In Leishmania major, the Homolog of the Oncogene PES1 May Play a Critical Role in Parasite Infectivity

Leishmaniasis is a neglected tropical disease caused by Leishmania spp. The improvement of existing treatments and the discovery of new drugs remain ones of the major goals in control and eradication of this disease. From the parasite genome, we have identified the homologue of the human oncogene PES1 in Leishmania major (LmjPES). It has been demonstrated that PES1 is involved in several processes such as ribosome biogenesis, cell proliferation and genetic transcription. Our phylogenetic studies showed that LmjPES encodes a highly conserved protein containing three main domains: PES N-terminus (shared with proteins involved in ribosomal biogenesis), BRCT (found in proteins related to DNA repair processes) and MAEBL-type domain (C-terminus, related to erythrocyte invasion in apicomplexan). This gene showed its highest expression level in metacyclic promastigotes, the infective forms; by fluorescence microscopy assay, we demonstrated the nuclear localization of LmjPES protein. After generating mutant parasites overexpressing LmjPES, we observed that these clones displayed a dramatic increase in the ratio of cell infection within macrophages. Furthermore, BALB/c mice infected with these transgenic parasites exhibited higher footpad inflammation compared to those inoculated with non-overexpressing parasites.

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The role of Leishmania GP63 in the modulation of innate inflammatory response to Leishmania major infection

PLoS ONE ◽

10.1371/journal.pone.0262158 ◽

2021 ◽

Vol 16 (12) ◽

pp. e0262158

Author(s):

Aretha Chan ◽

Jose-Mauricio Ayala ◽

Fernando Alvarez ◽

Ciriaco Piccirillo ◽

George Dong ◽

...

Keyword(s):

Inflammatory Response ◽

Leishmania Major ◽

Critical Role ◽

Protozoan Parasite ◽

Inflammatory Cells ◽

Host Cells ◽

Cell Infection ◽

Zinc Metalloprotease

Leishmaniasis is a disease caused by the protozoan parasite Leishmania and is known to affect millions of individuals worldwide. In recent years, we have established the critical role played by Leishmania zinc-metalloprotease GP63 in the modulation of host macrophage signalling and functions, favouring its survival and progression within its host. Leishmania major lacking GP63 was reported to cause limited infection in mice, however, it is still unclear how GP63 may influence the innate inflammatory response and parasite survival in an in vivo context. Therefore, we were interested in analyzing the early innate inflammatory events upon Leishmania inoculation within mice and establish whether Leishmania GP63 influences this initial inflammatory response. Experimentally, L. major WT (L. majorWT), L. major GP63 knockout (L. majorKO), or L. major GP63 rescue (L. majorR) were intraperitoneally inoculated in mice and the inflammatory cells recruited were characterized microscopically and by flow cytometry (number and cell type), and their infection determined. Pro-inflammatory markers such as cytokines, chemokines, and extracellular vesicles (EVs, e.g. exosomes) were monitored and proteomic analysis was performed on exosome contents. Data obtained from this study suggest that Leishmania GP63 does not significantly influence the pathogen-induced inflammatory cell recruitment, but rather their activation status and effector function. Concordantly, internalization of promastigotes during early infection could be influenced by GP63 as fewer L. majorKO amastigotes were found within host cells and appear to maintain in host cells over time. Collectively this study provides a clear analysis of innate inflammatory events occurring during L. major infection and further establish the prominent role of the virulence factor GP63 to provide favourable conditions for host cell infection.

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Recent Advances in the Function of the 67 kDa Laminin Receptor and its Targeting for Personalized Therapy in Cancer

Current Pharmaceutical Design ◽

10.2174/1381612823666170710125332 ◽

2017 ◽

Vol 23 (32) ◽

pp. 4745-4757 ◽

Cited By ~ 5

Author(s):

Ada Pesapane ◽

Pia Ragno ◽

Carmine Selleri ◽

Nunzia Montuori

Keyword(s):

Cell Adhesion ◽

Cancer Progression ◽

Ribosome Biogenesis ◽

Critical Role ◽

Protein Translation ◽

Basement Membranes ◽

Laminin Receptor ◽

Cell Surface Receptor ◽

Future Application ◽

Neoplastic Cells

The 67 kDa high affinity laminin receptor (67LR) is a non-integrin cell surface receptor for laminin, the major component of basement membranes. Interactions between 67LR and laminin play a major role in mediating cell adhesion, migration, proliferation and survival. 67LR derives from homo- or hetero-dimerization of a 37 kDa cytosolic precursor (37LRP), most probably by fatty acid acylation. Interestingly, 37LRP, also called p40 or OFA/iLR (oncofetal antigen/immature laminin receptor), is a multifunctional protein with a dual activity in the cytoplasm and in the nucleus. In the cytoplasm, 37LRP it is associated with the 40S subunit of ribosome, playing a critical role in protein translation and ribosome biogenesis while in the nucleus it is tightly associated with nuclear structures, and bound to components of the cytoskeleton, such as tubulin and actin. 67LR is mainly localized in the cell membrane, concentrated in lipid rafts. Acting as a receptor for laminin is not the only function of 67LR; indeed, it also acts as a receptor for viruses, bacteria and prions. 67LR expression is increased in neoplastic cells and correlates with an enhanced invasive and metastatic potential. The primary function of 67LR in cancer is to promote tumor cell adhesion to basement membranes, the first step in the invasion-metastasis cascade. Thus, 67LR is overexpressed in neoplastic cells as compared to their normal counterparts and its overexpression is considered a molecular marker of metastatic aggressiveness in cancer of many tissues, including breast, lung, ovary, prostate, stomach, thyroid and also in leukemia and lymphoma. Thus, inhibiting 67LR binding to laminin could be a feasible approach to block cancer progression. Here, we review the current understanding of the structure and function of this molecule, highlighting its role in cancer invasion and metastasis and reviewing the various therapeutic options targeting this receptor that could have a promising future application.

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Mutational Mapping of UL130 of Human Cytomegalovirus Defines Peptide Motifs within the C-Terminal Third as Essential for Endothelial Cell Infection

Journal of Virology ◽

10.1128/jvi.00572-10 ◽

2010 ◽

Vol 84 (18) ◽

pp. 9019-9026 ◽

Cited By ~ 13

Author(s):

Andrea Schuessler ◽

Kerstin Laib Sampaio ◽

Laura Scrivano ◽

Christian Sinzger

Keyword(s):

Endothelial Cell ◽

Human Cytomegalovirus ◽

Artificial Chromosome ◽

Protein Domain ◽

Cell Infection ◽

Peptide Motifs ◽

Charged Amino Acids ◽

C Terminus ◽

Charge Cluster ◽

A Charge

ABSTRACT The UL130 gene is one of the major determinants of endothelial cell (EC) tropism of human cytomegalovirus (HCMV). In order to define functionally important peptides within this protein, we have performed a charge-cluster-to-alanine (CCTA) mutational scanning of UL130 in the genetic background of a bacterial artificial chromosome-cloned endotheliotropic HCMV strain. A total of 10 charge clusters were defined, and in each of them two or three charged amino acids were replaced with alanines. While the six N-terminal clusters were phenotypically irrelevant, mutation of the four C-terminal clusters each caused a reduction of EC tropism. The importance of this protein domain was further emphasized by the fact that the C-terminal pentapeptide PNLIV was essential for infection of ECs, and the cell tropism could not be rescued by a scrambled version of this sequence. We conclude that the C terminus of the UL130 protein serves an important function for infection of ECs by HCMV. This makes UL130 a promising molecular target for antiviral strategies, e.g., the development of antiviral peptides.

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In Vitro Antileishmanial Activity of Nicotinamide

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.49.2.808-812.2005 ◽

2005 ◽

Vol 49 (2) ◽

pp. 808-812 ◽

Cited By ~ 35

Author(s):

D. Sereno ◽

A. Monte Alegre ◽

R. Silvestre ◽

B. Vergnes ◽

A. Ouaissi

Keyword(s):

Leishmania Major ◽

Growth Arrest ◽

Antileishmanial Activity ◽

Vitamin B ◽

Wild Type ◽

Intracellular Growth ◽

Cell Growth Arrest ◽

Transgenic Parasites

ABSTRACT Our study represents the first report demonstrating the antileishmanial activity of nicotinamide (NAm), a form of vitamin B3. A 5 mM concentration of NAm significantly inhibited the intracellular growth of Leishmania amastigotes and the NAD-dependent deacetylase activity carried by parasites overexpressing Leishmania major SIR2 (LmSIR2). However, the transgenic parasites were as susceptible as the wild-type parasites to NAm-induced cell growth arrest. Therefore, we conclude that NAm inhibits leishmanial growth and that overexpression of LmSIR2 does not overcome this inhibition. The mechanism of the inhibition is not defined but may include other in vivo targets. NAm may thus represent a new antileishmanial agent which could potentially be used in combination with other drugs during therapy.

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Prediction of GluN2B-CT1290-1310/DAPK1 Interaction by Protein–Peptide Docking and Molecular Dynamics Simulation

Molecules ◽

10.3390/molecules23113018 ◽

2018 ◽

Vol 23 (11) ◽

pp. 3018 ◽

Cited By ~ 6

Author(s):

Gao Tu ◽

Tingting Fu ◽

Fengyuan Yang ◽

Lixia Yao ◽

Weiwei Xue ◽

...

Keyword(s):

Molecular Dynamics ◽

Free Energy ◽

Electrostatic Interactions ◽

Binding Free Energy ◽

Computational Simulation ◽

Critical Role ◽

Dynamics Simulation ◽

Peptide Docking ◽

C Terminus ◽

Free Energy Decomposition

The interaction of death-associated protein kinase 1 (DAPK1) with the 2B subunit (GluN2B) C-terminus of N-methyl-D-aspartate receptor (NMDAR) plays a critical role in the pathophysiology of depression and is considered a potential target for the structure-based discovery of new antidepressants. However, the 3D structures of C-terminus residues 1290–1310 of GluN2B (GluN2B-CT1290-1310) remain elusive and the interaction between GluN2B-CT1290-1310 and DAPK1 is unknown. In this study, the mechanism of interaction between DAPK1 and GluN2B-CT1290-1310 was predicted by computational simulation methods including protein–peptide docking and molecular dynamics (MD) simulation. Based on the equilibrated MD trajectory, the total binding free energy between GluN2B-CT1290-1310 and DAPK1 was computed by the mechanics generalized born surface area (MM/GBSA) approach. The simulation results showed that hydrophobic, van der Waals, and electrostatic interactions are responsible for the binding of GluN2B-CT1290–1310/DAPK1. Moreover, through per-residue free energy decomposition and in silico alanine scanning analysis, hotspot residues between GluN2B-CT1290-1310 and DAPK1 interface were identified. In conclusion, this work predicted the binding mode and quantitatively characterized the protein–peptide interface, which will aid in the discovery of novel drugs targeting the GluN2B-CT1290-1310 and DAPK1 interface.

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The Nuclear Export Receptors TbMex67 and TbMtr2 Are Required for Ribosome Biogenesis in Trypanosoma brucei

mSphere ◽

10.1128/msphere.00343-19 ◽

2019 ◽

Vol 4 (4) ◽

Cited By ~ 3

Author(s):

Constance Rink ◽

Martin Ciganda ◽

Noreen Williams

Keyword(s):

Trypanosoma Brucei ◽

Nuclear Export ◽

Ribosome Biogenesis ◽

Biological Process ◽

Large Subunit ◽

Critical Role ◽

Ribosomal Subunits ◽

Content Type ◽

Protein Components ◽

Export Receptors

ABSTRACT Ribosomal maturation is a complex and highly conserved biological process involving migration of a continuously changing RNP across multiple cellular compartments. A critical point in this process is the translocation of individual ribosomal subunits (60S and 40S) from the nucleus to the cytoplasm, and a number of export factors participate in this process. In this study, we characterize the functional role of the auxiliary export receptors TbMex67 and TbMtr2 in ribosome biogenesis in the parasite Trypanosoma brucei. We demonstrate that depletion of each of these proteins dramatically impacts the steady-state levels of other proteins involved in ribosome biogenesis, including the trypanosome-specific factors P34 and P37. In addition, we observe that the loss of TbMex67 or TbMtr2 leads to aberrant ribosome formation, rRNA processing, and polysome formation. Although the TbMex67-TbMtr2 heterodimer is structurally distinct from Mex67-Mtr2 complexes previously studied, our data show that they retain a conserved function in ribosome biogenesis. IMPORTANCE The nuclear export of ribosomal subunits (60S and 40S) depends in part on the activity of the essential auxiliary export receptors TbMtr2 and TbMex67. When these proteins are individually depleted from the medically and agriculturally significant parasite Trypanosoma brucei, distinct alterations in the processing of the rRNAs of the large subunit (60S) are observed as well as aberrations in the assembly of functional ribosomes (polysomes). We also established that TbMex67 and TbMtr2 interact directly or indirectly with the protein components of the 5S RNP, including the trypanosome-specific P34/P37. The critical role that TbMex67 and TbMtr2 play in this essential biological process together with their parasite-specific interactions may provide new therapeutic targets against this important parasite.

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Molecular basis of the dual role of the Mlh1-Mlh3 endonuclease in MMR and in meiotic crossover formation

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2022704118 ◽

2021 ◽

Vol 118 (23) ◽

pp. e2022704118

Author(s):

Jingqi Dai ◽

Aurore Sanchez ◽

Céline Adam ◽

Lepakshi Ranjha ◽

Giordano Reginato ◽

...

Keyword(s):

Meiotic Recombination ◽

Molecular Basis ◽

Crystal Packing ◽

Critical Role ◽

Holliday Junctions ◽

Dual Roles ◽

Terminal Domain ◽

C Terminus ◽

Meiotic Crossover

In budding yeast, the MutL homolog heterodimer Mlh1-Mlh3 (MutLγ) plays a central role in the formation of meiotic crossovers. It is also involved in the repair of a subset of mismatches besides the main mismatch repair (MMR) endonuclease Mlh1-Pms1 (MutLα). The heterodimer interface and endonuclease sites of MutLγ and MutLα are located in their C-terminal domain (CTD). The molecular basis of MutLγ’s dual roles in MMR and meiosis is not known. To better understand the specificity of MutLγ, we characterized the crystal structure of Saccharomyces cerevisiae MutLγ(CTD). Although MutLγ(CTD) presents overall similarities with MutLα(CTD), it harbors some rearrangement of the surface surrounding the active site, which indicates altered substrate preference. The last amino acids of Mlh1 participate in the Mlh3 endonuclease site as previously reported for Pms1. We characterized mlh1 alleles and showed a critical role of this Mlh1 extreme C terminus both in MMR and in meiotic recombination. We showed that the MutLγ(CTD) preferentially binds Holliday junctions, contrary to MutLα(CTD). We characterized Mlh3 positions on the N-terminal domain (NTD) and CTD that could contribute to the positioning of the NTD close to the CTD in the context of the full-length MutLγ. Finally, crystal packing revealed an assembly of MutLγ(CTD) molecules in filament structures. Mutation at the corresponding interfaces reduced crossover formation, suggesting that these superstructures may contribute to the oligomer formation proposed for MutLγ. This study defines clear divergent features between the MutL homologs and identifies, at the molecular level, their specialization toward MMR or meiotic recombination functions.

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The C-terminus of ubiquitin plays a critical role in deamidase Lpg2148 recognition

Biochemical and Biophysical Research Communications ◽

10.1016/j.bbrc.2018.08.074 ◽

2018 ◽

Vol 503 (4) ◽

pp. 2943-2948

Author(s):

Min Zhu ◽

Xiuchang Ma ◽

Yongxiang Gao ◽

Xiaowu Li ◽

Jiyuan Ke ◽

...

Keyword(s):

Critical Role ◽

C Terminus

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Potential Role of Darvyadi Kwatha in the Management of Diabetes

The Healer ◽

10.51649/healer.26 ◽

2021 ◽

Vol 2 (1) ◽

pp. 80-86

Author(s):

Shankar Gautam ◽

Abhishek Upadhyay ◽

Rashmi Mutha ◽

BINOD KUMAR SINGH ◽

Ram Kishor Joshi

Keyword(s):

Causes Of Death ◽

Potential Role ◽

Critical Role ◽

Animal Experiments ◽

Safety Evaluation ◽

New Drugs ◽

Effective Management ◽

The Individual ◽

Glucose 6 Phosphate Dehydrogenase

Diabetes is a clinical condition characterized by a spike in blood glucose in plasma. It is one of the 21st century's greatest public health crises and is among the top 10 causes of death worldwide. Although new drugs and therapeutics are emerging for its management but the prevalence is increasing at an alarming pace; thus, every system must contribute for effective management. An effort is made to review the efficacy and safety evaluation of the individual herbs of Darvyadi Kwatha (DK), an Ayurvedic formulation mentioned in Charaka Samhita. The constituents of the DK has some strong efficient antidiabetic/hypoglycaemic chemical principle having insulin-triggering and insulin-like behaviors which increases the activity of glucose-6-phosphate dehydrogenase (G6PD) and glucokinase and decreases glucose-6-phosphatase activity, reduce oxidative stress and prevention of glutathione reductase, superoxide dismutase, and catalase activity play a critical role in glucose homeostasis. DK also improve biochemical parameters such as SGPT, SGOT, cholesterol and triglycerides and is found to be safe in animal experiments. The various evidences clearly indicates that DK has definite hypoglycemic potential as well as anti-diabetic activity.

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Sterol 14α-Demethylase from Trypanosomatidae Parasites as a Promising Target for Designing New Antiparasitic Agents

Current Topics in Medicinal Chemistry ◽

10.2174/1568026621666210303144448 ◽

2021 ◽

Vol 21 ◽

Author(s):

Paulo Fernando da Silva Santos-Júnior ◽

Martine Schmitt ◽

João Xavier de Araújo-Júnior ◽

Edeildo Ferreira da Silva-Júnior

Keyword(s):

Combined Therapy ◽

Experimental Studies ◽

New Drugs ◽

Host Cells ◽

Sar Studies ◽

Promising Target ◽

Ic50 Value ◽

Leishmania Spp ◽

Etiological Agents ◽

Antiparasitic Agents

: Trypanosomatidae family belongs to the Kinetoplastida order, which consists of obligatory parasites that affect plants and all classes of vertebrates, especially humans and insects. Among the heteroxenic parasites, Leishmania spp., Trypanosoma cruzi, and T. brucei are protozoa of most significant interest for medicinal chemistry, being etiological agents of Leishmaniasis, Chagas, and Sleep Sickness diseases, respectively. Currently, inefficient pharmacotherapy, especially in chronic phases and low selectivity towards parasite/host cells, justifies the need to discover new drugs to treat them effectively. Among other targets, the sterol 14α-demethylase (CYP51), an enzyme responsible for ergosterol's biosynthesis in Trypanosomatidae parasites, has received more attention in the development of new bioactive compounds. In this context, antifungal ravuconazole proved to be the most promising drug among this class against T. cruzi, being used in combined therapy with Bnz in clinic trials. Non-antifungal inhibitors, such as VFV and VNF, have shown promising results against T. cruzi and T.brucei, respectively, being tested in Bnz-combined therapies. Among the experimental studies involving azoles, compound (15) was found to be the most promising derivative, displaying an IC50 value of 0.002 µM against amastigotes from T. cruzi, in addition to being non-toxic and highly selective towards TcCYP51 (< 25 nM). Interestingly, imidazole analog (16) was active against infectious forms of these three parasites, demonstrating Ki values of 0.17, 0.02, and 0.36 nM for CYP51 from T. cruzi, T. brucei, and L. infantum. Finally, this review will address promising inhibitors targeting sterol 14α-demethylase (CYP51) from Trypanosomatidae parasites, highlighting SAR studies, interactions with this target, and recent contributions and advances in the field, as well.

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