The rockefeller university

The camelid single-domain antibody fragment, commonly referred to as a nanobody, achieves the targeting power of conventional monoclonal antibodies (mAbs) at only a fraction of their size. Isolated from camelid species (including llamas, alpacas, and camels), their small size at similar to 15 kDa, low structural complexity, and high stability compared with conventional antibodies have propelled nanobody technology into the limelight of biologic development. Nanobodies are proving themselves to be a potent complement to traditional mAb therapies, showing success in the treatment of, for example, autoimmune diseases and cancer, and more recently as therapeutic options to treat infectious diseases caused by rapidly evolving biological targets such as the SARS-CoV-2 virus. This review highlights the benefits of applying a proteomic approach to identify diverse nanobody sequences against a single antigen. This proteomic approach coupled with conventional yeast/phage display methods enables the production of highly diverse repertoires of nanobodies able to bind the vast epitope landscape of an antigen, with epitope sampling surpassing that of mAbs. Additionally, we aim to highlight recent findings illuminating the structural attributes of nanobodies that make them particularly amenable to comprehensive antigen sampling and to synergistic activity-underscoring the powerful advantage of acquiring a large, diverse nanobody repertoire against a single antigen. Lastly, we highlight the efforts being made in the clinical development of nanobodies, which have great potential as powerful diagnostic reagents and treatment options, especially when targeting infectious disease agents.

The prototypic IFN-inducible transcription factor, IRF1, not only controls inflammatory gene expression but also regulates T cell and macrophage fate specification and function. Using bone marrow chimeras (80% B6.129S2-Ighmtm1Cgn/J [mMT] + 20% B6.129S2-Irf1tm1Mak/J [Irf12/2]), we show that IRF1 expression in B cells is required for marginal zone B (MZB) cell development and T cell- independent Ab responses. Although IFNs can induce IRF1 expression in MZB precursors, deletion of the IFN-gR (C57BL/6J [B6], B6.129S7-Ifngr1tm1Agt/J) or IFN-aR (B6[Cg]-Ifnar1tm1Agt/J) did not affect MZB cell development. Instead, BCR and TLR signals promote IRF1 expression and nuclear translocation in MZB cell precursors. In turn, IRF1 is required for Notch2-dependent gene expression in BCR- and TLR-stimulated transitional B cells and development of the MZB cell compartment. Thus, IRF1 regulates MZB-driven T cell- independent Ab responses by regulating Notch programming in MZB precursors and facilitating commitment of these cells to the MZB lineage. The Journal of Immunology, 2024, 213: 1771-1786.

Mitochondria are semiautonomous organelles with diverse metabolic and cellular functions including anabolism and energy production through oxidative phosphorylation. Following the pioneering observations of Otto Warburg nearly a century ago, an immense body of work has examined the role of mitochondria in cancer pathogenesis and progression. Here, we summarize the current state of the field, which has coalesced around the position that functional mitochondria are required for cancer cell proliferation. In this review, we discuss how mitochondria influence tumorigenesis by impacting anabolism, intracellular signaling, and the tumor microenvironment. Consistent with their critical functions in tumor formation, mitochondria have become an attractive target for cancer therapy. We provide a comprehensive update on the numerous therapeutic modalities targeting the mitochondria of cancer cells making their way through clinical trials.

Mammalian retrotransposons constitute 40% of the genome. During tissue regeneration, adult stem cells coordinately repress retrotransposons and activate lineage genes, but how this coordination is controlled is poorly understood. Here, we observed that dynamic expression of histone methyltransferase SETDB1 (a retrotransposon repressor) closely mirrors stem cell activities in murine skin. SETDB1 ablation leads to the reactivation of endogenous retroviruses (ERVs, a type of retrotransposon) and the assembly of viral- like particles, resulting in hair loss and stem cell exhaustion that is reversible by antiviral drugs. Mechanistically, at least two molecularly and spatially distinct pathways are responsible: antiviral defense mediated by hair follicle stem cells and progenitors and antiviral-independent response due to replication stress in transient amplifying cells. ERV reactivation is promoted by DNA demethylase ten-eleven translocation (TET)mediated hydroxymethylation and recapitulated by ablating cell fate transcription factors. Together, we demonstrated ERV silencing is coupled with stem cell activity and essential for adult hair regeneration.

Signal transducer and activator of transcription 3 (STAT3) plays a key role in leukocytic and non-leukocytic cells. Germ line mutations in STAT3, which are mainly found in the SH2, DNA binding and transactivation domains, can be loss- or gain-of-function (LOF and GOF). STAT3 N-terminal domain (NTD) mutations are rare, and their biological effects remain incompletely understood. We explored the significance of STAT3 NTD p.Trp37* variant in a patient with chronic pulmonary aspergillosis and a low Hyper-IgE syndrome (HIES) score. In cell culture models, the expression of full-length p.Trp37* allele showed shorter STAT3 protein expression suggesting a re-initiation (Met99 or Met143). STAT3 activity using luciferase reporter assay showed a twofold-increased activity of the STAT3 p.Trp37* STAT3 protein compared with WT STAT3 at basal level and upon IL-6 stimulation. In contrast, the activity of the short pTrp37* peptide (amino acids 1 to 37) was amorphic but without dominant negative (DN) effect on transcriptional activity or STAT3 Tyr705 phosphorylation. The proteins initiated at Met99 and Met143 were surprisingly hypermorphic. In carriers' peripheral blood mononuclear cells (PBMCs), both WT and mutated STAT3 mRNA were equally present and the global amount of STAT3 protein was not significantly reduced. In stimulated heterozygous carriers' PBMCs, however, STAT3 Tyr705 phosphorylation and Th17 were reduced but not completely abolished. This suggests a DN effect of an unknown product of the p.Trp37* allele. Transcriptomics analysis of PBMCs from the index revealed selectively distinct gene expression. We conclude that heterozygosity for the NTD p.Trp37* STAT3 mutation defines a novel allelic form of STAT3 deficiency, associated with a chronic pulmonary aspergillosis and minor signs of HIES.

Background: International Dermatology Outcome Measures (IDEOM) is a non-profit organization whose mission is to improve the availability of evidence-based, consensus-driven outcome measures for dermatological diseases. IDEOM facilitates collaboration between stakeholders from various backgrounds, including researchers, patients, physicians, and industry representatives, to develop objective benchmark metrics that enable better treatment and management of dermatologic conditions. Summary: The 2023 IDEOM Annual Meeting was held June 23-24, 2023. Workgroups in psoriatic disease, hidradenitis suppurative, geriatric dermatology, connective tissue disease, vitiligo, itch, actinic keratosis, acne, and cutaneous T-cell lymphoma discussed the progress of their respective outcome-measures research. This report summarizes each workgroup's updates since the 2022 annual meeting and their next steps as established during the 2023 IDEOM Annual Meeting.

The azimuthal correlation angle, Delta phi, between the scattered lepton and the leading jet in deep inelastic e(+/-) p scattering at HERA has been studied using data collvected with the ZEUS detector at a centre-of-mass energy of root s = 318 GeV, corresponding to an integrated luminosity of 326 pb(-1). A measurement of jet cross sections in the laboratory frame was made in a fiducial region corresponding to photon virtuality 10 GeV2 < Q(2) < 350 GeV2, inelasticity 0.04 < y < 0.7, outgoing lepton energy Ee > 10 GeV, lepton polar angle 140 degrees pi for events with high jet multiplicity, due to limitations of the perturbative approach in describing soft phenomena in QCD. The data are equally well described by Monte Carlo predictions that supplement leading-order matrix elements with parton showering.

In insects, odorant receptors (ORs) are required for the detection of most olfactory cues. We investigated the function of a clade of four duplicated ORs in the hawkmoth Manduca sexta and found that these paralogs encode broadly tuned receptors with overlapping but distinct response spectra. Two paralogs, which arose after divergence from a related lineage, show high sensitivity to floral esters released by a nectar-rich plant frequently visited by M. sexta. Functional imaging in mutant moths lacking one of the paralogs suggests that olfactory sensory neurons expressing this OR target a previously identified feeding-associated glomerulus in the primary olfactory center of the brain. However, only the response of this glomerulus to the single ligand unique to the now mutated OR disappeared, suggesting neuronal coexpression of the paralogs. Our results suggest a link between the studied OR expansion and enhanced detection of odors emitted by valuable nectar sources in M. sexta.

Protein kinase A (PKA) is a key regulator of cellular functions by selectively phosphorylating numerous substrates, including ion channels, enzymes, and transcription factors. It has long served as a model system for understanding the eukaryotic kinases. Using cryoelectron microscopy, we present complex structures of the PKA catalytic subunit (PKA-C) bound to a full- length protein substrate, the cystic fibrosis transmembrane conductance regulator (CFTR)-an ion channel vital to human health. CFTR gating requires phosphorylation of its regulatory (R) domain. Unphosphorylated CFTR engages PKA-C at two locations, establishing two "catalytic stations" near to, but not directly involving, the R domain. This configuration, coupled with the conformational flexibility of the R domain, permits transient interactions of the eleven spatially separated phosphorylation sites. Furthermore, we determined two structures of the open- pore CFTR stabilized by PKA-C, providing a molecular basis to understand how PKA-C stimulates CFTR currents even in the absence of phosphorylation.

Introduction Cystic fibrosis (CF) is a hereditary autosomal recessive disease driven by deleterious variants of the CFTR gene, leading, among other symptoms, to increased lung infection susceptibility. Mucus accumulation in the CF lung is, as of yet, considered as one important factor contributing to its colonization by opportunistic pathogens such as Pseudomonas aeruginosa. However, in recent years evidence was provided that alveolar macrophages, which form the first line of defense against airborne pathogens, seem to be intrinsically defective with regard to bactericidal functionality in the CF lung. To assess the impact of CFTR deficiency in human macrophages only insufficient systems are available.Methods To address this problem and to evaluate the role of CFTR in human macrophages, we successfully differentiated human induced pluripotent stem cells (iPSC) from a CF p.Phe508del homozygous individual and a healthy donor into primitive macrophages (iMac Delta F508 and iMacWT), respectively, and compared the bactericidal functionality in the relevant cell type.Results iMac Delta F508 showed impaired P. aeruginosa clearance and intracellular killing capacity in comparison to iMacWT. Furthermore, iMac Delta F508 exhibited a less acidic lysosomal pH, and upon P. aeruginosa infection, there were signs of mitochondrial fragmentation and autophagosome formation together with a hyperinflammatory phenotype and deficient type I interferon response.Conclusion In summary, we present a defective phenotype in iMac Delta F508 upon P. aeruginosa infection, which will constitute an ideal platform to further study the role of macrophages in the context of CF.