title: Quantification of labile heme in live malaria parasites using a genetically encoded biosensor.
abstract: Heme is ubiquitous, yet relatively little is known about the maintenance of labile pools of this cofactor, which likely ensures its timely bioavailability for proper cellular function. Quantitative analysis of labile heme is of fundamental importance to understanding how nature preserves access to the diverse chemistry heme enables, while minimizing cellular damage caused by its redox activity. Here, we have developed and characterized a protein-based sensor that undergoes fluorescence quenching upon heme binding. By genetically encoding this sensor in the human malarial parasite,Plasmodium falciparum, we have quantified cytosolic labile heme levels in intact, blood-stage parasites. Our findings indicate that a labile heme pool (∼1.6 µM) is stably maintained throughout parasite development within red blood cells, even during a period coincident with extensive hemoglobin degradation by the parasite. We also find that the heme-binding antimalarial drug chloroquine specifically increases labile cytosolic heme, indicative of dysregulation of this homeostatic pool that may be a relevant component of the antimalarial activity of this compound class. We propose that use of this technology under various environmental perturbations inP. falciparumcan yield quantitative insights into fundamental heme biology.
authors.affiliation: Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139.
authors.forename: James R
authors.initials: JR
authors.lastname: Abshire
authors.affiliation: Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139.
authors.forename: Christopher J
authors.initials: CJ
authors.lastname: Rowlands
authors.affiliation: Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139.
authors.forename: Suresh M
authors.initials: SM
authors.lastname: Ganesan
authors.affiliation: Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139.|Department of Mechanical Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139.
authors.forename: Peter T C
authors.initials: PT
authors.lastname: So
authors.affiliation: Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139; jcniles@mit.edu.
authors.forename: Jacquin C
authors.initials: JC
authors.lastname: Niles
mesh_terms: D000818:Animals; D015374:Biosensing Techniques / Q000295:instrumentation / Q000379:methods; D015870:Gene Expression; D017930:Genes, Reporter; D006418:Heme / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D008856:Microscopy, Fluorescence; D010961:Plasmodium / Q000235:genetics / Q000378:metabolism*; D010963:Plasmodium falciparum / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D008164:Luminescent Proteins; D011993:Recombinant Fusion Proteins; D006418:Heme
keywords: Plasmodium falciparum; genetically encoded biosensor; heme; heme sensor; malaria