Biology subjects

Layer, N.

Publications and source records attributed to Layer, N..

5 recordsLinked to original sources

bioRxiv2026

Local translation drives glioblastoma heterogeneity and tumor invasion

O_FIG O_LINKSMALLFIG WIDTH=174 HEIGHT=200 SRC="FIGDIR/small/722387v1_ufig1.gif" ALT="Figure 1"> View larger version (96K): org.highwire.dtl.DTLVardef@1da020corg.highwire.dtl.DTLVardef@1bc9b32org.highwire.dtl.DTLVardef@171198aorg.highwire.dtl.DTLVardef@43ce6f_HPS_FORMAT_FIGEXP M_FIG C_FIG Glioblastoma is characterized by diffuse brain invasion, yet the subcellular mechanisms enabling this aggressive behavior remain poorly understood. A subpopulation of glioblastoma cells forms invasive tumor microtubes (TMs), neurite-like extensions that drive whole-brain colonization. Here, we establish local protein translation as a fundamental driver of TM dynamics and invasive cell states. Developing a subcellular transcriptomics approach - integrating subcellular organelle organization with spatially resolved transcriptomics and functional readouts - we reveal that TM gene expression drives cell state identity. Invasive cells further exhibit significantly elevated local translation in protruding TMs, directly linking subcellular protein synthesis to functional invasive states associated with neurodevelopmental programs of axonal growth cones. Targeted disruption of TM-localized translation via photoswitchable puromycin, and specific knockdowns of the TM-enriched proteins GPM6A and GAP43, impaired TM dynamics, suppressed invasion, and reduced tumor growth. Together, these findings define local translation as a key determinant of tumor heterogeneity and glioblastoma invasion.

cancer biology↗

bioRxiv2024

Characterizing and targeting glioblastoma neuron-tumor networks with retrograde tracing

Glioblastomas are invasive brain tumors with high therapeutic resistance. Neuron-to-glioma synapses have been shown to promote glioblastoma progression. However, a characterization of tumor-connected neurons has been hampered by a lack of technologies. Here, we adapted retrograde tracing using rabies viruses to investigate and manipulate neuron-tumor networks. Glioblastoma rapidly integrated into neural circuits across the brain engaging in widespread functional communication, with acetylcholinergic neurons driving glioblastoma invasion. We uncovered patient-specific and tumor cell state-dependent differences in synaptogenic gene expression associated with neuron-tumor connectivity and subsequent invasivity. Importantly, radiotherapy enhanced neuron-tumor connectivity by increased neuronal activity. In turn, simultaneous neuronal activity inhibition and radiotherapy showed increased therapeutic effects, indicative of a role for neuron-to-glioma synapses in contributing to therapeutic resistance. Lastly, rabies-mediated genetic ablation of tumor-connected neurons halted glioblastoma progression, offering a viral strategy to tackle glioblastoma. Together, this study provides a framework to comprehensively characterize neuron-tumor networks and target glioblastoma.

cancer biology↗

bioRxiv2023

Axonopathy and altered synaptic development in early hippocampal epileptogenesis of Dravet syndrome

Dravet syndrome caused by SCN1A variants is a severe developmental epileptic encephalopathy (DEE) characterized by pharmaco-resistant epileptic seizures and progressive neurodevelopmental decline with cognitive impairment and autism-spectrum-traits. Numerous preceding studies indicate that the initial pathophysiology due to impaired NaV1.1 function mainly derives from reduced interneuron firing leading to a network hyperexcitability (Bender et al. 2012). However, little is known how epileptogenesis and generally disease pathogenesis progress from the inborn molecular defect to infantile seizure onset. We address this question in a Dravet mouse model by comprehensive single-cell RNA sequencing and selected downstream analysis via single-cell electrophysiology, histology, live cell imaging and electron microscopy. Our data reveal a continuum of early primary (preseizure) and secondary (post-seizure onset) transcriptomic changes in various cell populations in the hippocampus. Focusing on cornu ammonis, we find a number of transcriptional pathways that are dysregulated including synaptic transmembrane adhesion molecules of the neurexin superfamily and voltage-gated ion channels. Further investigations support an ultrastructural and functional axonopathy and synaptopathy of parvalbumin interneurons. These processes precede somatic firing impairment and seizures suggesting they underlie fundamental early-phase disease mechanisms. Taken together we provide a cellularly resolved transcriptomic resource of early disease phases of Dravet syndrome and demonstrate epileptogenesis beyond NaV1.1 loss-of-function during an early developmental time window of CNS maturation. Altogether these data establish proof-of principle that the concept of epileptogenesis, originally devised for acquired forms of epilepsy, similarly applies to genetic epilepsies and DEEs.

neuroscience↗

bioRxiv2021

Dravet variant SCN1AA1783V impairs interneuron firing predominantly by altered channel activation

Dravet syndrome (DS) is a developmental epileptic encephalopathy mainly caused by functional NaV1.1 haploinsufficiency in interneurons (IN). Recently, a new conditional mouse model expressing the recurrent human p.A1783V missense variant has become available. Here we provide an electrophysiological characterization of this variant in tsA201 cells, revealing both altered voltage-dependence of activation and slow inactivation without reduced sodium peak current density. Simulating IN excitability in a Hodgkin-Huxley one-compartment model suggested surprisingly similar firing deficits for Scn1aA1783V and full haploinsufficiency as caused by heterozygous truncation variants. Impaired NaVA1783V channel activation was predicted to have a significantly larger impact on channel function than altered slow inactivation and is therefore proposed as the main mechanism underlying IN dysfunction. The computational model was validated in cortical organotypic slice cultures derived from conditional Scn1aA1783V mice. Pan-neuronal activation of the p.A1783V variant in vitro confirmed the predicted IN firing deficit while demonstrating normal excitability of pyramidal neurons. Taken together these data demonstrate that despite maintained physiological peak currents density LOF gating properties may match effects of full haploinsufficiency on neuronal level, thereby causing DS. HighlightsNaV1.1A1783V alters voltage-dependence of activation and slow inactivation while not affecting fast inactivation. Depolarizing and hyperpolarizing shifts of activation and slow inactivation curves result in combined channel loss of function (LOF). Simulations of NaV1.1A1783V interneuronal properties indicate reduced action potential firing rates comparable to full SCN1A haploinsufficiency, which is often found in Dravet syndrome. In silico modelling identifies impaired channel activation as the predominant mechanism of channel LOF. Panneuronal induction of Scn1a+/A1783V in a cortical slice culture model confirms restriction of loss of function and its restriction to interneurons.

neuroscience↗

bioRxiv2019

Human cerebrospinal fluid induce neuronal excitability changes in resected human neocortical and hippocampal brain slices

Human cerebrospinal fluid (hCSF) have proven advantageous over conventional medium when culturing both rodent and human brain tissue. Increased excitability and synchronicity, similar to the active state exclusively recorded in vivo, reported in rodent slice and cell-cultures with hCSF as recording medium, indicates properties of the hCSF not matched by the artificial cerebrospinal fluid (aCSF) commonly used for electrophysiological recording. To evaluate the possible importance of using hCSF as electrophysiological recording medium of human brain tissue, we compared the general excitability in ex vivo human brain tissue slice cultures during perfusion with hCSF and aCSF. For measuring the general activity from a majority of neurons within neocortical and hippocampal human ex vivo slices we used a microelectrode array (MEA) recording technique with 252 electrodes covering an area of 3.2 x 3.2 mm2 and a second CMOS-based MEA with 4225 electrodes on a 2 x 2 mm2 area for detailed mapping of action potential waveforms. We found that hCSF increase the number of active neurons and the firing rate of the neurons in the slices as well as increasing the numbers of bursts while leaving the duration of the bursts unchanged. Interestingly, not only an increase in the overall activity in the slices was observed, but a reconfiguration of the network functionality could be detected with specific activation and inactivation of subpopulations of neuronal ensembles. In conclusion, hCSF is an important component to consider for future human tissue studies, especially for experiments designed to mimic the in vivo situation.

neuroscience↗