The protozoan parasite Trypanosoma cruzi has a complex life cycle characterized by intracellular and extracellular forms alternating between invertebrate and mammals. To cope with these changing environments, T. cruzi undergoes rapid changes in gene expression, which are achieved essentially at the posttranscriptional level. At present, expanding families of small RNAs are recognized as key players in novel forms of posttranscriptional gene regulation in most eukaryotes. However, T. cruzi lacks canonical small RNA pathways. In a recent work, we reported the presence of alternate small RNA pathways in T. cruzi mainly represented by a homogeneous population of tRNA-derived small RNAs (tsRNAs). In T. cruzi epimastigotes submitted to nutrient starvation, tsRNAs colocalized with an argonaute protein distinctive of trypanosomatids (TcPIWI-tryp) and were recruited to particular cytoplasmic granules. Using epifluorescence and electronic microscopy, we observed that tsRNAs and the TcPIWI-tryp protein were recruited mainly to reservosomes and other intracellular vesicles including endosome-like vesicles and vesicular structures resembling the Golgi complex. These data suggested that, in T. cruzi, tsRNA biogenesis is probably part of endocytic/exocytic routes. We also demonstrated that epimastigotes submitted to nutrient starvation shed high levels of vesicles to the extracellular medium, which carry small tRNAs and TcPIWI-tryp proteins as cargo. At least a fraction of extracellular vesicle cargo was transferred between parasites and to mammalian susceptible cells. Our data afford experimental evidence, indicating that extracellular vesicles shed by T. cruzi promote not only life cycle transition of epimastigotes to trypomastigote forms but also infection susceptibility of mammalian cells.

With the advent of new and improved high-throughput sequencing technologies in the last few years, a growing number of novel classes of small RNA, other than miRNAs or siRNA, has emerged, which appear as new actors in gene expression regulation. tRNA-derived small RNAs represent one of these novel members that are, surprisingly, among the most conserved class of small RNAs throughout evolution. They could represent the most primitive small RNA pathways from which the well-known canonical RNA silencing pathways reported in higher eukaryotes evolved. This review aims to make a compilation of the most relevant research literature in this field with the purpose of shedding light on the relation of these primitive tRNA-derived molecules with the gene silencing machinery.

At present, noncoding small RNAs are recognized as key players in novel forms of posttranscriptional gene regulation in most eukaryotes. However, canonical small RNA pathways seem to be lost or excessively simplified in some unicellular organisms including Trypanosoma cruzi which lack functional RNAi pathways. Recently, we reported the presence of alternate small RNA pathways in T. cruzi mainly represented by homogeneous populations of tRNA- and rRNA-derived small RNAs, which are secreted to the extracellular medium included in extracellular vesicles. Extracellular vesicle cargo could be delivered to other parasites and to mammalian susceptible cells promoting metacyclogenesis and conferring susceptibility to infection, respectively. Here we analyzed the changes in gene expression of host HeLa cells induced by extracellular vesicles from T. cruzi. As assessed by microarray assays a large set of genes in HeLa cells were differentially expressed upon incorporation of T. cruzi-derived extracellular vesicles. The elicited response modified mainly host cell cytoskeleton, extracellular matrix, and immune responses pathways. Some genes were also modified by the most abundant tRNA-derived small RNAs included in extracellular vesicles. These data suggest that microvesicles secreted by T. cruzi could be relevant players in early events of the T. cruzi host cell interplay.

The lectin-like oxidized LDL receptor 1 (LOX-1) is a key player in the development of atherosclerosis. LOX-1 promotes endothelial activation and dysfunction by mediating uptake of oxidized LDL and inducing pro-atherogenic signaling. However, little is known about modulators of LOX-1–mediated responses. Here, we show that the function of LOX-1 is controlled proteolytically. Ectodomain shedding by the metalloprotease ADAM10 and lysosomal degradation generate membrane-bound N-terminal fragments (NTFs), which we identified as novel substrates of the intramembrane proteases signal peptide peptidase–like 2a and b (SPPL2a/b). SPPL2a/b control cellular LOX-1 NTF levels which, following self-association via their transmembrane domain, can activate MAP kinases in a ligand-independent manner. This leads to an up-regulation of several pro-atherogenic and pro-fibrotic targets including ICAM-1 and the connective tissue growth factor CTGF. Consequently, SPPL2a/b-deficient mice, which accumulate LOX-1 NTFs, develop larger and more advanced atherosclerotic plaques than controls. This identifies intramembrane proteolysis by SPPL2a/b as a novel atheroprotective mechanism via negative regulation of LOX-1 signaling.

Intramembrane proteolysis describes the cleavage of substrate proteins within their hydrophobic transmembrane segments. Several families of intramembrane proteases have been identified including the aspartyl proteases Signal peptide peptidase (SPP) and its homologues, the SPP-like (SPPL) proteases SPPL2a, SPPL2b, SPPL2c and SPPL3. As presenilin homologues, they employ a similar catalytic mechanism as the well-studied γ-secretase. However, SPP/SPPL proteases cleave transmembrane proteins with a type II topology. The characterisation of SPP/SPPL-deficient mouse models has highlighted a still growing spectrum of biological functions and also promoted the substrate discovery of these proteases. In this review, we will summarise the current hypotheses how phenotypes of these mouse models are linked to the molecular function of the enzymes. At the cellular level, SPP/SPPL-mediated cleavage events rather provide specific regulatory switches than unspecific bulk proteolysis. By this means, a plethora of different cell biological pathways is influenced including signal transduction, membrane trafficking and protein glycosylation.Publisher's Note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

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The lectin-like oxidized-LDL (oxLDL) receptor LOX-1, which is broadly expressed in vascular cells, represents a key mediator of endothelial activation and dysfunction in atherosclerotic plaque development. Being a member of the C-type lectin receptor family, LOX-1 can bind different ligands, with oxLDL being the best characterized. LOX-1 mediates oxLDL uptake into vascular cells and by this means can promote foam cell formation. In addition, LOX-1 triggers multiple signaling pathways, which ultimately induce a pro-atherogenic and pro-fibrotic transcriptional program. However, the molecular mechanisms underlying this signal transduction remain incompletely understood. In this regard, proteolysis has recently emerged as a regulatory mechanism of LOX-1 function. Different proteolytic cleavages within the LOX-1 protein can initiate its turnover and control the cellular levels of this receptor. Thereby, cleavage products with individual biological functions and/or medical significance are produced. Ectodomain shedding leads to the release of a soluble form of the receptor (sLOX1) which has been suggested to have diagnostic potential as a biomarker. Removal of the ectodomain leaves behind a membrane-bound N-terminal fragment (NTF), which despite being devoid of the ligand-binding domain is actively involved in signal transduction. Degradation of this LOX-1 NTF, which represents an athero-protective mechanism, critically depends on the aspartyl intramembrane proteases Signal peptide peptidase-like 2a and b (SPPL2a/b). Here, we present an overview of the biology of LOX-1 focusing on how proteolytic cleavages directly modulate the function of this receptor and, what kind of pathophysiological implications this has in cardiovascular disease.

Silent mutations are being intensively studied. We previously showed that the estrogen receptor alpha Ala87’s synonymous polymorphism affects its functional properties. Whereas a link has been clearly established between the effect of silent mutations, tRNA abundance and protein folding in prokaryotes, this connection remains controversial in eukaryotic systems. Although a synonymous polymorphism can affect mRNA structure or the interaction with specific ligands, it seems that the relative frequencies of isoacceptor tRNAs could play a key role in the protein-folding process, possibly through modulation of translation kinetics. Conformational changes could be subtle but enough to cause alterations in solubility, proteolysis profiles, functional parameters or intracellular targeting. Interestingly, recent advances describe dramatic changes in the tRNA population associated with proliferation, differentiation or response to chemical, physical or biological stress. In addition, several reports reveal changes in tRNAs’ posttranscriptional modifications in different physiological or pathological conditions. In consequence, since changes in the cell state imply quantitative and/or qualitative changes in the tRNA pool, they could increase the likelihood of protein conformational variants, related to a particular codon usage during translation, with consequences of diverse significance. These observations emphasize the importance of genetic code flexibility in the co-translational protein-folding process.