Genetic disruption of nucleoside transporter 4 reveals its critical roles in malaria parasite sporozoite functions

Deveci, Gözde; Kamil, Mohd; Kına, Ümit Yaşar; Temel, Binnur Aydogan; Aly, A. S.

doi:10.1080/20477724.2022.2112880

Cited by 3 publications

(6 citation statements)

References 22 publications

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“…By contrast, knockdown of PfNT4, PfGDH3 and PfCA did not have a significant impact on parasite growth, consistent with previous studies for PfNT4 and PfGDH3 19,20 .…”

Section: Further Characterisation Of Osm-s-106 Targetssupporting

confidence: 91%

“…A similar base level sensitisation has been observed for a PfHsp70 inhibitor in an aptamer-regulated PfHsp70 line 34 . PfNT4 is a putative purine transporter that has been shown to be dispensable for blood stage growth but required for sporozoite colonization of salivary glands 19,35,36 . It is possible that PfNT4 transports OSM-S-106 away from its primary site of action and that mutations in PfNT4 enhance the accumulation of OSM-S-106.…”

Section: Discussionmentioning

confidence: 99%

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Reaction hijacking inhibition of Plasmodium falciparum asparagine tRNA synthetase

Tilley,

Xie,

Wang

et al. 2023

Preprint

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Malaria poses an enormous threat to human health. With ever increasing resistance to currently deployed drugs, breakthrough compounds with novel mechanisms of action are urgently needed. Here, we explore pyrimidine-based sulfonamides as a new low molecular weight inhibitor class with drug-like physical parameters and a synthetically accessible scaffold. We show that the exemplar, OSM-S-106, has potent activity against parasite cultures, low mammalian cell toxicity and low propensity for resistance development. In vitro evolution of resistance using a slow ramp-up approach pointed to the Plasmodium falciparum cytoplasmic asparaginyl tRNA synthetase (PfAsnRS) as the target, consistent with our finding that OSM-S-106 inhibits protein translation and activates the amino acid starvation response. Targeted mass spectrometry confirms that OSM-S-106 is a pro-inhibitor and that inhibition of PfAsnRS occurs via enzyme-mediated production of an Asn-OSM-S-106 adduct. Human AsnRS is much less susceptible to this reaction hijacking mechanism. X-ray crystallographic studies of human AsnRS in complex with inhibitor adducts and docking of pro-inhibitors into a model of Asn-tRNA-bound PfAsnRS provide insights into the structure activity relationship and the selectivity mechanism.

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“…By contrast, knockdown of PfNT4, PfGDH3 and PfCA did not have a significant impact on parasite growth, consistent with previous studies for PfNT4 and PfGDH3 19,20 .…”

Section: Further Characterisation Of Osm-s-106 Targetssupporting

confidence: 91%

Section: Discussionmentioning

confidence: 99%

Reaction hijacking inhibition of Plasmodium falciparum asparagine tRNA synthetase

Tilley,

Xie,

Wang

et al. 2023

Preprint

View full text Add to dashboard Cite

show abstract

“…A similar base level sensitization has been observed for a Pf Hsp70 inhibitor in an aptamer-regulated Pf Hsp70 line 37 . Pf NT4 is a putative purine transporter that has been shown to be dispensable for blood stage growth but required for sporozoite colonization of salivary glands 20 , 38 , 39 . It is possible that Pf NT4 transports OSM-S-106 away from its primary site of action and that mutations in Pf NT4 enhance the accumulation of OSM-S-106.…”

Section: Discussionmentioning

confidence: 99%

Reaction hijacking inhibition of Plasmodium falciparum asparagine tRNA synthetase

Xie,

Wang,

Morton

et al. 2024

Nat Commun

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Malaria poses an enormous threat to human health. With ever increasing resistance to currently deployed drugs, breakthrough compounds with novel mechanisms of action are urgently needed. Here, we explore pyrimidine-based sulfonamides as a new low molecular weight inhibitor class with drug-like physical parameters and a synthetically accessible scaffold. We show that the exemplar, OSM-S-106, has potent activity against parasite cultures, low mammalian cell toxicity and low propensity for resistance development. In vitro evolution of resistance using a slow ramp-up approach pointed to the Plasmodium falciparum cytoplasmic asparaginyl-tRNA synthetase (PfAsnRS) as the target, consistent with our finding that OSM-S-106 inhibits protein translation and activates the amino acid starvation response. Targeted mass spectrometry confirms that OSM-S-106 is a pro-inhibitor and that inhibition of PfAsnRS occurs via enzyme-mediated production of an Asn-OSM-S-106 adduct. Human AsnRS is much less susceptible to this reaction hijacking mechanism. X-ray crystallographic studies of human AsnRS in complex with inhibitor adducts and docking of pro-inhibitors into a model of Asn-tRNA-bound PfAsnRS provide insights into the structure-activity relationship and the selectivity mechanism.

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“…The diluted blood samples were placed in a hemocytometer and incubated at room temperature for 10 min. The exflagellation centers in Pbp230p(−) , Pbtmem33(−) , Pbtmem33‐mNG , and Pb WT were counted at 400x magnification (Deveci et al., 2022). At the same time, blood from tail puncture was thin smeared and stained with Giemsa to determine the ratio of male versus female gametocytes.…”

Section: Methodsmentioning

confidence: 99%

“…The sequences of the primer pairs for the preparation of donor DNA plasmid and for the integration-specific diagnostic PCR are listed in Table S1. PbWT were counted at 400x magnification (Deveci et al, 2022).…”

Section: Analysis Of Amino Acid Sequencesmentioning

confidence: 99%

Endoplasmic reticulum localized TMEM33 domain‐containing protein is crucial for all life cycle stages of the malaria parasite

Kamil,

Kina,

Atmaca

et al. 2024

Molecular Microbiology

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Endoplasmic reticulum (ER) plays a pivotal role in the regulation of stress responses in multiple eukaryotic cells. However, little is known about the effector mechanisms that regulate stress responses in ER of the malaria parasite. Herein, we aimed to identify the importance of a transmembrane protein 33 (TMEM33)‐domain‐containing protein in life cycle of the rodent malaria parasite Plasmodium berghei. TMEM33 is an ER membrane‐resident protein that is involved in regulating stress responses in various eukaryotic cells. A C‐terminal tagged TMEM33 was localized in the ER throughout the blood and mosquito stages of development. Targeted deletion of TMEM33 confirmed its importance for asexual blood stages and ookinete development, in addition to its essential role for sporozoite infectivity in the mammalian host. Pilot scale analysis shows that the loss of TMEM33 results in the initiation of ER stress response and induction of autophagy. Our findings conclude an important role of TMEM33 in the development of all life cycle stages of the malaria parasite, which indicates its potential as an antimalarial target.

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Genetic disruption of nucleoside transporter 4 reveals its critical roles in malaria parasite sporozoite functions

Cited by 3 publications

References 22 publications

Reaction hijacking inhibition of Plasmodium falciparum asparagine tRNA synthetase

Reaction hijacking inhibition of Plasmodium falciparum asparagine tRNA synthetase

Reaction hijacking inhibition of Plasmodium falciparum asparagine tRNA synthetase

Endoplasmic reticulum localized TMEM33 domain‐containing protein is crucial for all life cycle stages of the malaria parasite

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