Biology subjects

Melia, C.

Publications and source records attributed to Melia, C..

3 recordsLinked to original sources

bioRxiv2026

Motile ciliophagy promotes ciliary recycling under stress

Motile cilia on eukaryotic cell surfaces are exposed to a wide array of external stresses. Ciliary remodeling in response to stress remains understudied despite being pivotal to cell homeostasis. Here, we apply a commonly used calcium perturbation to mimic hyper-osmotic cellular stress in the model ciliate Tetrahymena thermophila. We find that partially deciliated Tetrahymena, upon calcium shock, internalise entire ciliary axonemes into ring-like configurations we refer to as c-rings. Partially deciliated cells recover and regenerate new cilia over time. We propose that the recycling of c-rings releases axonemal building blocks to allow ciliary regeneration. We use time-course ultrastructure expansion microscopy (U-ExM) to show that the ciliary tubulin code undergoes a dynamic pattern of erasure during the bulk disassembly of doublet microtubules within the c-rings. Using whole cell quantitative proteomics, we find that partial deciliation induces the autophagy of c-rings and activates de novo dynein synthesis by upregulating several axonemal dynein assembly factors (DNAAFs). Transmission electron microscopy (TEM) and confocal imaging confirms that intact ciliary axonemes are encapsulated within VPS13A +ve autophagic vacuoles. We propose that internalised motile cilia undergo a regulated process of bulk degradation we term "motile ciliophagy" for the recycling of axonemal components for cilia regeneration. Our work establishes a link between ciliary turnover via macroautophagy and cellular homeostasis which may be a conserved response to stress.

cell biology↗

bioRxiv2024

DNAAF9 (Shulin) and ARL3 regulate the transport and activation of ciliary Outer Dynein Arms

Multiciliogenesis requires large-scale biosynthesis of motility-powering axonemal inner and outer dynein arm motors (IDA and ODA) prior to their intraflagellar transport (IFT) into cilia. ODAs are inhibited by the packaging chaperone Shulin during ciliogenesis in T. thermophila. How Shulin is released for ODAs to become active inside cilia remains unclear. We establish interactions between DNAAF9 (human Shulin) and mammalian ODA subunits, IFT proteins and the ciliary small GTPase ARL3 using proteomics and in vitro reconstitutions. Mutagenesis combined with biochemical and structural studies reveal that DNAAF9 and Shulin preferentially bind the active Arl3-GTP state. GTP-loaded Arl3 can access, bind and displace Shulin from the packaged ODA-Shulin complex. We propose that once the inhibited ODA complex enters growing cilia, Arl3-GTP displaces Shulin (DNAAF9) and sequesters it away from ODAs promoting activation of their motility specifically inside cilia.

cell biology↗

bioRxiv2021

Architecture of cell-cell junctions in situ reveals a mechanism for bacterial biofilm inhibition

Many bacteria, including the major human pathogen Pseudomonas aeruginosa, are naturally found in multicellular, antibiotic-tolerant biofilm communities, where cells are embedded in an extracellular matrix of polymeric molecules. Cell-cell interactions within P. aeruginosa biofilms are mediated by CdrA, a large, membrane-associated adhesin present in the extracellular matrix of biofilms, regulated by the cytoplasmic concentration of cyclic diguanylate. Here, using electron cryotomography of focused-ion beam milled specimens, we report the architecture of CdrA molecules in the extracellular matrix of P. aeruginosa biofilms at intact cell-cell junctions. Combining our in situ observations at cell-cell junctions with biochemistry, native mass spectrometry and cellular imaging, we demonstrate that CdrA forms an extended structure that projects from the outer membrane to tether cells together via polysaccharide binding partners. We go on to show the functional importance of CdrA using custom single-domain antibody (nanobody) binders. Nanobodies targeting the tip of functional cell-surface CdrA molecules could be used to inhibit bacterial biofilm formation or disrupt pre-existing biofilms in conjunction with bactericidal antibiotics. These results reveal a functional mechanism for cell-cell interactions within bacterial biofilms and highlight the promise of using inhibitors targeting biofilm cell-cell junctions to prevent or treat problematic, chronic bacterial infections.

microbiology↗