Identification and characterization of a new adhesin involved in the binding of Streptococcus suis to the extracellular matrix proteins

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TUNAR lncRNA Encodes a Microprotein that Regulates Neural Differentiation and Neurite Formation by Modulating Calcium Dynamics

Microproteínas; Diferenciación neural; Formación de neuritasMicroproteïnes; Diferenciació neural; Formació de neuritesMicroproteins; Neural differentiation; Neurite formationLong noncoding RNAs (lncRNAs) are regulatory molecules which have been traditionally considered as “non-coding”. Strikingly, recent evidence has demonstrated that many non-coding regions, including lncRNAs, do in fact contain small-open reading frames that code for small proteins that have been called microproteins. Only a few of them have been characterized so far, but they display key functions in a wide variety of cellular processes. Here, we show that TUNAR lncRNA encodes an evolutionarily conserved microprotein expressed in the nervous system that we have named pTUNAR. pTUNAR deficiency in mouse embryonic stem cells improves their differentiation potential towards neural lineage both in vitro and in vivo. Conversely, pTUNAR overexpression impairs neuronal differentiation by reduced neurite formation in different model systems. At the subcellular level, pTUNAR is a transmembrane protein that localizes in the endoplasmic reticulum and interacts with the calcium transporter SERCA2. pTUNAR overexpression reduces cytoplasmatic calcium, consistent with a possible role of pTUNAR as an activator of SERCA2. Altogether, our results suggest that our newly discovered microprotein has an important role in neural differentiation and neurite formation through the regulation of intracellular calcium. From a more general point of view, our results provide a proof of concept of the role of lncRNAs-encoded microproteins in neural differentiation.Work in the Abad lab is supported by VHIO, Fero Foundation, La Caixa Foundation (HR18-00256), Asociación Española Contra el Cancer (AECC), Cellex Foundation, Mutua Madrileña Foundation and by grants from the Spanish Ministry of Science and Innovation (SAF2015-69413-R; RTI2018-102046-B-I00). M.A. was recipient of a Ramon y Cajal contract from the Spanish Ministry of Science and Innovation (RYC-2013-14747). E.S. was recipient of a AECC Postdoctoral Fellowship. L.H-M. also acknowledges funding from grants SAF2017-88019-C3-1R y PID2020-116927RB-C21 from the Spanish Government. MG is supported by the advanced ERC grant NeuroCentro and the German Research Foundation (SFB870; SPP2202; SPP2306; SYNERGY; TRR274). DT is supported by the Ramón y Cajal program (RYC-2017-23486/RTI2018-095580-A-I00). MMA acknowledges funding from the Spanish Ministry of Science and Innovation PGC2018-094091-B-I00 co-funded by FEDER

Regulatory and coding sequences of TRNP1 co-evolve with brain size and cortical folding in mammals

Brain size and cortical folding have increased and decreased recurrently during mammalian evolution. Identifying genetic elements whose sequence or functional properties co-evolve with these traits can provide unique information on evolutionary and developmental mechanisms. A good candidate for such a comparative approach is TRNP1, as it controls proliferation of neural progenitors in mice and ferrets. Here, we investigate the contribution of both regulatory and coding sequences of TRNP1 to brain size and cortical folding in over 30 mammals. We find that the rate of TRNP1 protein evolution (omega) significantly correlates with brain size, slightly less with cortical folding and much less with body size. This brain correlation is stronger than for >95% of random control proteins. This co-evolution is likely affecting TRNP1 activity, as we find that TRNP1 from species with larger brains and more cortical folding induce higher proliferation rates in neural stem cells. Furthermore, we compare the activity of putative cis-regulatory elements (CREs) of TRNP1 in a massively parallel reporter assay and identify one CRE that likely co-evolves with cortical folding in Old World monkeys and apes. Our analyses indicate that coding and regulatory changes that increased TRNP1 activity were positively selected either as a cause or a consequence of increases in brain size and cortical folding. They also provide an example how phylogenetic approaches can inform biological mechanisms, especially when combined with molecular phenotypes across several species

Tbr1 Misexpression Alters Neuronal Development in the Cerebral Cortex

Changes in the transcription factor (TF) expression are critical for brain development, and they may also underlie neurodevelopmental disorders. Indeed, T-box brain1 (Tbr1) is a TF crucial for the formation of neocortical layer VI, and mutations and microdeletions in that gene are associated with malformations in the human cerebral cortex, alterations that accompany autism spectrum disorder (ASD). Interestingly, Tbr1 upregulation has also been related to the occurrence of ASD-like symptoms, although limited studies have addressed the effect of increased Tbr1 levels during neocortical development. Here, we analysed the impact of Tbr1 misexpression in mouse neural progenitor cells (NPCs) at embryonic day 14.5 (E14.5), when they mainly generate neuronal layers II-IV. By E18.5, cells accumulated in the intermediate zone and in the deep cortical layers, whereas they became less abundant in the upper cortical layers. In accordance with this, the proportion of Sox5+ cells in layers V-VI increased, while that of Cux1+ cells in layers II-IV decreased. On postnatal day 7, fewer defects in migration were evident, although a higher proportion of Sox5+ cells were seen in the upper and deep layers. The abnormal neuronal migration could be partially due to the altered multipolar-bipolar neuron morphologies induced by Tbr1 misexpression, which also reduced dendrite growth and branching, and disrupted the corpus callosum. Our results indicate that Tbr1 misexpression in cortical NPCs delays or disrupts neuronal migration, neuronal specification, dendrite development and the formation of the callosal tract. Hence, genetic changes that provoke ectopic Tbr1 upregulation during development could provoke cortical brain malformations

Trnp1 organizes diverse nuclear membrane‐less compartments in neural stem cells

TMF1‐regulated nuclear protein 1 (Trnp1) has been shown to exert potent roles in neural development affecting neural stem cell self‐renewal and brain folding, but its molecular function in the nucleus is still unknown. Here, we show that Trnp1 is a low complexity protein with the capacity to phase separate. Trnp1 interacts with factors located in several nuclear membrane‐less organelles, the nucleolus, nuclear speckles, and condensed chromatin. Importantly, Trnp1 co‐regulates the architecture and function of these nuclear compartments in vitro and in the developing brain in vivo. Deletion of a highly conserved region in the N‐terminal intrinsic disordered region abolishes the capacity of Trnp1 to regulate nucleoli and heterochromatin size, proliferation, and M‐phase length; decreases the capacity to phase separate; and abrogates most of Trnp1 protein interactions. Thus, we identified Trnp1 as a novel regulator of several nuclear membrane‐less compartments, a function important to maintain cells in a self‐renewing proliferative state

Helios expression coordinates the development of a subset of striatopallidal medium spiny neurons

Here we unravel the mechanism of action of Helios (He) during the development of striatal medium spiny neurons (MSNs). He regulates the second wave of striatal neurogenesis involved in the generation of striatopallidal neurons that express dopamine 2 receptor (D2R) and enkephalin (ENK). To exert this effect He is expressed in neural progenitor cells (NPCs) retaining them into the G1/G0 phase of the cell cycle. Thus, the lack of He produces an increase of S-phase entry and S-phase length of NPCs which in turn impairs striatal neurogenesis and produces an accumulation of the number of cycling NPCs in the germinal zone (GZ) that end up dying at postnatal stages. Therefore, He-/- mice show a reduction in the number of Dorso-Medial Striatal MSNs in the adulthood that produces deficits in motor skills acquisition. In addition, overexpression of He in NPCs induce DARPP32 phenotype when transplanted in mouse striatum.Present findings demonstrate that He is involved in the correct development of a subset of striatopallidal MSNs and reveal new cellular mechanisms for neuronal development

TUNAR lncRNA Encodes a Microprotein that Regulates Neural Differentiation and Neurite Formation by Modulating Calcium Dynamics

Long noncoding RNAs (lncRNAs) are regulatory molecules which have been traditionally considered as 'non-coding'. Strikingly, recent evidence has demonstrated that many non-coding regions, including lncRNAs, do in fact contain small-open reading frames that code for small proteins that have been called microproteins. Only a few of them have been characterized so far, but they display key functions in a wide variety of cellular processes. Here, we show that TUNAR lncRNA encodes an evolutionarily conserved microprotein expressed in the nervous system that we have named pTUNAR. pTUNAR deficiency in mouse embryonic stem cells improves their differentiation potential towards neural lineage both in vitro and in vivo. Conversely, pTUNAR overexpression impairs neuronal differentiation by reduced neurite formation in different model systems. At the subcellular level, pTUNAR is a transmembrane protein that localizes in the endoplasmic reticulum and interacts with the calcium transporter SERCA2. pTUNAR overexpression reduces cytoplasmatic calcium, consistent with a possible role of pTUNAR as an activator of SERCA2. Altogether, our results suggest that our newly discovered microprotein has an important role in neural differentiation and neurite formation through the regulation of intracellular calcium. From a more general point of view, our results provide a proof of concept of the role of lncRNAs-encoded microproteins in neural differentiation

Nolz1 promotes striatal neurogenesis through the regulation of retinoic acid signaling

Background: Nolz1 is a zinc finger transcription factor whose expression is enriched in the lateral ganglionic eminence (LGE), although its function is still unknown. Results: Here we analyze the role of Nolz1 during LGE development. We show that Nolz1 expression is high in proliferating neural progenitor cells (NPCs) of the LGE subventricular zone. In addition, low levels of Nolz1 are detected in the mantle zone, as well as in the adult striatum. Similarly, Nolz1 is highly expressed in proliferating LGE-derived NPC cultures, but its levels rapidly decrease upon cell differentiation, pointing to a role of Nolz1 in the control of NPC proliferation and/or differentiation. In agreement with this hypothesis, we find that Nolz1 over-expression promotes cell cycle exit of NPCs in neurosphere cultures and negatively regulates proliferation in telencephalic organotypic cultures. Within LGE primary cultures, Nolz1 over-expression promotes the acquisition of a neuronal phenotype, since it increases the number of β-III tubulin (Tuj1)- and microtubule-associated protein (MAP)2-positive neurons, and inhibits astrocyte generation and/or differentiation. Retinoic acid (RA) is one of the most important morphogens involved in striatal neurogenesis, and regulates Nolz1 expression in different systems. Here we show that Nolz1 also responds to this morphogen in E12.5 LGE-derived cell cultures. However, Nolz1 expression is not regulated by RA in E14.5 LGE-derived cell cultures, nor is it affected during LGE development in mouse models that present decreased RA levels. Interestingly, we find that Gsx2, which is necessary for normal RA signaling during LGE development, is also required for Nolz1 expression, which is lost in Gsx2 knockout mice. These findings suggest that Nolz1 might act downstream of Gsx2 to regulate RA-induced neurogenesis. Keeping with this hypothesis, we show that Nolz1 induces the selective expression of the RA receptor (RAR)β without altering RARα or RARγ. In addition, Nozl1 over-expression increases RA signaling since it stimulates the RA response element. This RA signaling is essential for Nolz1-induced neurogenesis, which is impaired in a RA-free environment or in the presence of a RAR inverse agonist. It has been proposed that Drosophila Gsx2 and Nolz1 homologues could cooperate with the transcriptional co-repressors Groucho-TLE to regulate cell proliferation. In agreement with this view, we show that Nolz1 could act in collaboration with TLE-4, as they are expressed at the same time in NPC cultures and during mouse development. Conclusions: Nolz1 promotes RA signaling in the LGE, contributing to the striatal neurogenesis during development

The Interaction of Canine Plasminogen with Streptococcus pyogenes Enolase: They Bind to One Another but What Is the Nature of the Structures Involved?

For years it has been clear that plasminogen from different sources and enolase from different sources interact strongly. What is less clear is the nature of the structures required for them to interact. This work examines the interaction between canine plasminogen (dPgn) and Streptococcus pyogenes enolase (Str enolase) using analytical ultracentrifugation (AUC), surface plasmon resonance (SPR), fluorescence polarization, dynamic light scattering (DLS), isothermal titration calorimetry (ITC), and simple pull-down reactions. Overall, our data indicate that a non-native structure of the octameric Str enolase (monomers or multimers) is an important determinant of its surface-mediated interaction with host plasminogen. Interestingly, a non-native structure of plasminogen is capable of interacting with native enolase. As far as we can tell, the native structures resist forming stable mixed complexes