Tauopathy-Associated Tau Fragment Ending at Amino Acid 224 Is Generated by Calpain-2 Cleavage

BACKGROUND: Tau aggregation in neurons and glial cells characterizes tauopathies as Alzheimer's disease (AD), progressive supranuclear palsy (PSP), and corticobasal degeneration (CBD). Tau proteolysis has been proposed as a trigger for tau aggregation and tau fragments have been observed in brain and cerebrospinal fluid (CSF). Our group identified a major tau cleavage at amino acid (aa) 224 in CSF; N-terminal tau fragments ending at aa 224 (N-224) were significantly increased in AD and lacked correlation to total tau (t-tau) and phosphorylated tau (p-tau) in PSP and CBD. OBJECTIVE: Previous studies have shown cleavage from calpain proteases at sites adjacent to aa 224. Our aim was to investigate if calpain-1 or -2 could be responsible for cleavage at aa 224. METHODS: Proteolytic activity of calpain-1, calpain-2, and brain protein extract was assessed on a custom tau peptide (aa 220-228), engineered with fluorescence resonance energy transfer (FRET) technology. Findings were confirmed with in-gel trypsination and mass spectrometry (MS) analysis of brain-derived bands with proteolytic activity on the FRET substrate. Finally, knock-down of the calpain-2 catalytic subunit gene (CAPN2) was performed in a neuroblastoma cell line (SH-SY5Y). RESULTS: Calpain-2 and brain protein extract, but not calpain-1, showed proteolytic activity on the FRET substrate. MS analysis of active gel bands revealed presence of calpain-2 subunits, but not calpain-1. Calpain-2 depletion and chemical inhibition suppressed proteolysis of the FRET substrate. CAPN2 knock-down caused a 76.4% reduction of N-224 tau in the cell-conditioned media. CONCLUSIONS: Further investigation of the calpain-2 pathway in the pathogenesis of tauopathies is encouraged

'I-I' and 'I-me' : Transposing Buber's interpersonal attitudes to the intrapersonal plane

Hermans' polyphonic model of the self proposes that dialogical relationships can be established between multiple I-positions1 (e.g., Hermans, 2001a). There have been few attempts, however, to explicitly characterize the forms that these intrapersonal relationships may take. Drawing on Buber's (1958) distinction between the 'I-Thou' and 'I-It' attitude, it is proposed that intrapersonal relationships can take one of two forms: an 'I-I' form, in which one I-position encounters and confirms another I-position in its uniqueness and wholeness; and an 'I-Me' form, in which one I-position experiences another I-position in a detached and objectifying way. This article argues that this I-Me form of intrapersonal relating is associated with psychological distress, and that this is so for a number of reasons: Most notably, because an individual who objectifies and subjugates certain I-position cannot reconnect with more central I-positions when dominance reversal (Hermans, 2001a) takes place. On this basis, it is suggested that a key role of the therapeutic process is to help clients become more able to experience moments of I-I intrapersonal encounter, and it is argued that this requires the therapist to confirm the client both as a whole and in terms of each of his or her different voices

Structure and function of mammalian cilia

In the past half century, beginning with electron microscopic studies of 9 + 2 motile and 9 + 0 primary cilia, novel insights have been obtained regarding the structure and function of mammalian cilia. All cilia can now be viewed as sensory cellular antennae that coordinate a large number of cellular signaling pathways, sometimes coupling the signaling to ciliary motility or alternatively to cell division and differentiation. This view has had unanticipated consequences for our understanding of developmental processes and human disease

Expression and secretion of synaptic proteins during stem cell differentiation to cortical neurons

Synaptic function and neurotransmitter release are regulated by specific proteins. Cortical neuronal differentiation of human induced pluripotent stem cells (hiPSC) provides an experimental model to obtain more information about synaptic development and physiology in vitro. In this study, expression and secretion of the synaptic proteins, neurogranin (NRGN), growth-associated protein-43 (GAP-43), synaptosomal-associated protein-25 (SNAP-25) and synaptotagmin-1 (SYT-1) were analyzed during cortical neuronal differentiation. Protein levels were measured in cells, modeling fetal cortical development, and cell-conditioned media which was used as a model of cerebrospinal fluid (CSF), respectively. Human iPSC-derived cortical neurons were maintained over a period of at least 150 days, which encompasses the different stages of neuronal development. The differentiation was divided into the following stages: hiPSC, neuro-progenitors, immature and mature cortical neurons. We show that NRGN was first expressed and secreted by neuro-progenitors while the maximum was reached in mature cortical neurons. GAP-43 was expressed and secreted first by neuro-progenitors and its expression increased markedly in immature cortical neurons. SYT-1 was expressed and secreted already by hiPSC but its expression and secretion peaked in mature neurons. SNAP-25 was first detected in neuro-progenitors and the expression and secretion increased gradually during neuronal stages reaching a maximum in mature neurons. The sensitive analytical techniques used to monitor the secretion of these synaptic proteins during cortical development make these data unique, since the secretion of these synaptic proteins has not been investigated before in such experimental models. The secretory profile of synaptic proteins, together with low release of intracellular content, implies that mature neurons actively secrete these synaptic proteins that previously have been associated with neurodegenerative disorders, including Alzheimer's disease. These data support further studies of human neuronal and synaptic development in vitro, and would potentially shed light on the mechanisms underlying altered concentrations of the proteins in bio-fluids in neurodegenerative diseases

A Cilia-inspired Closed-loop Sensor-actuator Array

© 2018, Jilin University. Cilia are finger-like cell-surface organelles that are used by certain varieties of aquatic unicellular organisms for motility, sensing and object manipulation. Initiated by internal generators and external mechanical and chemical stimuli, coordinated undulations of cilia lead to the motion of a fluid surrounding the organism. This motion transports micro-particles towards an oral cavity and provides motile force. Inspired by the emergent properties of cilia possessed by the pond organism P. caudatum, we propose a novel smart surface with closed-loop control using sensor-actuators pairings that can manipulate objects. Each vibrating motor actuator is controlled by a localised microcontroller which utilises proximity sensor information to initiate actuation. The circuit boards are designed to be plug-and-play and are infinitely up-scalable and reconfigurable. The smart surface is capable of moving objects at a speed of 7.2 millimetres per second in forward or reverse direction. Further development of this platform will include more anatomically similar biomimetic cilia and control