Summary of Study ST002926
This data is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench, https://www.metabolomicsworkbench.org, where it has been assigned Project ID PR001818. The data can be accessed directly via it's Project DOI: 10.21228/M8QH8M This work is supported by NIH grant, U2C- DK119886.
See: https://www.metabolomicsworkbench.org/about/howtocite.php
This study contains a large results data set and is not available in the mwTab file. It is only available for download via FTP as data file(s) here.
| Study ID | ST002926 |
| Study Title | Multi-“omics” analysis reveals the orphan P. falciparum protein kinase PfPK8 regulates multi-gene family expression |
| Study Summary | Protein kinases are a large group of proteins that serve regulatory and signalling functions in eukaryotic cells. Whilst kinases can be readily identified by highly conserved kinase domains, the downstream function of many protein kinases remains unknown, even more so for kinases of divergent eukaryotes, such as the Plasmodium parasites that cause malaria. PfPK8 (PF3D7_0203100) is an orphan kinase in Plasmodium falciparum that bears some homology to STE kinases but has no known function. To reveal the function of PfPK8 we investigated PfPK8 knockout parasites with an untargeted multi-omics workflow. Phosphoproteomics analysis identified six putative phosphorylation targets in the parasite nucleus, including another kinase, a histone acetyltransferase, and three transcription-associated proteins, including the transcription factor AP2-12 (PF3D7_1239200). Untargeted metabolomics and proteomics analysis demonstrated no impact of the PfPK8 knockout on metabolism but revealed differential regulation of exported surface proteins from multi-gene families. Transcriptomics analysis confirmed differential expression of these multi-gene family proteins, particularly de-repression of var genes encoding PfEMP1 variants. DAP-seq analysis of genes bound to the AP2-12 transcription factor also identified significant enrichment of var genes, with significant overlap between the group B/C and C var genes enriched in both the PfPK8-KO transcriptomics and AP2-12 DAP-seq analysis. Overall, this study revealed that the primary function of PfPK8 is to regulate transcription of antigenic variant genes via phosphorylation of nuclear targets including the AP2-12 transcription factor. |
| Institute | Monash University |
| Last Name | Siddiqui |
| First Name | Ghizal |
| Address | 381 Royal Parade, Parkville, Melbourne, Victoria, 3052, Australia |
| ghizal.siddiqui@monash.edu | |
| Phone | 99039282 |
| Submit Date | 2023-10-11 |
| Raw Data Available | Yes |
| Raw Data File Type(s) | raw(Thermo) |
| Analysis Type Detail | LC-MS |
| Release Date | 2023-10-26 |
| Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Project:
| Project ID: | PR001818 |
| Project DOI: | doi: 10.21228/M8QH8M |
| Project Title: | Multi-“omics” analysis reveals the orphan P. falciparum protein kinase PfPK8 regulates multi-gene family expression |
| Project Summary: | Protein kinases are a large group of proteins that serve regulatory and signalling functions in eukaryotic cells. Whilst kinases can be readily identified by highly conserved kinase domains, the downstream function of many protein kinases remains unknown, even more so for kinases of divergent eukaryotes, such as the Plasmodium parasites that cause malaria. PfPK8 (PF3D7_0203100) is an orphan kinase in Plasmodium falciparum that bears some homology to STE kinases but has no known function. To reveal the function of PfPK8 we investigated PfPK8 knockout parasites with an untargeted multi-omics workflow. Phosphoproteomics analysis identified six putative phosphorylation targets in the parasite nucleus, including another kinase, a histone acetyltransferase, and three transcription-associated proteins, including the transcription factor AP2-12 (PF3D7_1239200). Untargeted metabolomics and proteomics analysis demonstrated no impact of the PfPK8 knockout on metabolism but revealed differential regulation of exported surface proteins from multi-gene families. Transcriptomics analysis confirmed differential expression of these multi-gene family proteins, particularly de-repression of var genes encoding PfEMP1 variants. DAP-seq analysis of genes bound to the AP2-12 transcription factor also identified significant enrichment of var genes, with significant overlap between the group B/C and C var genes enriched in both the PfPK8-KO transcriptomics and AP2-12 DAP-seq analysis. Overall, this study revealed that the primary function of PfPK8 is to regulate transcription of antigenic variant genes via phosphorylation of nuclear targets including the AP2-12 transcription factor. |
| Institute: | Monash University |
| Last Name: | Siddiqui |
| First Name: | Ghizal |
| Address: | 381 Royal Parade, Parkville, Melbourne, Victoria, 3052, Australia |
| Email: | ghizal.siddiqui@monash.edu |
| Phone: | 99039282 |
Subject:
| Subject ID: | SU003039 |
| Subject Type: | Cultured cells |
| Subject Species: | Plasmodium falciparum |
| Taxonomy ID: | 5833 |
Factors:
Subject type: Cultured cells; Subject species: Plasmodium falciparum (Factor headings shown in green)
| mb_sample_id | local_sample_id | treatment |
|---|---|---|
| SA317891 | CP_10g2_2 | 10g2 clone PfPK8KO-1 |
| SA317892 | CP_10g2_3 | 10g2 clone PfPK8KO-2 |
| SA317893 | CP_10g2_4 | 10g2 clone PfPK8KO-3 |
| SA317894 | CP_10g2_5 | 10g2 clone PfPK8KO-4 |
| SA317895 | CP_11c_2 | 11c clone PfPK8KO-1 |
| SA317896 | CP_11c_3 | 11c clone PfPK8KO-2 |
| SA317897 | CP_11c_4 | 11c clone PfPK8KO-3 |
| SA317898 | CP_11c_5 | 11c clone PfPK8KO-4 |
| SA317899 | CP_2c_2 | 2c HA control-1 |
| SA317900 | CP_2c_3 | 2c HA control-2 |
| SA317901 | CP_2c_4 | 2c HA control-3 |
| SA317902 | CP_2c_5 | 2c HA control-4 |
| Showing results 1 to 12 of 12 |
Collection:
| Collection ID: | CO003032 |
| Collection Summary: | Highly synchronised parasites were harvested by magnet purification (Ribaut et al., 2008). 140 µL of 100% methanol was added to 5 x 107 parasites to extract metabolites. Samples were incubated at 4 °C for 1 h on a vortex shaker (Select Bioproducts). Samples were centrifuged at 21,000 x g for 5 min and the particle-free supernatant stored at – 80 °C until analysis. |
| Sample Type: | Blood (whole) |
Treatment:
| Treatment ID: | TR003048 |
| Treatment Summary: | untargeted metabolomics was performed on PfPK8-KO clone 10g2 and clone 11c, and on PfPK8-HA clone 2c (control). |
Sample Preparation:
| Sampleprep ID: | SP003045 |
| Sampleprep Summary: | Highly synchronised parasites were harvested by magnet purification (Ribaut et al., 2008). 140 µL of 100% methanol was added to 5 x 107 parasites to extract metabolites. Samples were incubated at 4 °C for 1 h on a vortex shaker (Select Bioproducts). Samples were centrifuged at 21,000 x g for 5 min and the particle-free supernatant stored at – 80 °C until analysis. |
Combined analysis:
| Analysis ID | AN004798 | AN004799 |
|---|---|---|
| Analysis type | MS | MS |
| Chromatography type | HILIC | HILIC |
| Chromatography system | QExactive | QExactive |
| Column | Merck SeQuant ZIC-pHILIC (150 x 4.6mm,5um) | Merck SeQuant ZIC-pHILIC (150 x 4.6mm,5um) |
| MS Type | ESI | ESI |
| MS instrument type | Orbitrap | Orbitrap |
| MS instrument name | Thermo Q Exactive Orbitrap | Thermo Q Exactive Orbitrap |
| Ion Mode | POSITIVE | NEGATIVE |
| Units | peak height | peak height |
Chromatography:
| Chromatography ID: | CH003628 |
| Instrument Name: | QExactive |
| Column Name: | Merck SeQuant ZIC-pHILIC (150 x 4.6mm,5um) |
| Column Temperature: | 25 |
| Flow Gradient: | 0% B to 50% B over 15 min, then to 5% B at 18 min until 21 min, increasing to 80% B at 24 min until 32 min. |
| Flow Rate: | 0.3 ml/min |
| Solvent A: | 100% water; 20 mM ammonium carbonate |
| Solvent B: | 100% acetonitrile |
| Chromatography Type: | HILIC |
MS:
| MS ID: | MS004544 |
| Analysis ID: | AN004798 |
| Instrument Name: | Thermo Q Exactive Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | The MS was fitted with a heated electrospray source that switched between positive and negative ionisation modes. The MS operated at a mass resolution of 35,000 from m/z 85 to 1,050. |
| Ion Mode: | POSITIVE |
| MS ID: | MS004545 |
| Analysis ID: | AN004799 |
| Instrument Name: | Thermo Q Exactive Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | The MS was fitted with a heated electrospray source that switched between positive and negative ionisation modes. The MS operated at a mass resolution of 35,000 from m/z 85 to 1,050. |
| Ion Mode: | NEGATIVE |
