Biology subjects

Sternlieb, T.

Publications and source records attributed to Sternlieb, T..

5 recordsLinked to original sources

bioRxiv2023

A PI(3,4,5)P3-dependent allosteric switch controls antigenic variation in trypanosomes

African trypanosomes evade host immune clearance by antigenic variation, causing persistent infections in humans and animals. These parasites express a homogeneous surface coat of variant surface glycoproteins (VSGs). They transcribe one out of hundreds of VSG genes at a time from telomeric expression sites (ESs) and periodically change the VSG expressed by transcriptional switching or recombination. The mechanisms underlying the control of VSG switching and its developmental silencing remain elusive. We report that telomeric ES activation and silencing entail an on/off genetic switch controlled by a nuclear phosphoinositide signaling system. This system includes a nuclear phosphatidylinositol 5-phosphatase (PIP5Pase), its substrate PI(3,4,5)P3, and the repressor-activator protein 1 (RAP1). RAP1 binds to ES sequences flanking VSG genes via its DNA binding domains and represses VSG transcription. In contrast, PI(3,4,5)P3 binds to the N-terminus of RAP1 and controls its DNA binding activity. Transient inactivation of PIP5Pase results in the accumulation of nuclear PI(3,4,5)P3, which binds RAP1 and displaces it from ESs, activating transcription of silent ESs and VSG switching. The system is also required for the developmental silencing of VSG genes. The data provides a mechanism controlling reversible telomere silencing essential for the periodic switching in VSG expression and its developmental regulation.

microbiology↗

bioRxiv2023

Helminth-derived metabolites induce tolerogenic functional, metabolic, and transcriptional signatures in dendritic cells that attenuate experimental colitis

Helminth infections can restore immune tolerance and suppress intestinal inflammation, yet the molecular mediators underlying these effects are poorly defined. Here, we show that nonpolar metabolites derived from the helminth Heligmosomoides polygyrus bakeri (HnpM) program dendritic cells (DCs) toward a tolerogenic DC (tolDC) phenotype. Adoptive transfer of HnpM-treated DCs significantly ameliorated DSS-induced intestinal inflammation in mice. Transcriptomic and metabolomic profiling revealed that HnpM induces early metabolic reprogramming followed by gene expression signatures characteristic of tolerogenic DCs that are shared by established tolDC-inducing agents. Together, these findings identify that helminth-derived metabolites are a novel source of small molecule therapeutics to restore immune tolerance in immune-mediated disorders. Summary SentenceMetabolites produced by Heligmosomoides polygyrus induce metabolic and transcriptional changes in DCs consistent with tolDCs, and adoptive transfer of these DCs attenuate DSS-induced intestinal inflammation.

immunology↗

bioRxiv2022

Genome-wide libraries for protozoan pathogens drug target screening using yeast surface display

The lack of genetic tools to manipulate protozoan pathogens has limited the use of genome-wide approaches to identify drug or vaccine targets and understand these organisms biology. We have developed an efficient method to construct genome-wide libraries for yeast surface display (YSD) and developed a YSD fitness screen (YSD-FS) to identify drug targets. We show the robustness of our method by generating genome-wide libraries for Trypanosoma brucei, Trypanosoma cruzi, and Giardia lamblia parasites. Each library has a diversity of [~]105 to 106 clones, representing [~]6 to 30-fold of the parasites genome. Nanopore sequencing confirmed the libraries genome coverage with multiple clones for each parasite gene. Western blot and imaging analysis confirmed surface expression of the G. lamblia library proteins in yeast. Using the YSD-FS assay, we identified bonafide interactors of metronidazole, a drug used to treat protozoan and bacterial infections. We also found enrichment in nucleotide-binding domain sequences associated with yeast increased fitness to metronidazole, indicating that this drug might target multiple enzymes containing nucleotide-binding domains. The libraries are valuable biological resources for discovering drug or vaccine targets, ligand receptors, protein-protein interactions, and pathogen-host interactions. The library assembly approach can be applied to other organisms or expression systems, and the YSD-FS assay might help identify new drug targets in protozoan pathogens.

microbiology↗

bioRxiv2020

AMP-activated protein kinase: A key enzyme to manage nutritional stress responses in parasites with complex life cycles

Trypanosoma cruzi, the etiological agent of Chagas disease, has a digenetic life cycle. In its passage from the insect vector to the mammalian host, and vice versa, it must be prepared to cope with abrupt changes in environmental conditions in order to survive. Sensing and signaling pathways that allow the parasite to adapt, have unique characteristics with respect to their hosts and other free-living organisms. Many of the canonical proteins involved in these transduction pathways have not yet been found in the genomes of these parasites, because they present divergences either at the functional, structural and/or protein sequence level. All of this makes these pathways promising targets for therapeutic drugs. The AMP-activated protein kinase (AMPK) is a serine/threonine kinase activated by environmental stresses that results in reduction of ATP and increase of AMP levels. Thus, AMPK is regarded as a fuel gauge, functioning both as a nutrient and an energy sensor, to maintain energy homeostasis and, eventually, to protect cells from death by nutrient starvation. In the present study, we report the characterization of AMPK complexes for the first time in T. cruzi and describe the function of TcAMPK as a novel regulator of nutritional stress in epimastigote forms. We demonstrate that this complex possesses specific AMPK kinase activity in epimastigotes, which is inhibited by Compound C and is modulated by carbon source availability. In addition, TcAMPK2 subunit has an unprecedented functional substitution (Ser x Thr) at the activation loop and its overexpression in epimastigotes led to higher autophagic activity during prolonged nutritional stress. Moreover, the over-expression of the catalytic subunits resulted in antagonistic phenotypes associated with proliferation. Together, these results point to a role of TcAMPK in autophagy and nutrient sensing, key processes for the survival of trypanosomatids and for its life cycle progression.

cell biology↗

bioRxiv2019

Intracellular cyclic AMP levels modulate differential adaptive responses on epimastigotes and cell culture trypomastigotes of Trypanosoma cruzi

Among the many environmental challenges the parasite Trypanosoma cruzi has to overcome to complete its life cycle through different hosts, oxidative stress plays a central role. Different stages of this parasite encounter distinct sources of oxidative stress, such as the oxidative burst of the immune system, or the Heme released from hemoglobin degradation in the triatomines midgut. Also, the redox status of the surroundings functions as a signal to the parasite, triggering processes coupled to differentiation or proliferation. Intracellular second messengers, like cAMP, are responsible for the transduction of environmental queues and initiating cellular processes accordingly. In trypanosomatids cAMP is involved in a variety of processes, including proliferation, differentiation, osmoregulation and quorum sensing. Trypanosomatid phosphodiesterases (PDE) show atypical pharmacological properties and some have been involved in key processes for the survival of the parasites, which validates them as attractive therapeutic targets. Our work here shows that cAMP modulates different processes according to parasite stage. Epimastigotes become more resistant to oxidative stress when pre-treated with cAMP analogs, while trypomastigotes do not alter their response to oxidative stress under the same treatment. However, cAMP analogs do increase trypomastigotes infectivity in vitro. Also, we show that TcrPDEA1, a functionally enigmatic phosphodiesterase with very high Km, is involved in the epimastigotes response to oxidative stress.

molecular biology↗