Clathrin-mediated hemoglobin endocytosis is essential for survival of Leishmania

Agarwal, Shruti; Rastogi, Ruchir; Gupta, Deepak Kumar; Patel, N.; Raje, Manoj; Mukhopadhyay, Asok

doi:10.1016/j.bbamcr.2013.01.006

Cited by 36 publications

(37 citation statements)

References 47 publications

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“…Most importantly, our results also showed that hexokinase and clathrin heavy chain (LdCHC) are associated with the LdRab5b interactome. These results are consistent with our previous finding that the HbR is a hexokinase (25), and Hb endocytosis a is clathrin-dependent process (23) in Leishmania. Subsequently, we validated these observations using pulldown assay.…”

Section: Mechanism Of Regulation Of Various Modes Of Endocytosis In Lsupporting

confidence: 83%

“…LdCHC of Leishmania specifically contains an insertion of 10 amino acid residues (MAQKQDTDLN) in the distal domain (23) and also interacts with LdRab5b, suggesting that LdRab5b-regulated clathrin-mediated endocytosis might be controlled by different effector(s). Taken together, these results suggest that LdRab5b plays a major role in regulating receptor-mediated endocytosis.…”

Section: Discussionmentioning

confidence: 99%

“…In this study, we have used the unicellular parasite, Leishmania, because this parasite offers a very good model to study endocytosis and intracellular trafficking (23,26,27). Moreover, depletion, overexpression, or generation of the null-mutant of target gene/protein shows the phenotype and affects the viability of the cells and thereby indicates the essential functions of the respective Rabs in an organism.…”

Section: Discussionmentioning

confidence: 99%

“…Moreover, depletion, overexpression, or generation of the null-mutant of target gene/protein shows the phenotype and affects the viability of the cells and thereby indicates the essential functions of the respective Rabs in an organism. Previously, we have shown that Hb endocytosis is a clathrin-mediated process in Leishmania (23)(24)(25) and is essential for parasite survival (40). Therefore, we have used Hb as a receptor-mediated endocytic probe and ϩ/Ϫ , and Leishmania:Rab5b ϩ/Ϫ .…”

Section: Discussionmentioning

confidence: 99%

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Rab5 Isoforms Specifically Regulate Different Modes of Endocytosis in Leishmania

Rastogi

Verma

Kapoor

et al. 2016

Journal of Biological Chemistry

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Differential functions of Rab5 isoforms in endocytosis are not well characterized. Here, we cloned, expressed, and characterized Rab5a and Rab5b from Leishmania and found that both of them are localized in the early endosome. To understand the role of LdRab5 isoforms in different modes of endocytosis in Leishmania, we generated transgenic parasites overexpressing LdRab5a, LdRab5b, or their dominant-positive (LdRab5a:Q93L and LdRab5b:Q80L) or dominant-negative mutants (LdRab5a: N146I and LdRab5b:N133I). Using LdRab5a or its mutants overexpressing parasites, we found that LdRab5a specifically regulates the fluid-phase endocytosis of horseradish peroxidase and also specifically induced the transport of dextran-Texas Red to the lysosomes. In contrast, cells overexpressing LdRab5b or its mutants showed that LdRab5b explicitly controls receptormediated endocytosis of hemoglobin, and overexpression of LdRab5b:WT enhanced the transport of internalized Hb to the lysosomes in comparison with control cells. To unequivocally demonstrate the role of Rab5 isoforms in endocytosis in Leishmania, we tried to generate null-mutants of LdRab5a and LdRab5b parasites, but both were lethal indicating their essential functions in parasites. Therefore, we used heterozygous LdRab5a ؉/؊ and LdRab5b ؉/؊ cells. LdRab5a ؉/؊ Leishmania showed 50% inhibition of HRP uptake, but hemoglobin endocytosis was uninterrupted. In contrast, about 50% inhibition of Hb endocytosis was observed in LdRab5b ؉/؊ cells without any significant effect on HRP uptake. Finally, we tried to identify putative LdRab5a and LdRab5b effectors. We found that LdRab5b interacts with clathrin heavy chain and hemoglobin receptor. However, LdRab5a failed to interact with the clathrin heavy chain, and interaction with hemoglobin receptor was significantly less. Thus, our results showed that LdRab5a and LdRab5b differentially regulate fluid phase and receptor-mediated endocytosis in Leishmania.

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Section: Mechanism Of Regulation Of Various Modes Of Endocytosis In Lsupporting

confidence: 83%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

See 2 more Smart Citations

Rab5 Isoforms Specifically Regulate Different Modes of Endocytosis in Leishmania

Rastogi

Verma

Kapoor

et al. 2016

Journal of Biological Chemistry

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“…Recently, we have shown that Ldgp63 is trafficked via Rab1 dependent conventional secretory pathway in Leishmania (36). Moreover, we have also shown that Leishmania has conserved trafficking pathways like higher eukaryotic cells (37)(38)(39)(40)(41)(42)(43)(44). Thus, it is tempting to speculate that Ldgp63 might exit from ER by Sar1-dependent COPII mediated process.…”

Section: Discussionmentioning

confidence: 99%

GTPase Sar1 regulates the trafficking and secretion of the virulence factor gp63 in Leishmania

Parashar

Mukhopadhyay

2017

Journal of Biological Chemistry

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Identification and characterization of the hemoglobin‐binding domain of hemoglobin receptor in Leishmania

Rastogi

Verma²,

Singh

et al. 2021

FEBS Letters

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Leishmania internalize hemoglobin (Hb) via a specific receptor (HbR) for their survival. To identify the Hb-binding domain of HbR, we cloned and expressed several truncated proteins of HbR and determined their ability to bind Hb. Our findings reveal that 90% of Hb-binding activity is retained in HbR 41-80 in comparison with HbR 1-471. We synthesized a 40 amino acid peptide (SSEKMKQLTMYMIHEMVEGLEGRPSTVRMLPSFVYTSDPA) corresponding to HbR 41-80 and found that it specifically binds Hb. Subsequently, we found that the HbR 41-80 peptide completely blocks Hb uptake in both promastigote and amastigote forms of Leishmania and, thereby, inhibits the growth of the parasite. These results demonstrate that HbR 41-80 is the Hb-binding domain of HbR, which might be used as a potential therapeutic agent to inhibit the growth of Leishmania.

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Clathrin-mediated hemoglobin endocytosis is essential for survival of Leishmania

Cited by 36 publications

References 47 publications

Rab5 Isoforms Specifically Regulate Different Modes of Endocytosis in Leishmania

Rab5 Isoforms Specifically Regulate Different Modes of Endocytosis in Leishmania

GTPase Sar1 regulates the trafficking and secretion of the virulence factor gp63 in Leishmania

Identification and characterization of the hemoglobin‐binding domain of hemoglobin receptor in Leishmania

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