Biology subjects

Borczyk, M.

Publications and source records attributed to Borczyk, M..

8 recordsLinked to original sources

bioRxiv2025

Risperidone regulates the expression of schizophrenia-related genes in the murine forebrain

Risperidone acts on monoaminergic signaling to alleviate psychosis. At the cellular level, through both direct and indirect effects, the drug induces a specific pattern of gene expression, which differs between the basal ganglia and frontal cortex. These risperidone-regulated changes influence neuronal plasticity and are crucial for both its antipsychotic and extrapyramidal effects. Here, we employed sequencing-based spatial transcriptomics to comprehensively characterize gene expression changes in the male mouse (Mus musculus L.) forebrain after an acute dose of risperidone (0.5 mg/kg, i.p.). The transcriptional patterns were structure-specific, and unsupervised clustering of spatial profiles accurately identified cortical divisions, layers, and basal ganglia subregions. Differential gene expression was subsequently analyzed within each anatomically defined cluster using a customized statistical framework. Risperidone significantly altered the levels of 95 transcripts across 12 brain regions. The largest number of changes was observed in ventral brain areas, including the olfactory tubercle (25 differentially regulated transcripts), the diagonal band nucleus (22), the corpus callosum and commissures (13), and the lateral septal nucleus (9). Notably, 21 of the 95 differentially expressed genes were previously associated with schizophrenia, including Olig2, Smpd3, and Cacna1i. Overall, our results indicate that the strongest effects of risperidone are in medial and ventral brain regions rich in oligodendrocytes and glial cells. Furthermore, the enrichment analysis provides robust evidence of a molecular link between the drugs mechanism of action and genetic factors involved in schizophrenia. HighlightsO_LIUnsupervised clustering accurately identifies transcripts localization in the brain C_LIO_LIAcute risperidone treatment alters spatial transcriptional patterns of 95 genes C_LIO_LIRisperidone-regulated transcripts include 21 genes previously linked to schizophrenia C_LIO_LISpatial gene expression analysis offers novel insight into the drug action mechanism C_LI

pharmacology and toxicology↗

bioRxiv2024

μ-Opioid receptor transcriptional variants in the murine forebrain and spinal cord

BackgroundOprm1, the gene encoding the -opioid receptor, has multiple reported transcripts, with a variable 3 region and many alternative sequences encoding the C-terminus of the protein. The functional implications of this variability remain mostly unexplored, though a recurring notion is that it could be exploited by developing selective ligands with improved clinical profiles. Here, we comprehensively examined Oprm1 transcriptional variants in the murine central nervous system. MethodsRNA-seq transcription analyses were performed based on Oxford Nanopore Sequencing (ONS) and 10x Genomics Visium spatial transcriptomics data. The spatial distribution of Oprm1 exons was evaluated via RNAscope in situ hybridization. Tissue and cell-type specificity was assessed based on reanalysis single-cell RNAseq databases. ResultsWe detected a mismatch between transcripts annotated in GRCm38/mm10 and RNA-seq results. Sequencing data indicated that the primary Oprm1 transcript has a 3 terminus located on chr10:6,860,027, which is ~9.5 kilobases downstream of the longest annotated exon 4 end. Long-read sequencing confirmed that the final Oprm1 exon included a 10.2 kilobase long 3 untranslated region. The presence of the long variant was unambiguously confirmed using RNAscope in situ hybridization. The long variant was observed in the thalamus, striatum, cortex and spinal cord. Expression of additional variants of the Oprm1 gene was close to the detection limit. Reanalysis of single-cell sequencing data confirmed these observations and indicated that Oprm1 was expressed mainly in parvalbumin-, somatostatin- and VIP-positive cells. ConclusionThe primary transcript of the Oprm1 mouse gene is a variant with a long 3 untranslated region. Author SummaryOpioids are essential for the management of pain and have multiple other medical indications; however, their addictive properties and widespread misuse have led to a severe modern health crisis. Accordingly, there has been a major effort to develop novel compounds that retain clinical effectiveness while diminishing their addictive potential and other adverse effects. One of the potential avenues for safer opioid drugs is developing compounds that are selective for a specific group of the main targets of opioid medications--the -opioid receptors. Multiple variants the -opioid receptor have been reported, encoded by different transcripts of the Oprm1 gene. Here, we used RNA transcript sequencing and in situ hybridization with probes to detect different parts of Oprm1 transcripts to validate the existence of various reported isoforms. Our main finding is that the primary transcript of the receptor is much longer than the current reference sequences annotated in the mouse genome and has an over 10,000-base-long noncoding sequence at the 3 terminus. Several other types of transcripts are also expressed; however, they represent approximately 15% or less of the total transcript content in each of the examined brain regions. In the context of future research on opioid drugs, these results indicate that it is unlikely that different subpopulations of receptors could be targeted.

genomics↗

bioRxiv2023

Phosphorylation of PSD-95 at Serine 73 in dCA1 is required for extinction of contextual fear

The updating of contextual memories is essential for survival in a changing environment. Accumulating data indicate that the dorsal CA1 area (dCA1) contributes to this process. However, the cellular and molecular mechanisms of contextual fear memory updating remain poorly understood. Postsynaptic density protein 95 (PSD-95) regulates the structure and function of glutamatergic synapses. Here, using dCA1-targeted genetic manipulations in vivo, combined with ex vivo 3D electron microscopy and electrophysiology, we identify a novel, synaptic mechanism that is induced during attenuation of contextual fear memories and involves phosphorylation of PSD-95 at Serine 73 in dCA1. Our data provide the proof that PSD-95-dependent synaptic plasticity in dCA1 is required for updating of contextual fear memory.

neuroscience↗

bioRxiv2022

Full-length dystrophin is expressed across human tissues and DMD downregulation commonly occurring in tumours coincides with Duchenne-like molecular alterations

Mutations of the DMD gene, encoding dystrophins, cause Duchenne muscular dystrophy (DMD). Some tumors also display altered dystrophin expression and recent studies identified a developmental onset of DMD. Given that embryogenesis and carcinogenesis share many mechanisms, we analyzed a broad spectrum of tumors to establish whether dystrophin loss evokes related outcomes. Transcriptomic, proteomic, and mutation datasets from fifty tumor tissues and matching controls (10,894 samples) and 140 corresponding tumor cell lines were analyzed. Interestingly, DMD expression was widespread across healthy tissues at levels comparable to housekeeping genes. In 80% of tumors, DMD expression was reduced due to transcriptional downregulation and not somatic mutations. The full-length transcript encoding Dp427 was decreased in 68% of tumors, while Dp71 variants showed variability of expression. Hierarchical clustering analysis of DMD transcripts distinguished malignant from control tissues. Transcriptomes of primary tumors and tumor cell lines with low DMD expression showed enrichment of specific pathways in the differentially expressed genes. Pathways consistently identified: ECM-receptor interaction, calcium signaling and PI3K-Akt, are also altered in DMD muscle. Notably, low DMD expression was associated with a more advanced stage, older age of onset, and reduced survival across different tumors. Thus, DMD transcription occurs throughout a spectrum of normal tissues. The molecular signature associated with its frequent downregulation in malignancies is concordant with changes found in Duchenne muscles, even though these malignancies originate from tissues never previously associated with dystrophin expression or function. Therefore, the importance of this largest known gene extends beyond its roles identified in DMD, certainly into oncology.

genetics↗

bioRxiv2021

Loss of full-length dystrophin expression results in major cell-autonomous abnormalities in proliferating myoblasts

Duchenne muscular dystrophy (DMD) affects myofibers and muscle stem cells (SC), causing progressive muscle degeneration and repair defects. It was not known whether dystrophic myoblasts--the effector cells of muscle growth and regeneration--are affected. Using a combination of transcriptomic, molecular, functional analyses, and genome-scale metabolic modelling, we demonstrate, for the first time, convergent cell-autonomous abnormalities in primary mouse and human dystrophic myoblasts. In Dmdmdx mouse myoblasts lacking full-length dystrophin transcripts, the expression of 170 other genes was significantly altered. Myod1 (p=2.9e-21) and key muscle genes controlled by MyoD (Myog, Mymk, Mymx, epigenetic regulators, ECM interactors, calcium signalling and fibrosis genes) were significantly downregulated. Gene ontology enrichment analysis indicated significant alterations in genes involved in muscle development and function. These transcriptomic abnormalities translated into functional alterations such as increased proliferation (p=3.0e-3), reduced chemotaxis towards both sera-rich (p=3.8e-2) and cytokine-containing medium (p=1.0e-2), and significantly accelerated differentiation in 3D organotypic cultures. These altered myoblast functions are essential for muscle regeneration. The defects were caused by the loss of expression of full-length dystrophin, as strikingly similar and not exacerbated alterations were also observed in dystrophin-null Dmdmdx-{beta}geo myoblasts. Corresponding abnormalities were identified in an established dystrophic mouse muscle (SC5) cell line and human DMD primary myoblasts, confirming universal, cross-species and cell-autonomous nature of these defects. The genome-scale metabolic analysis in human DMD myoblasts indicated significant alteration in the rate of glycolysis/gluconeogenesis (log2FC = 4.8), leukotriene metabolism (log2FC = 4.754), mitochondrial beta-oxidation of branched-chain, odd-chain, and di-unsaturated fatty acids (n-6) (log2FC = -1.187, log2FC = -0.8295 and log2FC = -0.655). These results demonstrate the disease continuum: DMD defects in satellite cells cause myoblast dysfunctions affecting muscle regeneration, which is essential to counteract myofiber loss. Contrary to the established belief, our data demonstrate that typical DMD alterations occur in myoblasts, making these cells a novel therapeutic target for the treatment of this lethal disease.

pathology↗

bioRxiv2020

PSD-95 in dorsal CA1 contributes to the persistence of fear memory

The ability to extinguish fearful memories is essential for survival. Accumulating data indicate that the dorsal CA1 area (dCA1) contributes to this process. However, the cellular and molecular basis of fear memory extinction remains poorly understood. Postsynaptic density protein 95 (PSD-95) regulates the structure and function of glutamatergic synapses. Here, using dCA1-targeted genetic and chemogenetic manipulations in vivo combined with PSD-95 immunostaining and 3D electron microscopy ex vivo, we demonstrate that phosphorylation of PSD-95 at serine 73 PSD-95(S73) is necessary for contextual fear extinction-induced expression of PSD-95 and synaptic plasticity. Moreover, PSD-95(S73) phosphorylation is not necessary for fear memory formation and recall but is required for extinction of contextual fear. Overall, our data shows how PSD-95-dependent synaptic plasticity in the hippocampus contributes to the persistence of fear memories.

neuroscience↗

bioRxiv2020

Effects of L-DOPA on gene expression in the frontal cortex of rats with unilateral lesion of midbrain dopaminergic neurons

The development of Parkinsons disease (PD) causes dysfunction of the frontal cortex, which contributes to hallmark motor symptoms and is regarded as one of the primary causes of the affective and cognitive impairments observed in PD. Treatment with L-DOPA alleviates motor symptoms but has mixed efficacy in restoring normal cognitive functions, which is further complicated by the psychoactive effects of the drug. In this study, we investigated how L-DOPA affects gene expression in the frontal cortex in an animal model of unilateral PD. We performed an RNA-seq analysis of gene expression in the frontal cortex of rats with 6-hydroxydopamine (6-OHDA)-induced unilateral dopaminergic lesion that were treated with L-3,4-dihydroxyphenylalanine (L-DOPA), for 2 weeks. We used analysis of variance to identify differentially expressed genes and found 48 genes with significantly altered transcript abundance after L-DOPA treatment. We also performed a weighted gene coexpression network analysis (WGCNA), which resulted in the detection of 5 modules consisting of genes with similar expression patterns. The analyses led to three primary observations. First, the changes in gene expression induced by L-DOPA were bilateral, although only one hemisphere was lesioned. Second, the changes were not restricted to neurons but also appeared to emerge in immune or endothelial cells. Finally, comparisons with databases of drug-induced gene expression signatures revealed multiple nonspecific effects, which indicates that a part of the observed response is a common pattern activated by multiple types of pharmaceuticals in different target tissues. Taken together, our results identify cellular mechanisms in the frontal cortex that are involved in the response to L-DOPA treatment.

neuroscience↗

bioRxiv2019

Contribution of PSD-95 protein to reward location memory

The molecular mechanisms involved in formation of memory are still poorly understood. We focus here on the function of post-synaptic density protein 95 (PSD-95) and its phosphorylation by CaMKII in spontaneous learning about reward location in female mice. We show that formation of reward location memory leads to downregulation of PSD-95 protein in dendritic spines of the stratum radiatum, area CA1, and selective shrinkage of dendritic spines that contain PSD-95. ShRNA-driven, long-term downregulation of PSD-95 in the area CA1 decreases precision of memory. Autophosphorylation deficient CaMKII mutant mice (CaMKII:T286A) need more time than wild-type animals to learn the location of reward. The same impairment is observed after CA1-targeted overexpression of CaMKII phosphorylation-deficient form of PSD-95 (PSD-95:S73A). In contrast to young adult mice, in aged animals reward location learning affects only spines that lack PSD-95. The frequency and size of the spines without PSD-95 are increased, while shRNA targeted to PSD-95 affects neither speed of learning nor precision of memory indicating alternative mechanisms to support successful memory formation in old mice. Altogether, our data suggest that dynamic regulation of PSD-95 expression is a mechanism that accelerates learning and improves precision of reward location memory in young mice. The function of PSD-95 in memory processes changes in aged animals.

neuroscience↗