Glycogen is the principal carbon reserve in Synechocystis sp. PCC 6803. We reconstituted its biosynthetic pathway in vitro-GlgC (glucose-1-phosphate adenylyltransferase), two glycogen synthase isoenzymes (GlgA1, GlgA2) a...Glycogen is the principal carbon reserve in Synechocystis sp. PCC 6803. We reconstituted its biosynthetic pathway in vitro-GlgC (glucose-1-phosphate adenylyltransferase), two glycogen synthase isoenzymes (GlgA1, GlgA2) and the branching enzyme GlgB-to define how supply, polymerization and branching set flux and product structure. GlgA2 shows higher specific activity and cooperates with GlgB-generated branched primers, whereas GlgA1 has higher substrate affinity and responds more to primer concentration. Product profiling links mechanism to architecture: GlgA1 produces more-branched glycogen, while GlgA2 yields longer, less-branched polymers, with GlgB biasing utilization toward GlgA2. The complementary behaviors of GlgA1 and GlgA2 provide capacity for rapid accumulation versus steady-state maintenance and offer dynamic metabolic levers to tune glycogen content and architecture in cyanobacteria.
Mycobacterium tuberculosis (Mtb) employs multiple strategies to evade host immunity, including disruption of antigen presentation. Dendritic cells (DCs) are crucial for effective antigen presentation and T-cell activatio...Mycobacterium tuberculosis (Mtb) employs multiple strategies to evade host immunity, including disruption of antigen presentation. Dendritic cells (DCs) are crucial for effective antigen presentation and T-cell activation. In this study, we show that the mycobacterial protein ESAT-6 impairs monocyte to DC differentiation, with reduced expression of the DC markers CD209 and CD1a. ESAT-6 treatment elevated IL-6 and IL-10 levels, but blocking the biological activity of these cytokines failed to restore DC differentiation. Mechanistically, ESAT-6 suppressed phosphorylation of p65, establishing that ESAT-6 impairs DC differentiation by inhibiting NF-κB activation, a function dependent on the last six amino acids of its C-terminal domain. This mechanism may represent a novel immune evasion strategy employed by Mtb to subvert host adaptive immune responses during infection.
DNA double-strand break (DSB) repair is critical for genome stability and requires chromatin remodeling for efficient processing. Fun30, an ATP-dependent chromatin remodeler, promotes long-range DNA end resection to gene...DNA double-strand break (DSB) repair is critical for genome stability and requires chromatin remodeling for efficient processing. Fun30, an ATP-dependent chromatin remodeler, promotes long-range DNA end resection to generate 3' overhangs, a key step in homologous recombination. Persistent DSBs relocate to the nuclear periphery, particularly through interactions with the inner nuclear membrane protein Mps3 and the nuclear pore complex component Nup84. By tracking a single irreparable break, we show that Fun30 facilitates this relocation. In fun30Δ cells, Mps3 and Nup84 enrichment at DSBs was reduced, indicating impaired tethering. We further demonstrate that Fun30 promotes deposition of the histone variant H2A.Z at DSBs. Thus, Fun30 favors relocation of persistent DSBs to the nuclear periphery by supporting resection and H2A.Z incorporation.
Phosphodiesterase 5 (PDE5) regulates several physiological processes, including cardiovascular function. A familial PDE5A variant resulting in an N-terminal truncation (∆M1-Q91) in PDE5A2 has been linked to premature isc...Phosphodiesterase 5 (PDE5) regulates several physiological processes, including cardiovascular function. A familial PDE5A variant resulting in an N-terminal truncation (∆M1-Q91) in PDE5A2 has been linked to premature ischemic heart disease, but its functional impact is unclear. Using computational analysis and BRET-based biosensors, we show that ∆M1-Q91 deletion alters structural dynamics and reduces the efficacy of cGMP-induced conformational change in PDE5. Molecular dynamics simulations and normal mode analysis using structural models revealed altered dynamics and correlated motions in the mutant. BRET assays showed a higher EC for cGMP-induced, but not sildenafil-induced, conformational change in the ∆M1-Q91 mutant PDE5A2. These findings suggest that M1-Q91 deletion impairs cGMP-mediated allosteric regulation in PDE5A2 without altering inhibitor sensitivity, offering insights into potential precision therapies targeting this variant.
Semaphorin 6A (SEMA6A) and Plexin A2 (PLXNA2) are interacting transmembrane proteins dually serving as both receptor and ligand. SEMA6A drives forward signaling through Plexin A2 while PLXNA2 drives reverse signaling thr...Semaphorin 6A (SEMA6A) and Plexin A2 (PLXNA2) are interacting transmembrane proteins dually serving as both receptor and ligand. SEMA6A drives forward signaling through Plexin A2 while PLXNA2 drives reverse signaling through SEMA6A. We have shown in zebrafish that forward signaling is essential for proper retinal size and lamination, whereas reverse signaling is necessary for retinal maintenance. To further characterize reverse signaling, we performed mass spectrometry analysis of immunopurified murine SEMA6A co-expressed with ABL, a tyrosine kinase implicated in SEMA6A reverse signaling. ABL induced SEMA6A phosphorylation at three intracellular tyrosines, two of which are conserved across vertebrates. Expression of murine wild-type SEMA6A mRNA, but not a tyrosine-to-phenylalanine mutant at the three sites, rescued reverse signaling retinal defects observed in sema6a morphants.
The mechanisms supporting progression of metastatic prostate cancer (PCa) in adipocyte-rich bone marrow remain unclear. We hypothesized that stearoyl-coenzyme A desaturase (SCD) promotes PCa survival in bone by modulatin...The mechanisms supporting progression of metastatic prostate cancer (PCa) in adipocyte-rich bone marrow remain unclear. We hypothesized that stearoyl-coenzyme A desaturase (SCD) promotes PCa survival in bone by modulating stress responses and regulating lipid peroxidation. We show that SCD-high PCa cells are sensitive to SCD loss, showing smaller spheroids, reduced mTOR signaling, and elevated endoplasmic reticulum (ER) stress. SCD expression is further augmented by adipocytes, and SCD loss induces DNA damage and repair activation only with adipocyte exposure. In vivo, pharmacological SCD inhibition reduces tumor size and increases ER stress and DNA damage in SCD-high-expressing bone tumors. These findings suggest SCD plays a role in redox regulation and DNA repair sensitivity, with therapeutic potential for targeting DNA repair pathways in combination with SCD inhibition. Impact statement This study reveals that stearoyl-CoA desaturase (SCD) supports prostate cancer growth in adipocyte-rich bone by regulating redox balance and DNA repair responses, uncovering a metabolic mechanism linking lipid metabolism to genomic stability and suggesting therapeutic potential for combining SCD and DNA repair pathway inhibition.
Universal stress proteins (USPs) have remained an enigma since their first description by Nystrom and Neidhardt in 1992. Despite being upregulated under diverse stresses and found across a range of bacterial species, dec...Universal stress proteins (USPs) have remained an enigma since their first description by Nystrom and Neidhardt in 1992. Despite being upregulated under diverse stresses and found across a range of bacterial species, decades of studies suggested only general and potentially redundant protective functions for USPs. Recent studies have uncovered that USPs are critical regulators of bacterial survival processes in Actinobacteria, most notably in Mycobacterium tuberculosis, one of the most prolific and lethal of human pathogens. This brief review places these recent studies in the context of earlier publications and discusses their importance for future USP research, our understanding of these regulatory proteins, and novel therapeutic options that these proteins present in Mycobacterium tuberculosis, related Actinobacteria, and across diverse bacterial species. Impact Statement Universal stress proteins (USPs) have recently been directly implicated in survival processes in Mycobacteria, related Actinobacteria, and multiple bacterial pathogens. This new understanding identifies these stress-responsive proteins as important targets for mechanistic studies in bacterial survival and promising targets for novel antimicrobial therapeutics.
Circadian regulation in peripheral cells depends on calcium dynamics, but the upstream mechanisms remain unclear. We identify endoplasmic reticulum lipid raft-associated protein 2 (ERLIN2) as a regulator of the periphera...Circadian regulation in peripheral cells depends on calcium dynamics, but the upstream mechanisms remain unclear. We identify endoplasmic reticulum lipid raft-associated protein 2 (ERLIN2) as a regulator of the peripheral clock. Knockdown and overexpression of ERLIN2 in C2C12 skeletal muscle cells show that ERLIN2 positively regulates cryptochrome circadian regulator 1/2 (CRY1/2) transcription and maintains rhythmicity. ERLIN2 regulates inositol 1,4,5-trisphosphate receptor (IPR)-mediated Ca release and activates the calcium/calmodulin-dependent protein kinase II (CaMKII)-mitogen-activated protein kinase (MAPK)-cAMP response element-binding protein (CREB) pathway. ATP induced IPR-dependent Ca transients, CREB phosphorylation, and Per1 expression, reshaping circadian rhythm, effects blocked by IPR, Ca, or CaMKII inhibition. CRY1 enhances and CRY2 suppresses CREB signaling, establishing a feedback loop with ERLIN2. This ERLIN2-Ca-CREB-CRY1/2 axis couples membrane contact sites to circadian regulation. Impact statement This study reveals ERLIN2 as a key regulator linking calcium signaling to circadian rhythms, establishing an ERLIN2-Ca-CREB-CRY1/2 axis that advances understanding of cellular clock control.
Recently, we showed that the Per-ARNT-Sim (PAS) domain-containing phosphoglycerate kinase from Leishmania major (LmPAS-PGK) is imported into both the glycosome and the lysosome; however, the mechanism underlying its dual...Recently, we showed that the Per-ARNT-Sim (PAS) domain-containing phosphoglycerate kinase from Leishmania major (LmPAS-PGK) is imported into both the glycosome and the lysosome; however, the mechanism underlying its dual targeting has remained unclear. LmPAS-PGK contains a C-terminal tripeptide sequence and two dileucine base motifs. To investigate the roles of these sorting signals, we generated different complement cell lines from null mutants by transfecting constructs encoding the wild-type protein, an L230A/L231A mutant, the C-terminal tripeptide deleted variant (∆), a dileucine motif deleted variant (∆), and a double-deletion mutant (∆/∆). Our results demonstrate that the dileucine motif governs the lysosomal targeting of LmPAS-PGK, whereas the C-terminal tripeptide is required for glycosomal localization. Deletion of both motifs abolished trafficking to either organelle, leading to cytosolic redistribution. Notably, ∆, ∆, and ∆/∆ cells displayed lower virulence compared with control cells in infected macrophages, underscoring the importance of proper LmPAS-PGK localization in Leishmania pathogenicity. Impact statement Here, we show the dileucine motif of PAS-PGK of Leishmania major governs its lysosomal targeting, whereas its C-terminal tripeptide is required for glycosomal localization. Cells lacking these domains displayed lower virulence compared with control cells in infected macrophages. These results increase our understanding of intracellular trafficking, as well as host-parasite interactions.
Biological systems are fundamentally rhythmic, with oscillations emerging at multiple scales, from intracellular gene circuits to organ-level coordination. Many of these rhythms, including the circadian clock, arise from...Biological systems are fundamentally rhythmic, with oscillations emerging at multiple scales, from intracellular gene circuits to organ-level coordination. Many of these rhythms, including the circadian clock, arise from feedback-driven genetic networks that interact to produce coherent temporal organisation. In this review, we examine the circadian system as a model for understanding the dynamics of coupled biological oscillators. We introduce the core theoretical concepts of delayed feedback, nonlinearity and coupling, and show how these principles govern the emergence of synchronisation, entrainment, and complex dynamics across cellular populations and tissues. Drawing on tools from nonlinear dynamics, we explore how oscillator models help explain robustness, plasticity, and failure modes in circadian systems. Finally, we discuss how this theoretical framework informs experimental design and translational applications in circadian medicine, from optimising drug timing to understanding rhythm disruptions in disease.
Understanding molecular interactions at the host-pathogen interface is essential to decipher infection mechanisms and develop new therapies. Bacterial surface proteins and host-derived bacterial binding proteins (HBBPs)...Understanding molecular interactions at the host-pathogen interface is essential to decipher infection mechanisms and develop new therapies. Bacterial surface proteins and host-derived bacterial binding proteins (HBBPs) govern colonization, adhesion, and immune modulation, but are difficult to study due to low abundance and transient interactions. Advances in chemical biology and proteomics now enable high-resolution mapping of these dynamic surfaces. Techniques such as bioorthogonal labeling, photo-crosslinking, click chemistry, and enzymatic proximity labeling expand our ability to identify surface-exposed and transient complexes. Combined with mass spectrometry and bioinformatics, they offer an integrated view of host-microbe crosstalk, revealing novel virulence factors and antigenic targets. This review highlights innovative labeling strategies advancing infection biology and immune recognition.
Cystathionine γ-synthase (CGS) and cystathionine γ-lyase (CGL) have highly similar amino acid sequences. CGS catalyzes the generation of cystathionine from acylated l-homoserine and l-cysteine, whereas CGL catalyzes the...Cystathionine γ-synthase (CGS) and cystathionine γ-lyase (CGL) have highly similar amino acid sequences. CGS catalyzes the generation of cystathionine from acylated l-homoserine and l-cysteine, whereas CGL catalyzes the decomposition of cystathionine to produce l-cysteine. Lactobacillus plantarum is a unique bacterium containing two open reading frames of CGL/CGS enzymes in its genome. Structural studies of LpCGS and LpCGL may provide insights into their reaction specificities. In the present study, we elucidated the structure and enzymatic function of LpCGS. We found that LpCGS has substrate specificity toward acetylated rather than succinylated l-homoserine. LpCGS has the characteristic residues E55 and V232 in the substrate-binding pocket, which synergistically confer substrate specificity toward acetylated l-homoserine. These results may facilitate the development of inhibitors of l-methionine and l-cysteine biosynthetic pathways.
The THO complex was initially identified in Saccharomyces cerevisiae with five subunits: Hpr1p, Tho2p, Mft1p, Thp2p, and Tex1p. It plays a major role in mRNA processing and nuclear export. Here, we aimed to identify the...The THO complex was initially identified in Saccharomyces cerevisiae with five subunits: Hpr1p, Tho2p, Mft1p, Thp2p, and Tex1p. It plays a major role in mRNA processing and nuclear export. Here, we aimed to identify the putative homologs in Schizosaccharomyces pombe. Among eight candidates, genetic analysis showed tho1, tho2, and pci2 are essential, while mutants of tho5 and tho7 exhibited growth defects along with genome instability and impaired mRNA export. Subcellular localization studies showed all putative homologs except Tho3 are localized to the nucleus, whereas Pci2 localizes to the nuclear envelope. Yeast two-hybrid and immunoprecipitation-mass spectrometry confirmed Tho1, Tho2, Tho5, and Tho7 form the core THO complex. This work defines the THOC complex in S. pombe and supports Pci2 as a component of TREX-2 at the nuclear periphery during mRNA export.
The inositol phosphate signaling pathway has emerged as a compelling therapeutic target in a broad range of diseases, including osteoporosis, viral infections, metabolic disorders, and cancer metastasis. Inositol phospha...The inositol phosphate signaling pathway has emerged as a compelling therapeutic target in a broad range of diseases, including osteoporosis, viral infections, metabolic disorders, and cancer metastasis. Inositol phosphates regulate essential cellular processes such as insulin signaling, nucleotide synthesis, DNA damage response, and phosphate homeostasis. Given this wide spectrum of physiological roles, the kinases responsible for inositol phosphate biosynthesis-namely IP3Ks, IPMK, ITPK1, IP5-2 K, IP6Ks, and PPIP5Ks-have attracted increased interest over the past decade. Accumulating evidence supports their potential as drug targets in the treatment of obesity, cancer, and aging-related conditions. In this review, structure-guided strategies, particularly those informed by high-resolution crystal structures, are examined for their role in accelerating the discovery and development of small-molecule inhibitors targeting inositol phosphate kinases. Structural insights, advances in therapeutic development, and future directions for improving inhibitor specificity and efficacy are discussed.
COL4A1/A2 disorders are rare, congenital, multisystem disorders caused by mutations in the COL4Α1 or COL4Α2 genes, which encode α chains of collagen IV. There are no curative treatments at present, and intervention is fo...COL4A1/A2 disorders are rare, congenital, multisystem disorders caused by mutations in the COL4Α1 or COL4Α2 genes, which encode α chains of collagen IV. There are no curative treatments at present, and intervention is focused on managing the symptoms. Associazione Famiglie COL4A1/A2 was established in 2021 to provide support for patients and their families, and to promote research into the basic mechanisms of the disorders. As part of FEBS Letters's series on patient advocacy for rare disorders, we interviewed Francesca Manodoro, Vice-President and Treasurer of Associazione Famiglie COL4A1-A2, Tom Van Agtmael, Professor of Matrix Biology and Disease at the University of Glasgow, and Simona Balestrini, Associate Professor of Child Neurology at the University of Florence, on the history of the organisation, ongoing research into these conditions, and the challenges in securing funding for research and translating basic research findings into the clinic.
Foam cells derived from macrophages and smooth muscle cells are formed by the uncontrolled uptake of modified low-density lipoprotein (LDL) and are the main cellular components of atherosclerotic lesions. Uptake of oxidi...Foam cells derived from macrophages and smooth muscle cells are formed by the uncontrolled uptake of modified low-density lipoprotein (LDL) and are the main cellular components of atherosclerotic lesions. Uptake of oxidized LDL (oxLDL) by macrophages occurs via receptor-mediated endocytosis through various scavenger receptors. Although resting macrophages internalize modified LDL mainly via SR-A and CD36 receptors, evidence suggests an important role for LOX-1 in the transformation of macrophages into foam cells, despite the low level of LOX-1 on the surface membrane of resting macrophages. Here we describe novel positive feedback loops involving anaphylatoxin C3a and its receptor, which lead to increased LOX-1 levels in macrophages and reveal the molecular mechanisms underlying these processes. Impact statement Little is known about processes which control the transformation of macrophages into foam cells in atherosclerotic lesions. Here, we describe novel positive feedback loops associated with anaphylatoxin C3a and its receptor, which lead to escalation of oxLDL uptake by macrophages, and reveal the central role of the LOX-1 receptor in this process.
Collaboration has become an essential pillar of modern biological research. From international genome initiatives to interdisciplinary multi-omics projects, research in the life sciences increasingly relies on (multi)ins...Collaboration has become an essential pillar of modern biological research. From international genome initiatives to interdisciplinary multi-omics projects, research in the life sciences increasingly relies on (multi)institutional teamwork. Yet, many collaborations fail to deliver on their promises of innovation, efficiency, and scientific impact. Morten T. Hansen's concept of 'disciplined collaboration' (2009) offers a valuable framework for understanding why collaboration sometimes hinders rather than helps research productivity. In this article, Hansen's principles are repurposed to the context of biological research in universities and research institutes. It is substantiated that selective, well-managed, and strategically aligned collaborations, rather than indiscriminate cooperations, lead to sustainable scientific advancement. The discussion of this paper explores the four major barriers to effective collaboration in academia, the three organizational levers proposed by Hansen, and the evaluative processes necessary for implementing disciplined collaboration in research environments. Finally, Hansen's views on institutional strategies are adapted to cultivate collaborative excellence within life science research in academic institutions.
Circadian clocks are endogenous timekeeping mechanisms that are phylogenetically widespread. Despite the immense diversity of bacterial life, to date, clocks have been identified in few bacterial species. The cyanobacter...Circadian clocks are endogenous timekeeping mechanisms that are phylogenetically widespread. Despite the immense diversity of bacterial life, to date, clocks have been identified in few bacterial species. The cyanobacterial clock is understood in great detail, and the roles of its clock proteins in other types of timing mechanisms and in stress resistance are being studied in an ever-growing range of species. Studies of host-associated microbiomes have shown that host and microbial rhythmicity impact one another reciprocally. However, bacterial rhythms have primarily been studied in species in isolation or in host-associated microbiomes. Here, we summarize the state of the field of microbial chronobiology and propose the hypothesis that rhythmicity could be an emergent property of microbial interactions in free-living bacterial communities.
SETDB1, a H3K9 methyltransferase involved in nuclear transcriptional silencing, also localizes to the cytoplasm through unclear mechanisms. Here, we identify cell density as key regulator of SETDB1 subcellular localizati...SETDB1, a H3K9 methyltransferase involved in nuclear transcriptional silencing, also localizes to the cytoplasm through unclear mechanisms. Here, we identify cell density as key regulator of SETDB1 subcellular localization and demonstrate its role in modulating the Hippo signaling pathway. Under low-density culture, SETDB1 distributes between nucleus and cytoplasm, whereas high-density culture triggers nuclear exclusion and proteasomal degradation. SETDB1 depletion reduces YAP1 phosphorylation and increases nuclear YAP1 accumulation. Transcriptomic analysis of SETDB1 knockout cells revealed upregulation of YAP1-TEAD1 target genes (YTGs). Immunoprecipitation experiments showed that SETDB1 is recruited to YTG promoters via TEAD1 and competes with YAP1 for TEAD1 binding. These findings reveal that SETDB1 regulates Hippo pathway output through YAP1 phosphorylation modulation and competitive transcriptional repression.
The GPN-loop GTPase Npa3 plays a critical role in RNA polymerase II (RNAPII) assembly and nuclear import. We employed here the npa3ΔC mutant, which supports normal RNAPII localization and function, to investigate potenti...The GPN-loop GTPase Npa3 plays a critical role in RNA polymerase II (RNAPII) assembly and nuclear import. We employed here the npa3ΔC mutant, which supports normal RNAPII localization and function, to investigate potential links between Npa3 and target of rapamycin complex I (TORC1) signaling. The npa3ΔC cells exhibited increased sensitivity to rapamycin, a synthetic sickness interaction with tor1Δ, and a delayed growth recovery rate from rapamycin-induced G1 arrest. Co-expression analysis identified LTV1, a gene involved in TORC1 signaling and ribosome nuclear export, as one of the top genes co-expressed with NPA3. Furthermore, overexpression of eukaryotic translation initiation factor 1A (eIF1A, TIF11) or regulator of heterotrimeric G-protein signaling (RGS2) restored growth in npa3ΔC cells under rapamycin treatment. Interestingly, RGS2 also rescued growth under hygromycin B stress. Our findings suggest a genetic interplay between Npa3 and TORC1.