Software for analyzing the expression of TEs at locus-specific resolution
By onWe enable the locus-specific analysis of transposable elements, with curated GTF files to improve detection of yound transposable elements, such as HERVs and young L1s. Parts of this software will be used for single-cell TE anlaysis protocols, which are under development
Immunofluorescence of RAB5 and FLAG-EEA1 puncta after Dynamin-1 and -2 inhibition with Dyngo4a
By onSelective purification of early endosomes can be achieved through affinity capture of the early endosome-associated protein EEA1 (termed Endo-IP) (Park et al. in submission). These purified endosomes can be used for proteomic and lipidomic studies to obtain snapshots of early endosomes. Here we present an immunofluorescence protocol to assess the extent of colocalization between FLAG-EEA1 and RAB5 with and without the Dynamin-1 and -2 (DNM1/2) inhibitor Dyngo4a.
Sex-Specific Microglial Responses to Glucocerebrosidase Inhibition: Relevance to GBA1-Linked Parkinson’s Disease
By onMicroglia are heterogenous cells characterized by distinct populations each contributing to specific biological processes in the nervous system, including neuroprotection. To elucidate the impact of sex-specific microglia heterogenicity to the susceptibility of neuronal stress, we video-recorded with time-lapse microscopy the changes in shape and motility occurring in primary cells derived from mice of both sexes in response to pro-inflammatory or neurotoxic stimulations. With this morpho-functional analysis, we documented distinct microglia subpopulations eliciting sex-specific responses to stimulation: male microglia tended to have a more pro-inflammatory phenotype, while female microglia showed increased sensitivity to conduritol-B-epoxide (CBE), a small molecule inhibitor of glucocerebrosidase, the enzyme encoded by the GBA1 gene, mutations of which are the major risk factor for Parkinson’s Disease (PD). Interestingly, glucocerebrosidase inhibition particularly impaired the ability of female microglia to enhance the Nrf2-dependent detoxification pathway in neurons, attenuating the sex differences observed in this neuroprotective function. This finding is consistent with the clinical impact of GBA1 mutations, in which the 1.5–2-fold reduced risk of developing idiopathic PD observed in female individuals is lost in the GBA1 carrier population, thus suggesting a sex-specific role for microglia in the etiopathogenesis of PD-GBA1
pCAG-OSF-ATG13 (2-197)-F16D
By onPlasmid for expression of human ATG13 HORMA F16D mutant in mammalian cells.
Toward a standard model for autophagosome biogenesis
By onHere, we discuss two papers focusing on autophagosome biogenesis in mammals: Olivas et al. confirming ATG9A as an autophagosome component using biochemistry, while Broadbent et al. showing autophagy protein dynamics using particle tracking.
A computational pipeline to quantify primary cilia in mouse embryonic fibroblasts with CellProfiler
By onThis protocol works with .czi format images which are acquired using a Zeiss laser scanning confocal microscope and are maximum intensity projected.
Manual Tracking/measure_ECM.ijm
By onA script to generate circular ROIs in each frame of a timelapse, using a previously generated track as reference. Used to measure fluorescence intensity (i.e., extracellular matrix content) along a microglia track. .
Manual Tracking/tracking.ijm
By onA script to store in ImageJ’s ROImanager the tracks generated using Manual Tracking plugin, for later use (drawing tracks, frame-by-frame analysis). To be used as a continuation of 1_image_prep.ijm.
Manual Tracking/image_prep.ijm
By onA script to register 2-channel timelapse images using HyperStackReg, using only 1 of the channels to compute transformation. The script also enhances contrast and prepares the image for manual tracking.
Vibrational Stabilization of Complex Network Systems
By onMany natural and man-made network systems need to maintain certain patterns, such as working at equilibria or limit cycles, to function properly. The authors provide some numerical results that demonstrate the validity of the theoretical findings.
Novel green fluorescent polyamines to analyze ATP13A2 and ATP13A3 activity in the mammalian polyamine transport system
By onBiochemical evidence presented here shows that fluorescently labeled polyamines are genuine substrates of ATP13A2.
Detecting rhythmic spiking through the power spectra of point process model residuals
By onThe work highlighted in this manuscript improves the ability to characterize oscillatory activity and detect pathological changes at the level of individual neurons.
Genetic meta-analysis of levodopa induced dyskinesia in Parkinson’s disease
By onBased on a functional annotation analysis on chromosome 1, the authors determined that changes in DNAJB4 gene expression is an additional potential cause of increased susceptibility to LiD.
Preparation of unilamellar liposomes
By onA protocol for preparing liposomes that can be used to monitor binding of proteins to vesicles of various membrane curvatures.
State-dependent GABAergic regulation of striatal spiny projection neuron excitability
By onSynaptic transmission mediated by GABAA receptors (GABAARs) in adult, principal striatal spiny projection neurons (SPNs) can suppress ongoing spiking, but its effect on synaptic integration at sub-threshold membrane potentials is less well characterized, particularly those near the resting down-state. To fill this gap, a combination of molecular, optogenetic, optical and electrophysiological approaches were used to study SPNs in mouse ex vivo brain slices, and computational tools were used to model somatodendritic synaptic integration. Activation of GABAARs, either by uncaging of GABA or by optogenetic stimulation of GABAergic synapses, evoked currents with a reversal potential near −60 mV in perforated patch recordings from both juvenile and adult SPNs. Molecular profiling of SPNs suggested that this relatively positive reversal potential was not attributable to NKCC1 expression, but rather to a dynamic equilibrium between KCC2 and Cl-/HCO3- cotransporters. Regardless, from down-state potentials, optogenetic activation of dendritic GABAergic synapses depolarized SPNs. This GABAAR-mediated depolarization summed with trailing ionotropic glutamate receptor (iGluR) stimulation, promoting dendritic spikes and increasing somatic depolarization. Simulations revealed that a diffuse dendritic GABAergic input to SPNs effectively enhanced the response to coincident glutamatergic input. Taken together, our results demonstrate that GABAARs can work in concert with iGluRs to excite adult SPNs when they are in the resting down-state, suggesting that their inhibitory role is limited to brief periods near spike threshold. This state-dependence calls for a reformulation of the role intrastriatal GABAergic circuits.
MAPL regulates gasdermin-mediated release of mtDNA from lysosomes to drive pyroptotic cell death
By onThese data place mitochondria-to-lysosome transport as a driver of pyroptosis and link multiple PD proteins along a common inflammatory pathway.
Mitochondrial degradation: Mitophagy and beyond
By onMitochondria are vital for cellular function. Degradation pathways like mitophagy regulate their quality and activity. Various mechanisms, such as mitophagy and mitochondrial extrusion, ensure cellular homeostasis by removing unwanted mitochondria.
Standard Operating Procedure (SOP) for Combined ICA and Systemic Administration of MPTP: The Overlesioned (Bilateral Asymmetric) Non-human Primate Model
By onThis standard operating procedure (SOP) describes safe methods for the use of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) in non-human primates (adult macaque monkeys).
Mitophagy induction using Difereprone
By onProtocol describing the procedure for mitophagy induction in HeLa cells using Difereprone (DFP).
