Biology subjects

Milenkovic, I.

Publications and source records attributed to Milenkovic, I..

7 recordsLinked to original sources

bioRxiv2026

Evolutionary and biological insights into the paternally expressed Snord116-Ipw-Snord115 gene array at the imprinted Prader-Willi syndrome domain

Placental mammal-specific box C/D small nucleolar RNA (SNORD) genes within the imprinted human 15q11q13 domain have garnered increasing attention because their poorly understood roles in the brain and their potential involvement in Prader-Willi syndrome (PWS). Using two novel knockout (KO) mouse models, we demonstrate that combined deletion of Snord116 and Snord115 genes, but not the intervening Ipw ncRNA gene, leads to partially penetrant perinatal lethality (40-50%). Snord116/115-deficient neonates display postnatal growth impairment, hypoglycemia and endocrine dysregulation, including failure of the postnatal leptin surge. Despite showing no overt alterations in feeding behavior, adult Snord116/115-KO mice recapitulate most phenotypes previously reported in Snord116-KO models. RNA-seq analyses of the hypothalamus, prefrontal cortex and cerebellum reveal limited global changes. However, we observed upregulation of two of the most compelling putative RNA targets of Snord116 and Snord115 (Dgkk and Htr2c, respectively) in the postnatal hypothalamus. Nevertheless, no evidence was found to support efficient Snord115-guided ribose methylation of Htr2c mRNA. Finally, comparative analyses across 64 representative placental mammal species reveal that many PWS-associated SNORD genes, including SNORD115, display greater evolutionary changes than previously appreciated, raising questions about the functional relevance and evolutionary selective pressures that have shaped the diversification of certain family members across species. Overall, our study provides an unbiased re-assessment of the evolutionary, molecular and physiological significance of the paternally expressed Snord116-Ipw-Snord115 genomic interval and highlights the early postnatal period as a critical, yet largely underexplored, developmental window during which recently evolved SNORDs likely function as dispensable fine-tuners of gene expression.

genetics↗

bioRxiv2026

Presynaptic GABAA Receptors Mediate Ethanol-Induced Suppression at a Central Auditory Synapse

Acute alcohol consumption impairs speech perception, particularly in noisy environments, but the synaptic mechanisms underlying this auditory deficit remain unknown. Here, we investigated how low-dose ethanol modulates the endbulb of Held synapse in the Mongolian gerbil cochlear nucleus, the first central relay of the auditory pathway. In vivo recordings showed that ethanol suppresses sound-evoked firing, an effect blocked by GABAA receptor antagonists. Direct presynaptic patch-clamp recordings revealed that ethanol potentiates GABAA currents at the endbulb of Held terminal but not at the postsynaptic spherical bushy cell. A high intra-terminal chloride concentration drives a GABA-mediated depolarization that activates low-threshold Kv1 potassium channels and shunts the presynaptic action potential. The reduced action potential amplitude limits calcium influx and glutamate release, effectively suppressing synaptic transmission. These findings identify ethanol-enhanced presynaptic shunting inhibition at the first central auditory synapse as a cellular mechanism for alcohol-induced auditory dysfunction.

neuroscience↗

bioRxiv2025

Variation in Synaptic Inputs Drives Functional Versatility for Sound Encoding in Globular Bushy Cells

Synaptic convergence is fundamental to neuronal circuit function, underpinning computations such as coincidence detection and signal integration. Across sensory systems, convergence architecture and synaptic input strengths are key for extracting stimulus features and processing of sensory information. In the cochlear nucleus, globular bushy cells (GBCs) receive convergent inputs from multiple auditory nerve fibers via large endbulb of Held terminals. While these inputs vary considerably in size, even among those targeting the same cell, the functional consequences of this variation for sound encoding remain unclear. Here, we investigated how synaptic input variation shapes sound encoding in GBCs of Mongolian gerbils using in vitro conductance-clamp recordings and computational modeling. By simulating synaptic inputs with variable strength distributions, we found that increasing input variation enhances rate coding at the expense of temporal precision. These findings suggest that endbulb strength heterogeneity allows the GBC population to operate along a functional continuum, generating diverse information streams to downstream targets.

neuroscience↗

bioRxiv2025

Systematic benchmarking of basecalling models for RNA modification detection with highly multiplexed nanopore sequencing

Nanopore direct RNA sequencing (DRS) holds promise for advancing our understanding of the epitranscriptome by detecting RNA modifications in native RNA molecules. Recently, Oxford Nanopore Technologies (ONT) has released basecalling models capable of detecting several RNA modifications. However, their accuracy, sensitivity and specificity, as well as cross-reactivity against other modification types, remains largely unexplored. Here, we systematically benchmark modification-aware models by evaluating their performance on a highly-multiplexed panel of synthetic molecules covering all possible sequence contexts, as well as on biological samples from a diverse set of species. We find that modification-aware models reliably detect diverse RNA modification types across a broad range of sequence contexts. However, they are prone to elevated false positive rates and exhibit notable cross-reactivity with other RNA modification types. We show that the use of modification-free controls allows significant, yet incomplete removal of false positives, thus constituting an essential control. Finally, we demonstrate that basecalling error patterns and alterations in current features can identify differentially modified sites, for modifications for which modification-aware models are absent. Overall, our results underscore the utility and accuracy of modification-aware basecalling models for RNA modification detection, while highlighting the importance of including diverse control samples to mitigate false positive rates.

bioinformatics↗

bioRxiv2024

Epitranscriptomic rRNA fingerprinting reveals tissue-of-origin and tumor-specific signatures

Mammalian ribosomal RNA (rRNA) molecules are highly abundant RNAs, decorated with over 220 rRNA modifications. Previous works have shown that some rRNA modification types can be dynamically regulated; however, how and when the mammalian rRNA modification landscape is remodeled remains largely unexplored. Here, we employ direct RNA nanopore sequencing to chart the human and mouse rRNA epitranscriptome across tissues, developmental stages, cell types and disease. Our analyses reveal multiple rRNA sites that are differentially modified in a tissue- and/or developmental stage-specific manner, including previously unannotated modified sites. We demonstrate that rRNA modification patterns can be used for tissue and cell type identification, which we hereby term epitranscriptomic fingerprinting. We then explore rRNA modification patterns in normal-tumor matched samples from lung cancer patients, finding that epitranscriptomic fingerprinting accurately classifies clinical samples into normal and tumor groups from only 250 reads per sample, demonstrating the potential of rRNA modifications as diagnostic biomarkers.

molecular biology↗

bioRxiv2023

Monocyte-derived microglia with Dnmt3a mutation cause motor pathology in aging mice

Microglia are established in embryogenesis forming a self-containing cellular compartment resisting seeding with cells derived from adult definitive hematopoiesis. We report that monocyte-derived macrophages (MoM{Phi}) accumulate in the brain of aging mice with distinct topology, including the nigrostriatum and medulla, but not the frontal cortex. Parenchymal MoM{Phi} adopt bona fide microglia expression profiles. Unlike microglia, these monocyte-derived microglia (MoMg) are due to their hematopoietic origin targets of clonal hematopoiesis (CH). Using a chimeric transfer model, we show that hematopoietic expression of DNMT3AR822H, a prominent mutation in human CH, renders MoMg pathogenic promoting motor deficits resembling atypical Parkinsonian disorders. Collectively, these data establish in a mouse model that MoMg progressively seed the brains of aging healthy mice, accumulate in selected areas, and, when carrying a somatic mutation associated with CH, can contribute to brain pathology.

immunology↗

bioRxiv2021

RPL3L-containing ribosomes modulate mitochondrial activity in the mammalian heart

The existence of naturally occurring ribosome heterogeneity is now a well-acknowledged phenomenon. However, whether this heterogeneity leads to functionally diverse specialized ribosomes is still a controversial topic. Here, we explore the biological function of RPL3L (uL3L), a ribosomal protein (RP) paralog of RPL3 (uL3) that is exclusively expressed in muscle and heart tissues, by generating a viable homozygous Rpl3l knockout mouse strain. We identify a rescue mechanism in which, upon RPL3L depletion, RPL3 becomes upregulated, yielding RPL3-containing ribosomes instead of RPL3L-containing ribosomes that are typically found in cardiomyocytes. Using both ribosome profiling (Ribo-Seq) and a novel orthogonal approach consisting of ribosome pulldown coupled to nanopore sequencing (Nano-TRAP), we find that RPL3L neither modulates translational efficiency nor ribosome affinity towards a specific subset of transcripts. By contrast, we show that depletion of RPL3L leads to increased ribosome-mitochondria interactions in cardiomyocytes, which is accompanied by a significant increase in ATP levels, potentially as a result of mitochondrial activity fine-tuning. Our results demonstrate that the existence of tissue-specific RP paralogs does not necessarily lead to enhanced translation of specific transcripts or modulation of translational output. Instead, we reveal a complex cellular scenario in which RPL3L modulates the expression of RPL3, which in turn affects ribosomal subcellular localization and, ultimately, mitochondrial activity.

molecular biology↗