Khakpour, P. | Search bioRxiv

bioRxiv2026

Nuclear Cx43 restrains microglial neurotoxicity during brain development

Microglia are essential for sculpting the developing brain, yet the molecular mechanisms that select beneficial overreactive phagocytosis remain incompletely understood. Connexin 43 (Cx43, encoded by GJA1 in humans) is best known as a gap junction and hemichannel protein, although its non-canonical, channel-independent functions are increasingly recognized. We found that Cx43 is highly expressed in microglia during the perinatal period in human and mouse, whereas proportion of full-length multimeric Cx43 unexpectedly localizes to the nucleoplasm. Deletion of microglial Cx43 in mice during development instigates a transient neurotoxic state with microgliosis, upregulated phagocytic and complement pathways, excessive neuronal apoptosis, translating into depressive-like and cognitive deficits in the adulthood. Notably, neither microglia-specific deletion of Cx43 in adulthood nor hemichannel blockade recapitulate these changes, indicating a channel-independent, developmental stage-specific neuroprotective mechanism. Nucleus-targeted Cx43 overexpression suppresses neurotoxic markers and neural apoptosis. Nuclear Cx43 interacts with transcriptional regulators to restrain proinflammatory gene programs, nuclear import of Cx43 is driven by neurogenic niche-derived bFGF, which triggers AKT-mediated phosphorylation of a C-terminal nucleus localization signal (NLS), 14-3-3 binding, and importin-dependent nucleus translocation. These findings reveal a developmentally restricted nuclear Cx43 function that restrains microglial neurotoxicity while promoting microglial physiological functions thus expanding connexin biology to transcriptional co-regulation and pointing to a potential avenue for therapeutic intervention.

neuroscience↗

bioRxiv2023

Investigation of microglial diversity in a mouse model of Parkinson's disease pathology

Microglia, the central nervous system resident immune cells, are now recognized to critically impact homeostasis maintenance and contribute to the outcomes of various pathological conditions including Parkinsons disease (PD). Microglia are heterogenous, with a variety of states recently identified in aging and neurodegenerative disease models, including the disease-associated microglia (DAM) which present a selective enrichment of CLEC7A encoding the CLEC7A or DECTIN1 protein, and the dark microglia (DM) displaying markers of cellular stress at the ultrastructural level. However, the roles of CLEC7A-positive microglia and DM in the pathology of PD have remained largely elusive. By applying immunofluorescence and scanning electron microscopy, we aimed to characterize 1) the CLEC7A -positive cell population, and 2) their possible relationships to DM in a mouse model harboring a G2019S pathogenic mutation of the LRRK2 gene, the most common mutation linked to PD. We examined 18-month-old mice, comparing between LRRK2 G2019S knock-in mice and wild-type controls. In the dorsal striatum, a region affected by PD pathology, extensive ultrastructural features of cellular stress (e.g., endoplasmic reticulum and Golgi apparatus dilation), as well as reduced direct cellular contacts, were observed for microglia from LRRK2 G2019S mice versus controls. CLEC7A-positive microglia exhibited extensive phagocytic ultrastructural characteristics in the LRRK2 G2019S mice. Additionally, the LRRK2 G2019S mice presented a higher proportion of DM. Lastly, immunofluorescence and biochemical analysis revealed higher number of CLEC7A-positive cells in Lrrk2 G2019S genotype versus controls both in tissues and in primary microglia cells. Of note, CLEC7A-positive cells present a selective enrichment of ameboid morphology and tend to cluster in the pathogenic animal. In summary, we provide novel insights into the involvement of recently-defined microglial states, CLEC7A-positive cells and DM, in the context of LRRK2 G2019S PD pathology.

neuroscience↗