The Plasmodium falciparum MESA erythrocyte cytoskeleton-binding (MEC)motif binds to erythrocyte ankyrin

Geoffrey Kimiti Kilili, Bikash Shakya, Patrick T. Dolan, Ling Wang, Monica L. Husby, Robert Stahelin, Ernesto S. Nakayasu, Douglas J. LaCount

Research output: Contribution to journalArticle

Abstract

The MESA erythrocyte cytoskeleton binding (MEC)motif is a 13-amino acid sequence found in 14 exported Plasmodium falciparum proteins. First identified in the P. falciparum Mature-parasite-infected Erythrocyte Surface Antigen (MESA), the MEC motif is sufficient to target proteins to the infected red blood cell cytoskeleton. To identify host cell targets, purified MESA MEC motif was incubated with a soluble extract from uninfected erythrocytes, precipitated and subjected to mass spectrometry. The most abundant co-purifying protein was erythrocyte ankyrin (ANK1). A direct interaction between the MEC motif and ANK1 was independently verified using co-purification experiments, the split-luciferase assay, and the yeast two-hybrid assay. A systematic mutational analysis of the core MEC motif demonstrated a critical role for the conserved aspartic acid residue at the C-terminus of the MEC motif for binding to both erythrocyte inside-out vesicles and to ANK1. Using a panel of ANK1 constructs, the MEC motif binding site was localized to the ZU5C domain, which has no known function. The MEC motif had no impact on erythrocyte deformability when introduced into uninfected erythrocyte ghosts, suggesting the MEC motif's primary function is to target exported proteins to the cytoskeleton. Finally, we show that PF3D7_0402100 (PFD0095c)binds to ANK1 and band 4.1, likely through its MEC and PHIST motifs, respectively. In conclusion, we have provided multiple lines of evidence that the MEC motif binds to erythrocyte ANK1.

Original languageEnglish (US)
Article number111189
JournalMolecular and Biochemical Parasitology
Volume231
DOIs
StatePublished - Jul 1 2019
Externally publishedYes

Fingerprint

Ankyrins
Erythrocyte Membrane
Plasmodium falciparum
Surface Antigens
Parasites
Erythrocytes
Cytoskeleton
Proteins
Erythrocyte Deformability
Two-Hybrid System Techniques
Luciferases
Aspartic Acid
Amino Acid Sequence

Keywords

  • Ankyrin
  • Band 4.1
  • Exported protein
  • Host-pathogen interaction
  • MESA
  • PF3D7_0402100
  • PF3D7_0500800
  • Plasmodium

ASJC Scopus subject areas

  • Parasitology
  • Molecular Biology

Cite this

The Plasmodium falciparum MESA erythrocyte cytoskeleton-binding (MEC)motif binds to erythrocyte ankyrin. / Kilili, Geoffrey Kimiti; Shakya, Bikash; Dolan, Patrick T.; Wang, Ling; Husby, Monica L.; Stahelin, Robert; Nakayasu, Ernesto S.; LaCount, Douglas J.

In: Molecular and Biochemical Parasitology, Vol. 231, 111189, 01.07.2019.

Research output: Contribution to journalArticle

Kilili, Geoffrey Kimiti ; Shakya, Bikash ; Dolan, Patrick T. ; Wang, Ling ; Husby, Monica L. ; Stahelin, Robert ; Nakayasu, Ernesto S. ; LaCount, Douglas J. / The Plasmodium falciparum MESA erythrocyte cytoskeleton-binding (MEC)motif binds to erythrocyte ankyrin. In: Molecular and Biochemical Parasitology. 2019 ; Vol. 231.
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abstract = "The MESA erythrocyte cytoskeleton binding (MEC)motif is a 13-amino acid sequence found in 14 exported Plasmodium falciparum proteins. First identified in the P. falciparum Mature-parasite-infected Erythrocyte Surface Antigen (MESA), the MEC motif is sufficient to target proteins to the infected red blood cell cytoskeleton. To identify host cell targets, purified MESA MEC motif was incubated with a soluble extract from uninfected erythrocytes, precipitated and subjected to mass spectrometry. The most abundant co-purifying protein was erythrocyte ankyrin (ANK1). A direct interaction between the MEC motif and ANK1 was independently verified using co-purification experiments, the split-luciferase assay, and the yeast two-hybrid assay. A systematic mutational analysis of the core MEC motif demonstrated a critical role for the conserved aspartic acid residue at the C-terminus of the MEC motif for binding to both erythrocyte inside-out vesicles and to ANK1. Using a panel of ANK1 constructs, the MEC motif binding site was localized to the ZU5C domain, which has no known function. The MEC motif had no impact on erythrocyte deformability when introduced into uninfected erythrocyte ghosts, suggesting the MEC motif's primary function is to target exported proteins to the cytoskeleton. Finally, we show that PF3D7_0402100 (PFD0095c)binds to ANK1 and band 4.1, likely through its MEC and PHIST motifs, respectively. In conclusion, we have provided multiple lines of evidence that the MEC motif binds to erythrocyte ANK1.",
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AU - Kilili, Geoffrey Kimiti

AU - Shakya, Bikash

AU - Dolan, Patrick T.

AU - Wang, Ling

AU - Husby, Monica L.

AU - Stahelin, Robert

AU - Nakayasu, Ernesto S.

AU - LaCount, Douglas J.

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KW - PF3D7_0500800

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