Dataset Information

Analysis of proteins immunoprecipitated with anti-FLAG antibodies from Leishmania donovani promastigotes expressing Cdc45-FLAG or carrying empty vector

ABSTRACT: This study was carried out to identify the interacting partners of Leishmania donovani Cdc45, a protein that is conserved in eukaryotic DNA replication. Accordingly, Cdc45 was expressed in Leishmania promastigotes with a C terminal FLAG tag and whole cell lysates were isolated from these Ld1S transfectant promastigotes. Whole cell lysates were also isolated from promastigotes harbouring empty vector. These lysates were used in immunoprecipitation reactions with anti-FLAG antibodies.

INSTRUMENT(S):

ORGANISM(S): Leishmania Donovani

SUBMITTER: Swati Saha

LAB HEAD: Professor Swati Saha

PROVIDER: PXD064001 | Pride | 2025-10-27

REPOSITORIES: Pride

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Publications

Identification of Cdc45-interacting partners uncovers two Leishmania donovani proteins involved in DNA replication.

Dashora Vishal V Singh Ashish A Pandey Adarsh A Saha Swati S

BMC microbiology 20251021 1

<h4>Background</h4>Among fatalities due to parasitic diseases, deaths due to Leishmaniases rank second only to those caused by malaria. Endemic to 90 countries, more than a billion people are at risk of Leishmania infection. While there are no vaccines against Leishmaniases, treatments are available. However, the complexities of the treatment regimens, their high costs and toxic side effects, and most importantly the emerging problem of drug resistance, have continued to thrust forward research ...[more]

PMID: 41120844

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Project description:Protozoa of the genus Leishmania are the causative agents of leishmaniasis in humans. These parasites cycle between promastigotes in the sand fly mid-gut and amastigotes in phagolysosome of mammalian macrophages. During infection, host up-regulate nitric oxide synthase and parasite induce host arginase expression, both of which use arginine as a substrate. These elevated activities deplete macrophage arginine pools, a situation that invading Leishmania must overcome since it is an essential amino acid. Leishmania donovani imports exogenous arginine via a mono-specific amino acid transporter (AAP3) and utilizes it primarily through the polyamine pathway to provide precursors for trypanothione biosynthesis. Here we report the discovery of a pathway whereby promastigote and amastigote forms of the Leishmania sense the lack of environmental arginine and respond with rapid up-regulation in AAP3 expression and activity, as well as several other transporters. Significantly, this arginine deprivation response is also activated in parasites during macrophage infection. Phosphoproteomic analyses of L. donovani promastigotes have implicated a Mitogen-Activated Protein Kinase 2 (MPK2)-mediated signaling cascade in this response and L. mexicana mutants lacking MPK2 are unable to respond to arginine deprivation. In this study, we established that Leishmania cells sense the absence of arginine in their environment; both in culture (axenic promastigotes and amastigotes) and in macrophages during infection (amastigotes). This study describes the first amino acid deprivation sensing mechanism and the pathway that transduce this response, and reveals a novel host-pathogen metabolic interplay. Total RNA from Ten Leishmania donovani samples were analyzed using RNA-Seq. Cells from two life stages (promastigotes and amastigotes) were grown in axenic culture in the presence and absense of arginine. For each condition two biological replicates were grown and analyzed. In addition two macrophage grown amastigotes were analyzed.

			Action	DRS
	Experiment1_control.msf	Msf
	Experiment1_control.raw	Raw
	Experiment1_sample.msf	Msf
	Experiment1_sample.raw	Raw
	Experiment2_control.msf	Msf