scholarly journals The 3-phosphoinositide-dependent protein kinase 1 is an essential upstream activator of protein kinase A in malaria parasites

2021 ◽  
Author(s):  
Eva Hitz ◽  
Natalie Wiedemar ◽  
Armin Passecker ◽  
Nicolas M. B. Brancucci ◽  
Ioannis Vakonakis ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Protein Kinase A ◽  
Blood Stage ◽  
Targeted Mutagenesis ◽  
Missense Mutations ◽  
Dependent Protein Kinase ◽  
Gene Encoding ◽  
Upstream Regulator ◽  
Dependent Protein ◽  
Kinase A

Cyclic AMP (cAMP) signalling is crucial for the propagation of asexual malaria blood stage parasites. Recent work on Plasmodium falciparum demonstrated that phosphorylation of the invasion ligand AMA1 by the catalytic subunit of cAMP-dependent protein kinase A (PfPKAc) is an essential step during parasite invasion into red blood cells. However, the exact mechanisms regulating PfPKAc activity are only partially understood and PfPKAc function has not been extensively studied in gametocytes, the sexual blood stage forms that are essential for malaria transmission. By studying a conditional PfPKAc knockdown mutant, we confirm the essential role for PfPKAc in erythrocyte invasion and demonstrate that PfPKAc is involved in regulating gametocyte deformability. Interestingly, we observed that the conditional overexpression of PfPKAc also caused a profound lethal phenotype by preventing intra-erythrocytic parasite multiplication. Whole genome sequencing of parasites selected to tolerate increased PfPKAc expression levels identified missense mutations exclusively in the gene encoding the putative parasite orthologue of 3-phosphoinositide-dependent protein kinase-1 (PfPDK1). Using targeted mutagenesis, we show that PfPDK1 is essential for PfPKAc activation, most likely by phosphorylating T189 in the PfPKAc activation loop. In summary, our results corroborate the importance of tight regulation of PfPKA signalling for parasite survival and identify PfPDK1 as a crucial upstream regulator in this pathway and potential new drug target.

scholarly journals The 3-phosphoinositide–dependent protein kinase 1 is an essential upstream activator of protein kinase A in malaria parasites

PLoS Biology ◽  
2021 ◽  
Vol 19 (12) ◽  
pp. e3001483
Author(s):  
Eva Hitz ◽  
Natalie Wiedemar ◽  
Armin Passecker ◽  
Beatriz A. S. Graça ◽  
Christian Scheurer ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Protein Kinase A ◽  
Cyclic Adenosine Monophosphate ◽  
Adenosine Monophosphate ◽  
Blood Stage ◽  
Targeted Mutagenesis ◽  
Missense Mutations ◽  
Dependent Protein Kinase ◽  
Dependent Protein ◽  
Kinase A

Cyclic adenosine monophosphate (cAMP)-dependent protein kinase A (PKA) signalling is essential for the proliferation of Plasmodium falciparum malaria blood stage parasites. The mechanisms regulating the activity of the catalytic subunit PfPKAc, however, are only partially understood, and PfPKAc function has not been investigated in gametocytes, the sexual blood stage forms that are essential for malaria transmission. By studying a conditional PfPKAc knockdown (cKD) mutant, we confirm the essential role for PfPKAc in erythrocyte invasion by merozoites and show that PfPKAc is involved in regulating gametocyte deformability. We furthermore demonstrate that overexpression of PfPKAc is lethal and kills parasites at the early phase of schizogony. Strikingly, whole genome sequencing (WGS) of parasite mutants selected to tolerate increased PfPKAc expression levels identified missense mutations exclusively in the gene encoding the parasite orthologue of 3-phosphoinositide–dependent protein kinase-1 (PfPDK1). Using targeted mutagenesis, we demonstrate that PfPDK1 is required to activate PfPKAc and that T189 in the PfPKAc activation loop is the crucial target residue in this process. In summary, our results corroborate the importance of tight regulation of PfPKA signalling for parasite survival and imply that PfPDK1 acts as a crucial upstream regulator in this pathway and potential new drug target.

scholarly journals AniScan Using Extracellular Cyclic AMP-Dependent Protein Kinase A as a Serum Biomarker Assay for the Diagnosis of Malignant Tumors in Dogs

Sensors ◽  
2020 ◽  
Vol 20 (15) ◽  
pp. 4075
Author(s):  
Ji-Eun Lee ◽  
Woo-Jin Song ◽  
Hunjoo Lee ◽  
Byung-Gak Kim ◽  
Taeho Kim ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Protein Kinase A ◽  
Cancer Detection ◽  
Malignant Tumors ◽  
Cost Effective ◽  
Benign Tumors ◽  
Dependent Protein Kinase ◽  
Diagnostic Significance ◽  
Dependent Protein ◽  
Kinase A

The early detection of tumors improves chances of decreased morbidity and prolonged survival. Serum biomarkers are convenient to use and have several advantages over other approaches, such as accuracy and straightforward protocols. Reliable biomarkers from easily accessible sources are warranted for the development of cost-effective assays for routine screening, particularly in veterinary medicine. Extracellular c-AMP-dependent protein kinase A (ECPKA) is a cytosolic leakage enzyme. The diagnostic accuracy of detecting autoantibodies against ECPKA was found to be higher than that of ECPKA activity from enzymatic assays, which use a complicated method. Here, we investigated the diagnostic significance of measuring serum ECPKA autoantibody levels using an in-house kit (AniScan cancer detection kit; Biattic, Anyang, Korea). We used sera from 550 dogs, including healthy dogs and those with malignant and benign tumors. Serum ECPKA and immunoglobulin G were determined using the AniScan cancer detection kit. ECPKA autoantibody levels were significantly higher (p < 0.01) in malignant tumors than in benign tumors, non-tumor diseases, and healthy controls. On the basis of sensitivity and specificity values, AniScan ECPKA is a rapid and easy-to-use assay that can be applied to screen malignant tumors from benign tumors or other diseases in dogs.

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