Transcriptional co-activator complexes play essential roles in regulating gene transcription by integrating signals from transcription factors and chromatin. Among these, the SAGA (Spt-Ada-Gcn5 acetyltransferase) and ATAC (Ada Two-A Containing) complexes represent related histone acetyltransferase assemblies that contain both shared and specific subunits. SAGA and ATAC possess similar and divergent enzymatic activities that shape their contributions to transcriptional control. Here, we review emerging evidence that highlights differential interactions of transcriptional activators with SAGA or ATAC and their differential chromatin-reading capabilities through specific recognition of histone modifications. Distinct requirements for SAGA and for ATAC have been observed in gene regulation important for maintaining cellular homeostasis and for oncogenic transformation, indicating that dysregulation of these complexes can contribute to cancer progression. The development of small-molecule inhibitors targeting key components of SAGA and/or ATAC offers promising strategies to disrupt their recruitment and transcriptional activity in cancer cells. Our review dissects the unique and overlapping functions of SAGA and ATAC, which are crucial for understanding regulation by these complexes in order to exploit them as targets in precision oncology.

Cytoplasmic lattices (CPLs) are filamentous assemblies essential for mammalian embryonic development. They are known to regulate organelle organization, spindle assembly, and protein homeostasis, but their molecular functions remain unclear. Here, we develop a strategy combining cryo-focused ion beam milling and cryo-electron tomography to resolve macromolecular complexes directly in mammalian embryos. Using this approach, we determine the in situ structure of cytoplasmic lattices within 6/8-cell mouse embryos at ~4.7 Å resolution. CPL filaments are built from multiple copies of at least fourteen proteins arranged into a ~4.5 MDa repeating unit. The repeat contains a central cavity that is open at the back and lined with multiple FBXW-SKP1 complexes and three modules, each containing the E2 ubiquitin-conjugating enzyme UBE2D and the E3 ligase UHRF1. We resolve two CPL states: one is consistent with a ubiquitin-charged UBE2D, where ubiquitin is held in an open, inactive conformation by binding the scaffold protein PADI6; the second lacks discernible ubiquitin density and shows structural changes compatible with ubiquitin becoming available for transfer. Our findings support a model in which CPLs function as large ubiquitin ligase assemblies during early embryonic development.

Ion channels possess selectivity filters that are hardwired to ensure the selective passage of ions. Lysosomal two-pore channels are unusual as they are able to switch their cation selectivity in an agonist-specific manner, allowing differential control of organellar activity. TPC2 is permeable to Ca[2+] when activated by the calcium-mobilizing messenger NAADP, but largely Na[+]-selective when activated by the signaling lipid PI(3,5)P2. Co-stimulation increases Ca[2+] but not Na[+] permeability; however, the molecular basis for these specificity switches is not well understood. Here we show that mutation of TPC2 residues within the distal cytosolic linker, which connects the first voltage-sensing-like domain to the pore, rendered TPC2 largely unable to discriminate its agonists and highly calcium-permeable, even in the presence of PI(3,5)P2. This mutation induced a co-activated-like state by disrupting a network of residues that connects the linker to the activation gate. Such deregulated agonist action increased lysosomal Ca[2+] flux and compromised locomotion and viability when expressed in C. elegans. A proximal disease-linked mutation perturbed agonist action in a similar way both in vitro and in vivo. Biased signaling through TPC2 thus proceeds through molecular determinants that are remote from the selectivity filter, affecting Ca[2+] permeability, endo-lysosomal integrity and disease.

Inositol is an essential nutrient for most living organisms, as combinatorial phosphorylation on this cyclic sugar generates key cellular messengers, such as lipid-bound phosphoinositides (PtdInsPs), water-soluble inositol phosphates (InsPs), and high-energy inositol pyrophosphates (PP-InsPs). Although the kinases and phosphatases modifying inositol-derived molecules are well-characterised, the molecular pathways controlling the cellular homeostasis of the inositol backbone and transport carriers remain unclear. Using a combination of LC-MS analysis and a screen based on the inositol-exporting opi1Δ mutant yeast, we here discovered that inositol export is tightly regulated by PP-InsPs and that the high-affinity phosphate transporter Pho84 also acts as an inositol exporter. We further expanded these observations to the mammalian system and revealed that inositol export in human cells is similarly controlled by PP-InsPs, and that the human homolog of the yeast Pho84, GLUT2, contributes to inositol export. In summary, we discovered an evolutionarily conserved crosstalk pathway linking PP-InsPs to both phosphate and inositol homeostasis.

Progressive aging of bone marrow hematopoietic stem cells (HSCs) underlies clonal hematopoiesis and age-associated hematologic disorders. Defining early molecular events driving HSC functional decline is essential for rejuvenation strategies. Here, we identify P-selectin (Selp) as a surface marker that stratifies HSCs into conserved functional and transcriptional states during organismal aging in humans and mice. P-selectin expression increases early during aging and remains elevated in old HSCs. Selp[high]-HSCs exhibit increased DNA damage and bias towards megakaryocytic/myeloid lineage fate, whereas Selp[low]-HSCs maintain metabolic integrity, enhanced antioxidant capacity, and reduced myeloid skewing. Transcriptomic analysis revealed that Selp[high]-HSCs adopt megakaryocytic-primed, pro-inflammatory, and oxidative stress programs, while Selp[low]-HSCs retain lymphoid-associated and redox-balanced signatures consistent with a more preserved stem-cell state. Further ATAC-seq analysis demonstrates distinct chromatin landscapes with Selp[high]-HSCs being enriched for inflammatory and platelet-related regulatory elements and CTCF motifs, but Selp[low]-HSCs displaying accessible ETS-driven networks linked to metabolic fitness and stem cell resilience. Together, these findings uncover conserved heterogeneity during blood stem cell aging and establish P-selectin as an early biomarker and potential therapeutic target to mitigate age-associated hematopoietic decline.

Tissue regeneration after injury is crucial for restoring epithelial structure and function. Upon damage, a regenerative microenvironment forms that provides signalling cues that stimulate stem cell proliferation to replace lost cells. While this process is well understood, how stem cells themselves sense damage, translate this input into their proliferation and shape the regenerative microenvironment remains unclear. Here we show that Draper-Src-Shark signalling in Drosophila intestinal stem cells (ISCs) recognises tissue damage by sensing externalised phosphatidylserine on dying midgut epithelial cells and is required for STAT activation in ISCs to promote their proliferation. Unlike its role in phagocytosis, Draper in ISCs does not promote the clearance of these apoptotic enterocytes but rather facilitates ISC proliferation in the presence of damaged enterocytes. Moreover, Draper-Src-Shark signalling in progenitors regulates STAT transcriptional activity in the adjacent visceral muscle, indicating that progenitors can shape the regenerative microenvironment beyond tissue boundaries. As Src and STAT are also activated in the mammalian intestinal epithelium after damage and tumour formation, these findings may help develop therapies for tissue regeneration, inflammatory diseases and cancer.

The Wnt family ligands carry a lipid moiety that must be shielded from the aqueous environment. During secretion, this is achieved by the cargo receptor Wntless. The effect of acute endocytosis inhibition suggests that Wingless dissociates from Wntless at the apical plasma membrane but is prevented from being released extracellularly by rapid endocytosis. Super-resolution microscopy shows that Wingless then traffics, without Wntless, through specialized Rab7/Rab4-positive endosomes before basolateral release. In the absence of endocytosis, apical release is allowed, albeit in punctae. Similar punctae are formed upon abrogation of the ceramide synthase Schlank. We suggest that they represent insoluble aggregates since they are 'solubilized' by the lipid-binding protein Dally-like protein. Moreover, purified Wingless forms similar-looking structures that are solubilised by liposomes in vitro. These findings suggest that membrane lipid composition could modulate the stability of the Wingless-Wntless complex and/or the lipid bilayer's ability to accommodate the Wingless lipid. Together with rapid endocytosis, this ensures orderly transfer of Wingless from Wntless to the membrane, preventing aggregation, and promoting basolateral secretion.

Oxidative stress and genotoxic damage activate NF-κB signaling through intracellular pathways distinct from those initiated by membrane receptors. DNA damage selectively induces post-translational modifications at lysine residues 277 and 309 of the human ubiquitin-binding protein NEMO to promote NF-κB signaling, but the physiological importance of these modifications in vivo remains unclear. Here, we show that a newly developed mouse model (NEMO[DK]) carrying germline arginine substitutions of the corresponding NEMO lysine residues exhibits B-cell-intrinsic defects in germinal center formation and anti-viral humoral responses. Mechanistically, we identify in NEMO[DK] B-cells a CD40-specific NF-κB signaling defect that is not linked to the well-characterized canonical or noncanonical NF-κB pathways. These B-cells fail to secure sustained NEMO monoubiquitination following CD40-induced ROS generation, which specifically reduces downstream RelA (p65) signaling required for the transcriptomic and epigenetic remodeling underlying homotypic B-cell aggregation, cell proliferation, class-switch recombination, and antibody-secreting cell generation. Our results establish a physiological role of murine NEMO K270 and K302 in linking CD40 engagement to B-cell responses through enabling sustained NEMO modification and RelA transcriptional activity.

Mitotic centrosome maturation requires Polo/PLK1-dependent expansion of the pericentriolar material (PCM). In Drosophila, Centrosomin (Cnn) assembles a scaffold around mitotic centrioles through interactions between its PReM and CM2 domains. Here, we show that PReM adopts an autoinhibited helical hairpin conformation that prevents CM2 binding. Polo/PLK1 phosphorylation relieves this autoinhibition, enabling scaffold assembly, whereas phospho-blocking mutations disrupt PReM-CM2 binding in vitro and Cnn scaffold assembly in vivo. Potential functionally analogous domains have been identified in the human and C. elegans Cnn homologues CDK5RAP2 and SPD-5. We find that the human protein appears to share a structurally similar mechanism for scaffold assembly, but the worm protein does not. Consistent with this, deletion of these domains alters the dynamics of Cnn condensates in vitro, but has little effect on SPD-5 condensate dynamics. We conclude that Polo/PLK1 promotes mitotic centrosome assembly, at least in part, by relieving autoinhibitory intramolecular interactions.

Plant plasma membrane proteins are organized into distinct domains; however, the principles that govern this spatial organization are not fully understood. HYPERSENSITIVE-INDUCED REACTION (HIR) proteins are plant-specific members of the stomatin/prohibitin/ flotillin/HflK/C (SPFH) family implicated in membrane organization. Arabidopsis thaliana HIR2 interacts with multiple plasma membrane proteins, including receptor kinases such as BRASSINOSTEROID INSENSITIVE 1-ASSOCIATED KINASE (BAK1)-INTERACTING RECEPTORS 2 and 3 (BIR2 and 3) and BAK1. These interactions connect HIR2 to BAK1-mediated signaling pathways, as evidenced by impaired growth and immunity phenotypes in hir2 mutants. HIR2 is anchored to the inner leaflet of the plasma membrane through a hydrophobic domain and S-acylation. Single-particle-tracking photoactivated localization microscopy (sptPALM) revealed that HIR2 affects BAK1 mobility and clustering, exemplifying a role in regulating plasma membrane dynamics. Structural modeling with AlphaFold 3 predicts a multimeric cup-like assembly for HIR2, consistent with high molecular weight complexes identified through blue-native PAGE. These findings indicate that HIR2 contributes to the formation of membrane sub-compartments, providing a potential structural framework for spatial membrane organization that influences the dynamics and function of membrane-resident receptors.

Nerve nets-characterised by scattered neurons and mesh-like projections-represent a widespread organisation of nervous tissue. Nerve nets have uniquely diffuse conductivity and an apparent simplicity that features in many theories on the evolution of nervous systems. Here we review the anatomical and functional organisation of nerve nets and revisit evolutionary theories in light of new information. We discuss how transgenesis and volume electron microscopy begin to reveal the fine-scale cellular organisation of nerve nets. On the functional level, we describe how excitation can spread within nerve nets and how non-synaptic volume transmission enable multi-layered signalling, driving complex behaviours. As a convergent solution to tissue-scale coordination, we introduce ctenophores, where syncytial nerve nets implement distributed control. We argue that nerve nets are not primitive remnants in some animals or only historical preludes but represent an enduring organisational principle for animal coordination with many unique advantages.

The nucleolus is organized around actively transcribed ribosomal RNA genes (rDNA), where high RNA polymerase I (Pol I) activity creates intrinsic susceptibility to replication stress and DNA damage. Here, we identify the DNA translocase RAD54L as a critical regulator of the nucleolar DNA damage response (nDDR) to rDNA double-strand breaks (DSBs) and replication stress. We show that RAD54L localizes to the nucleolus under basal conditions and is recruited to nucleolar caps following CRISPR-Cas9-induced rDNA-DSBs to promote repair. RAD54L loss results in persistent RAD51 foci, increased nucleolar γH2AX, and micronuclei formation, indicating defective resolution of rDNA lesions and genome instability. Under baseline conditions and replication stress induced by the Pol I transcription inhibitor CX-5461, RAD54L limits the accumulation of ssDNA and coordinates nDDR signaling. We further show that rDNA-DSBs induce RNA polymerase II-dependent RNA-DNA hybrids (R-loops) at intergenic rDNA regions, which facilitate nucleolar reorganization and cap formation and repair factor recruitment. Together, these findings establish RAD54L as a key regulator that coordinates replication stress response and rDNA repair, maintaining rDNA stability and genome integrity.

FUS is an RNA-binding protein mutated in amyotrophic lateral sclerosis (ALS), a neurodegenerative disease characterized by progressive muscle weakness. We show in this work that a heterozygous knock-in mutation in the mouse Fus gene leads to cell-autonomous ultrastructural defects in skeletal muscle, with disruption of sarcomeres and mitochondria. Studies in mouse and Drosophila models demonstrate an evolutionarily conserved cell-autonomous function of FUS in muscle development. Mechanistically, FUS is required for the transcription of MEF2 target genes, binds to the promoter of genes bound by ETS transcription factors, in particular ETV5, and co-activates the transcription of MEF2-dependent genes with ETV5. FUS phase-separates with ETV5 and MEF2A, and stimulation of MEF2-dependent transcription by FUS is dependent upon its phase separation properties. Finally, Etv5 haploinsufficiency exacerbates muscle weakness and atrophy in Fus knock-in mice. Our findings establish a key role for FUS in skeletal muscle differentiation through its phase separation-dependent recruitment of ETV5 and MEF2, defining a novel pathway compromised in FUS-ALS.