Intramembrane proteolytic cleavage by human signal peptide peptidase like 3 and malaria signal peptide peptidase

Department of Neuroscience, Mayo Clinic Jacksonville, Mayo Clinic College of Medicine, 4500 San Pablo Rd., Jacksonville, Florida 32224, USA.
The FASEB Journal (Impact Factor: 5.04). 09/2006; 20(10):1671-9. DOI: 10.1096/fj.06-5762com
Source: PubMed


Signal peptide peptidase (SPP) is an intramembrane cleaving protease (I-CLiP) identified by its cleavage of several type II membrane signal peptides. To date, only human SPP has been directly shown to have proteolytic activity. Here we demonstrate that the most closely related human homologue of SPP, signal peptide peptidase like 3 (SPPL3), cleaves a SPP substrate, but a more distantly related homologue, signal peptide peptidase like 2b (SPPL2b), does not. These data provide strong evidence that the SPP and SPPL3 have conserved active sites and suggest that the active sites SPPL2b is distinct. We have also synthesized a cDNA designed to express the single SPP gene present in Plasmodium falciparum and cloned this into a mammalian expression vector. When the malaria SPP protein is expressed in mammalian cells it cleaves a SPP substrate. Notably, several human SPP inhibitors block the proteolytic activity of malarial SPP (mSPP). Studies from several model organisms that express multiple SPP homologs demonstrate that the silencing of a single SPP homologue is lethal. Based on these data, we hypothesize that mSPP is a potential a novel therapeutic target for malaria.

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Available from: Thomas Kukar, May 26, 2014
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    • "While HSP101 is essential for parasite protein export and translocation, a functional role of its signal peptide is not yet known [26]. We utilized a transient transfection assay previously used to demonstrate the enzyme activity of PfSPP against a model substrate (Fig. 3A) [4]. The cleavage assay demonstrated cleavage of HSP101 signal peptide by PfSPP (Fig. 3B). "
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    ABSTRACT: Previously we described the identification of a Plasmodium falciparum signal peptide peptidase (PfSPP) functioning at the blood stage of malaria infection. Our studies also demonstrated that mammalian SPP inhibitors prevent malaria parasite growth at the late-ring/early trophozoite stage of intra-erythrocytic development. Consistent with its role in development, we tested the hypothesis that PfSPP functions at the endoplasmic reticulum of Plasmodium falciparum where it cleaves membrane-bound signal peptides generated following the enzyme activity of signal peptidase. The localization of PfSPP to the endoplasmic reticulum was confirmed by immunofluorescence microscopy and immunogold electron microscopy. Biochemical analysis indicated the existence of monomer and dimer forms of PfSPP in the parasite lysate. A comprehensive bioinformatics screen identified several candidate PfSPP substrates in the parasite genome. Using an established transfection based in vivo luminescence assay, malaria heat shock protein 101 (HSP101) was identified as a substrate of PfSPP, and partial inhibition of PfSPP correlated with the emergence of gametocytes. This finding unveils the first known substrate of PfSPP, and provides new perspectives for the function of intra-membrane proteolysis at the erythrocyte stage of malaria parasite life cycle.
    Full-text · Article · Jul 2014 · Biochemical and Biophysical Research Communications
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    • "Two homologs, SPP-like proteases, SPPL2a and SPPL2b, apparently cleave tumor necrosis factor α (Fluhrer et al., 2006) and the dementia-associated Bri2 protein (Martin et al., 2008), although the biological roles of these proteolytic events are unknown. SPP has also been identified in malarial parasites and may represent a worthwhile therapeutic target (Nyborg et al., 2006). "
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    ABSTRACT: Presenilin is the catalytic component of gamma-secretase, a complex aspartyl protease and a founding member of intramembrane-cleaving proteases. gamma-Secretase is involved in the pathogenesis of Alzheimer's disease and a top target for therapeutic intervention. However, the protease complex processes a variety of transmembrane substrates, including the Notch receptor, raising concerns about toxicity. Nevertheless, gamma-secretase inhibitors and modulators have been identified that allow Notch processing and signaling to continue, and promising compounds are entering clinical trials. Molecular and biochemical studies offer a model for how this protease hydrolyzes transmembrane domains in the confines of the lipid bilayer. Progress has also been made toward structure elucidation of presenilin and the gamma-secretase complex by electron microscopy as well as by studying cysteine-mutant presenilins. The signal peptide peptidase (SPP) family of proteases are distantly related to presenilins. However, the SPPs work as single polypeptides without the need for cofactors and otherwise appear to be simple model systems for presenilin in the gamma-secretase complex. SPP biology, structure, and inhibition will also be discussed.
    Preview · Article · Aug 2010 · Biological Chemistry
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    • "Here we report that LY411,575 reduces development of P. berghei ANKA in human hepatoma cells, primary hepatocytes and mouse livers. Though developed as a potent GSI, LY411,575 also blocks the activity of all human SPP homologues and P. falciparum SPP with reasonable potency [18], [19]. LY411575 is not unique and one of many compounds developed as potential therapeutic agents for the treatment of Alzheimer's disease (AD) that appear capable of blocking GSI and SPP activity. "
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    ABSTRACT: The liver stage of Plasmodium's life cycle is the first, obligatory step in malaria infection. Decreasing the hepatic burden of Plasmodium infection decreases the severity of disease and constitutes a promising strategy for malaria prophylaxis. The efficacy of the gamma-secretase and signal peptide peptidase inhibitor LY411,575 in targeting Plasmodium liver stages was evaluated both in human hepatoma cell lines and in mouse primary hepatocytes. LY411,575 was found to prevent Plasmodium's normal development in the liver, with an IC(50) of approximately 80 nM, without affecting hepatocyte invasion by the parasite. In vivo results with a rodent model of malaria showed that LY411,575 decreases the parasite load in the liver and increases by 55% the resistance of mice to cerebral malaria, one of the most severe malaria-associated syndromes. Our data show that LY411,575 does not exert its effect via the Notch signaling pathway suggesting that it may interfere with Plasmodium development through an inhibition of the parasite's signal peptide peptidase. We therefore propose that selective signal peptide peptidase inhibitors could be potentially used for preventive treatment of malaria in humans.
    Full-text · Article · Feb 2009 · PLoS ONE
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