each P. falciparum subdomain is graphically represented by the green shading. The relative positions of nine low-complexity regions (LCX 1–9) detected in 3D7, are indicated as dark green bars, the height of which indicates AT% content of the corresponding DNA fragment. LCXs 5, 7 and 9 correspond to DNA repeats. Six highly conserved (HC 1–6) and five poorly conserved (PC 1–5) regions are depicted in the lower green bar, in scale to the graph. B. Diagram of Pf_calpain domains and subdomains. Regions are in scale to the graph in (A). The highly conserved regions in the N-terminal domain are, in order, a conserved myristoylation consensus sequence (Myr seq), a subdomain common to some nucleic acid-interacting proteins (NA interac) and a putative nuclear localization signal (NLS). The central domain has homology (E-value 9.59 e−34) to calpain catalytic domains (domain II) containing the catalytic triad C, H and N (black arrows indicate their relative positions). The C-terminus shows homology (E-value 1.84 e−3) to calpain domain III. Positions of repeats (shaded boxes) and peptides used to generate rabbit antisera #19 and #35 (bars) are shown.
Mentions: The averaged overall homology among Plasmodium calpain amino acid sequences is 67% and the identity 55%. However, the homology is not evenly distributed along the protein. Therefore, in order to analyse the conservation distribution, we designed a PERL program that calculates homology in a sliding 20-amino-acid window. The result shows that Pf_calpain has six highly conserved regions (HC) interspersed with five poorly conserved ones (PC) (Fig. 4A).

Bottom Line: Palmitoylation status has an important role in dictating P. falciparum calpain localization.The targeting signals function in mammalian cells as well as in the parasite.P. falciparum calpain is a unique nucleolar protein with an interesting mechanism of targeting.

View Article: PubMed Central - PubMed

Affiliation: Howard Hughes Medical Institute, Washington University School of Medicine, Department of Molecular Microbiology, St Louis, Missouri 63110, USA.

ABSTRACT
The Plasmodium falciparum genome encodes a single calpain. By generating P. falciparum clones expressing C-terminally tagged calpain, we localized this protein to the nucleolus. Pf_calpain possesses an unusual and long N-terminal domain in which we identified three subregions that are highly conserved among Plasmodium species. Two have putative targeting signals: a myristoylation motif and a nuclear localization sequence. We assessed their functionality. Our data show that the nuclear localization sequence is an active nuclear import motif that contains an embedded signal conferring nucleolar localization on various chimeras. The N-terminus is myristoylated at Gly2 and palmitoylated at Cys3 and Cys22. Palmitoylation status has an important role in dictating P. falciparum calpain localization. The targeting signals function in mammalian cells as well as in the parasite. P. falciparum calpain is a unique nucleolar protein with an interesting mechanism of targeting.

Show MeSH
Related in: MedlinePlus