Biology subjects

Segawa, M.

Publications and source records attributed to Segawa, M..

4 recordsLinked to original sources

bioRxiv2026

Single-nucleoid imaging in whole cells defines the dynamics of the mtDNA life cycle

The mitochondrial genome (mtDNA) is essential for oxidative phosphorylation, and mammalian cells typically contain hundreds or thousands of copies. Although specific aspects of mtDNA replication and degradation have been explored, a complete account of the mtDNA life cycle has remained elusive, particularly for nondividing cells. Using super-resolution and 4D lattice light-sheet imaging, we quantified the full mtDNA life cycle in quiescent primary human cells. We show that cells maintain steady state by replicating and degrading 1.5 {+/-} 0.2% of total mtDNA content each hour, a remarkably rapid flux. Younger mtDNA molecules are closer to the nucleus, spared from degradation, and serve as foci for further replication events. Non-proliferative cells regulate nucleoid density within the mitochondrial network, and the mitochondrial membrane potential sustains mtDNA copy number. These findings provide a foundational understanding of the dynamics underlying mtDNA homeostasis and a mechanism explaining how mutations accumulate in aging and disease.

cell biology↗

bioRxiv2026

Chemogenetic activation induces persistent transcriptomic remodeling and functional plasticity in cultured astrocytes

Astrocytes are essential regulators of neuronal function and brain homeostasis. Recent methodological advances have increasingly revealed their active roles through precise manipulation of astrocyte function in the brain. While chemogenetic strategies, such as Designer Receptors Exclusively Activated by Designer Drugs (DREADDs), have been widely used to recapitulate endogenous G-protein-coupled receptors (GPCRs) signaling and selectively modulate astrocyte activity, the molecular mechanisms accompanying DREADD activation remain incompletely understood. Here, we characterized the transcriptional responses of cultured cortical astrocytes and found that DREADD activation induces profound changes in astrocytic gene expression profiles. Activation of Gi-, Gq-, and Gs-coupled DREADDs elicited distinct transcriptomic responses. Transcription factor activity profiling further revealed that each DREADD subtype selectively modulates a distinct set of transcription factors that collectively shape astrocytic transcriptomic responses, thereby indicating subtype-specific transcriptional mechanisms. Importantly, DREADD activation induced persistent changes in the astrocytic transcriptome that remained detectable even three days after stimulation. These transcriptomic alterations were associated with sustained changes in astrocytic calcium responses to extracellular ATP, indicating plastic changes in astrocytic physiological function. Together, these findings provide a molecular framework for interpreting astrocytic DREADD manipulations and reveal a mechanistic basis for functional plasticity and heterogeneity of astrocytes.

neuroscience↗

bioRxiv2020

High-Throughput Image Analysis of Lipid-Droplet-Bound Mitochondria

Changes to mitochondrial architecture are associated with various adaptive and pathogenic processes. However, quantification of changes to mitochondrial structures are limited by the yet unmet challenge of defining the borders of each individual mitochondrion within an image.. Here, we describe a novel method for segmenting Brown Adipose Tissue (BAT) images. We describe a granular approach to quantifying subcellular structures, particularly mitochondria in close proximity to lipid droplets, peri-droplet mitochondria. In addition, we lay out a novel machine-learning-based mitochondrial segmentation method that eliminates the bias of manual mitochondrial segmentation and improves object recognition compared to conventional thresholding analyses. By applying these methods, we discovered a significant difference between cytosolic and peridroplet BAT mitochondrial H2O2 production, and validated the machine learning algorithm in BAT via norepinephrine-induced mitochondrial fragmentation and comparing manual analyses to the automated analysis. This approach provides a higher-throughput analysis protocol to quantify ratiometric probes in subpopulations of mitochondria in adipocytes.

cell biology↗

bioRxiv2019

Cristae undergo continuous cycles of fusion and fission in a MICOS-dependent manner

The mitochondrial inner membrane can reshape under different physiological conditions. How and at which frequency this occurs in vivo and what are the molecular players involved is unknown. Here we show using state-of-the-art live-cell stimulated emission depletion (STED) super-resolution nanoscopy that crista junctions (CJs) are dynamically fusing and dividing in a reversible and balanced manner at a timescale of seconds. CJ dynamics is strongly reduced in the absence of the MICOS subunit MIC13. Staining of the cristae membrane using different protein markers or two inner mitochondrial membrane-specific dyes revealed that cristae also undergo continuous cycles of fusion and fission. These processes are dependent on MIC13 and occur at a timescale of seconds, resembling CJ dynamics. Our data further suggest that MIC60 acts as a docking platform pioneering CJ formation. Overall, by employing a variety of advanced imaging techniques including FRAP (Fluorescence-Recovery-After Photobleaching), SPT (Single-Particle-Tracking), live-cell STED and confocal Airyscan microscopy we demonstrate that cristae undergo continuous cycles of fusion and fission in a manner that is mechanistically linked to CJ formation and dynamics.

cell biology↗