Biology subjects

Cattaneo, A.

Publications and source records attributed to Cattaneo, A..

6 recordsLinked to original sources

bioRxiv2018

Nuclear Tau modulates VGluT1 expression: a new function for Tau

Summary\n\nTau displacement from microtubules is the first step in the onset of tauopathies, and is followed by toxic protein aggregation. However, other non-canonical functions of Tau might have a role in these pathologies. Here, we demonstrate that a small amount of Tau localizes in the nuclear compartment and accumulates in both the soluble and DNA-bound fractions. We show that nuclear Tau regulates the expression of VGluT1, a disease-relevant gene directly involved in glutamatergic synaptic transmission. Impeding Tau/tubulin interaction in the cytosol favours its nuclear translocation and increases VGluT1 expression. Remarkably, the P301L mutation impairs this mechanism leading to a loss of function. Altogether, our results provide the demonstration of a direct physiological role of Tau on gene expression. Alterations of this mechanism may be at the basis of the onset of neurodegeneration.

molecular biology↗

bioRxiv2018

The structure of the Pro-domain of mouse proNGF in contact with the NGF domain

Nerve Growth Factor (NGF) is an important neurotrophic factor involved in the regulation of cell differentiation, maintenance, growth and survival of target neurons. Expressed as a proNGF precursor, NGF is then matured by furin-mediated protease cleavage. Increasing evidence suggests that NGF and proNGF have distinct cellular partners which account for different functional roles. While the structure of mature NGF is available, little is known about the structure of the pro-domain within the context of proNGF because the dynamical and structural features of the protein have so far prevented its structure determination. We have exploited a new hybrid strategy based on nuclear magnetic resonance and modelling validated by small angle X-ray scattering to gain novel insights on the pro-domain, both in isolation and in the context of proNGF. We show that the isolated pro-domain is intrinsically unstructured but has a clear tertiary structure propensity and forms transient tertiary intramolecular contacts. It is also able to interact, albeit weakly, with mature NGF and has per se the ability to induce growth cone collapse, indicating functional independence. Based on paramagnetic relaxation enhancement data and advanced molecular modelling, we have then reconstructed the overall properties of the pro-domain in the context of proNGF and showed that it has a compact structure. Our data represent an important step towards the structural and functional characterization of the properties of proNGF and its pro-domain.

biophysics↗

bioRxiv2018

A congenital pain insensitivity mutation in the nerve growth factor gene uncouples nociception from affective pain in heterozygous humans and mice

Pain is an unpleasant but necessary sensory experience, which facilitates adaptive behaviours, such as fear. Despite recent advances, the question of how the pain experience influences learning of the fear response is still debated1,2. Genetic disorders rendering patients congenitally unable to feel pain have been described, and are usually explained by defects in peripheral nociceptors3-5. It is not known how growing up without pain affects central emotional and motivational responses to aversive stimuli. The rare autosomal recessive Hereditary Sensory and Autonomic Neuropathy type V (HSAN V) is caused by a mutation in the nerve growth factor (NGF) gene (R100W). HSAN V homozygous patients display a congenital indifference to painful events, with deficits in peripheral nociceptors but without overt cognitive impairment6. In contrast, heterozygous carriers do not present with pain-related deficits and have been identified only through pedigree and genetic screening7,8. We exploited this clinically silent population to dissociate nociceptive from affective features of pain. To address this we investigated both heterozygous knock-in mice bearing the R100W mutation (NGFR100W/m mice) and a cohort of HSAN V heterozygous human carriers. Surprisingly, we found that NGFR100W/m mice, despite normal responses to a noxious conditioning stimulus, show a deficit in learned fear while their social and innate fear responses are normal. The lack of pain-related fear response was linked to a reduced activation of anterior cingulate and motor cortices and of striatum, but not in primary somatosensory cortex. Likewise, human heterozygous R100W carriers, despite perceiving noxious stimuli and reporting subjective pain thresholds, show increased reaction latencies in response to painful stimulation, alongside a decreased subjective urgency to react. Functional magnetic resonance imaging (fMRI) revealed, comparably to the mouse data, an altered processing of painful stimuli in rostral anterior cingulate, medial premotor cortical regions and striatum. These findings from both human and mouse HSAN V carriers uncover behavioural and motivational consequences of a mild genetic pain insensitivity on the establishment of pain-dependent affective responses and memories.

neuroscience↗

bioRxiv2017

The unfolded protein response turns from pro-survival to pro-death to cap the extent of endoplasmic reticulum expansion

Insufficient folding capacity of the endoplasmic reticulum (ER) activates the unfolded protein response (UPR) to restore homeostasis. Yet, how the UPR achieves and evaluates ER homeostatic readjustment is poorly understood. In a HeLa cell model we show that, upon a severe proteostatic insult that eclipses the ER chaperone BiP, the UPR transitions from acute full-gear activation to chronic submaximal activation, when BiP is in excess again. As such, the UPR-driven ER expansion is pro-survival, primarily via the UPR transducer ATF6. Simultaneous abrogation of ER-associated degradation (ERAD) leads to chronic full-gear UPR activation and further ER expansion, but the UPR transducers IRE1 and PERK turn pro-apoptotic. Pro-survival XBP1 mRNA processing by IRE1 is then trumped since unspliced XBP1 mRNA is depleted and IRE1s endonuclease activity is unleashed against other RNAs. Thus, the IRE1/XBP1 relay serves as a capacitor of ER expansion that heralds cell death if the expansion is deemed excessive.

cell biology↗

bioRxiv2016

Activity-dependent expression of reporter proteins at dendritic spines for synaptic activity mapping and optogenetic stimulation

Increasing evidence points to the importance of dendritic spines in the formation and allocation of memories, and alterations of spine number and physiology are associated to memory and cognitive disorders. Synaptic connections and pathways constitute the physical substrate that conveys information in the brain, and different combinations of active synaptic connections are believed to be responsible for the encoding of specific memories. In addition, modifications of the activity of such subsets of synapses are believed to be crucial for memory establishment, but a way to directly test this hypothesis, by selectively controlling the activity of potentiated spines, is currently lagging behind. Therefore it would be important to develop methods to tag active synapses for mapping functionally active connections and to selectively stimulate or interfere with active synapses. Here we introduce an approach to express light-sensitive membrane channels at synapses in an activity-dependent way by means of RNA and protein regulatory sequences. This approach is based on the local expression of reporter proteins, including optogenetic probes, at activated synapses and will allow the mapping of previously active synapses and the re-activation of the neuron only at these sites. This will allow extending the investigation of memory processes beyond the current neuron tagging technologies, whose resolution is limited at the cellular scale. Thus, it will be possible to unveil and recall the synaptic engram out of the global set of synapses.

neuroscience↗

bioRxiv2016

Assessment of Antibody Library Diversity through Next Generation Sequencing and Technical Error Compensation.

Antibody libraries are important resources to derive antibodies to be used for a wide range of applications, from structural and functional studies to intracellular protein interference studies to developing new diagnostics and therapeutics. Whatever the goal, the key parameter for an antibody library is its diversity, i.e. the number of distinct elements in the collection, which directly reflects the probability of finding in the library an antibody against a given antigen, of sufficiently high affinity. Quantitative evaluation of antibody library diversity and quality has been for a long time inadequately addressed, due to the high similarity and length of the sequences of the library. Diversity was usually inferred by the transformation efficiency and tested either by fingerprinting and/or sequencing of a few hundred random library elements. Inferring diversity from such a small sample is, however, very rudimental and gives limited information about the real complexity, because complexity does not scale linearly with sample size. Next-generation sequencing (NGS) has opened new ways to tackle the antibody library diversity quality assessment. However, much remains to be done to fully exploit the potential of NGS for the quantitative analysis of antibody repertoires and to overcome current limitations. To obtain a more reliable antibody library complexity estimate here we show a new, PCR-free, NGS approach to sequence antibody libraries on Illumina platform, coupled to a new bioinformatic analysis and software (Diversity Estimator of Antibody Library, DEAL) that allows to reliably estimate the diversity, taking in consideration the sequencing error.

bioinformatics↗