Proximity labeling reveals new functional relationships between meiotic recombination proteins in S. cerevisiae.

Several protein ensembles facilitate crossover recombination and the associated assembly of synaptonemal complex (SC) during meiosis. In yeast, meiosis-specific factors including the DNA helicase Mer3, the "ZZS" complex consisting of Zip4, Zip2, and Spo16, the RING-domain protein Zip3, and the MutSγ heterodimer collaborate with crossover-promoting activity of the SC component, Zip1, to generate crossover-designated recombination intermediates. These ensembles also promote SC formation - the organized assembly of Zip1 with other structural proteins between aligned chromosome axes.

Heat-induced structural and chemical changes to a computationally designed miniprotein.

The de novo design of miniprotein inhibitors has recently emerged as a new technology to create proteins that bind with high affinity to specific therapeutic targets. Their size, ease of expression, and apparent high stability makes them excellent candidates for a new class of protein drugs. However, beyond circular dichroism melts and hydrogen/deuterium exchange experiments, little is known about their dynamics, especially at the elevated temperatures they seemingly tolerate quite well.

The methylome of motile cilia.

Cilia are highly complex motile, sensory, and secretory organelles that contain perhaps 1000 or more distinct protein components, many of which are subject to various posttranslational modifications such as phosphorylation, N-terminal acetylation, and proteolytic processing. Another common modification is the addition of one or more methyl groups to the side chains of arginine and lysine residues. These tunable additions delocalize the side-chain charge, decrease hydrogen bond capacity, and increase both bulk and hydrophobicity.

Pain/Stress, Mitochondrial Dysfunction, and Neurodevelopment in Preterm Infants.

Introduction: Preterm infants experience tremendous early life pain/stress during their neonatal intensive care unit (NICU) hospitalization, which impacts their neurodevelopmental outcomes. Mitochondrial function/dysfunction may interface between perinatal stress events and neurodevelopment. Nevertheless, the specific proteins or pathways linking mitochondrial functions to pain-induced neurodevelopmental outcomes in infants remain unidentified.

Methylation of ciliary dynein motors involves the essential cytosolic assembly factor DNAAF3/PF22.

Axonemal dynein motors drive ciliary motility and can consist of up to twenty distinct components with a combined mass of ~2 MDa. In mammals, failure of dyneins to assemble within the axonemal superstructure leads to primary ciliary dyskinesia. Syndromic phenotypes include infertility, rhinitis, severe bronchial conditions, and situs inversus.

CD8+ T Cell-Dependent Antitumor Activity In Vivo of a Mass Spectrometry-Identified Neoepitope despite Undetectable CD8+ Immunogenicity In Vitro.

Identification of neoepitopes that can control tumor growth in vivo remains a challenge even 10 y after the first genomics-defined cancer neoepitopes were identified. In this study, we identify a neoepitope, resulting from a mutation in the junction plakoglobin (Jup) gene (chromosome 11), from the mouse colon cancer line MC38-FABF (C57BL/6). This neoepitope, Jup mutant (JupMUT), was detected during mass spectrometry of MHC class I-eluted peptides from the tumor.

N-Terminal Processing and Modification of Ciliary Dyneins.

Axonemal dyneins are highly complex microtubule motors that power ciliary motility. These multi-subunit enzymes are assembled at dedicated sites within the cytoplasm. At least nineteen cytosolic factors are specifically needed to generate dynein holoenzymes and/or for their trafficking to the growing cilium.

Proteomic interrogation of the meninges reveals the molecular identities of structural components and regional distinctions along the CNS axis.

The meninges surround the brain and spinal cord, affording physical protection while also serving as a niche of neuroimmune activity. Though possessing stromal qualities, its complex cellular and extracellular makeup has yet to be elaborated, and it remains unclear whether the meninges vary along the neuroaxis. Hence, studies were carried-out to elucidate the protein composition and structural organization of brain and spinal cord meninges in normal, adult Biozzi ABH mice.

Associations of Mitochondrial Function, Stress, and Neurodevelopmental Outcomes in Early Life: A Systematic Review.

Early life stress is commonly experienced by infants, especially preterm infants, and may impact their neurodevelopmental outcomes in their early and later lives. Mitochondrial function/dysfunction may play an important role underlying the linkage of prenatal and postnatal stress and neurodevelopmental outcomes in infants. This review aimed to provide insights on the relationship between early life stress and neurodevelopment and the mechanisms of mitochondrial function/dysfunction that contribute to the neuropathology of stress.

Poor maternal diet during gestation alters offspring muscle proteome in sheep.

Poor maternal nutrition during gestation can result in reduced offspring muscle growth and altered muscle metabolism. We hypothesized that over- or restricted-nutrition during gestation would alter the longissimus dorsi muscle (LM) proteome of offspring. Pregnant ewes were fed 60% (restricted), 100% (control), or 140% (over) of National Research Council requirements for total digestible nutrients from day 30 of gestation until parturition.

The Okur-Chung Neurodevelopmental Syndrome Mutation CK2 Leads to a Rewiring of Kinase Specificity.

Okur-Chung Neurodevelopmental Syndrome (OCNDS) is caused by heterozygous mutations to the gene, which encodes the alpha subunit of protein kinase CK2. The most frequently occurring mutation is lysine 198 to arginine (K198R). To investigate the impact of this mutation, we first generated a high-resolution phosphorylation motif of CK2, including the first characterization of specificity for tyrosine phosphorylation activity.

KCNQ2 and KCNQ5 form heteromeric channels independent of KCNQ3.

KCNQ2 and KCNQ3 channels are associated with multiple neurodevelopmental disorders and are also therapeutic targets for neurological and neuropsychiatric diseases. For more than two decades, it has been thought that most KCNQ channels in the brain are either KCNQ2/3 or KCNQ3/5 heteromers. Here, we investigated the potential heteromeric compositions of KCNQ2-containing channels.

Notch signaling enhances bone regeneration in the zebrafish mandible.

Loss or damage to the mandible caused by trauma, treatment of oral malignancies, and other diseases is treated using bone-grafting techniques that suffer from numerous shortcomings and contraindications. Zebrafish naturally heal large injuries to mandibular bone, offering an opportunity to understand how to boost intrinsic healing potential. Using a novel her6:mCherry Notch reporter, we show that canonical Notch signaling is induced during the initial stages of cartilage callus formation in both mesenchymal cells and chondrocytes following surgical mandibulectomy.

Lymphocyte Activation Gene-3 Regulates Dendritic Cell Metabolic Programing and T Cell Priming Function.

Deficiency of lymphocyte activation gene-3 (LAG3) is significantly associated with increased cardiovascular disease risk with in vitro results demonstrating increased TNF-α and decreased IL-10 secretion from LAG3-deficient human B lymphoblasts. The hypothesis tested in this study was that Lag3 deficiency in dendritic cells (DCs) would significantly affect cytokine expression, alter cellular metabolism, and prime naive T cells to greater effector differentiation. Experimental approaches used included differentiation of murine bone marrow-derived DCs (BMDCs) to measure secreted cytokines, cellular metabolism, RNA sequencing, whole cell proteomics, adoptive OT-II donor cells into wild-type (WT) C57BL/6 and recipient mice, and ex vivo measurements of IFN-γ from cultured splenocytes.

An unbiased approach to defining bona fide cancer neoepitopes that elicit immune-mediated cancer rejection.

Identification of neoepitopes that are effective in cancer therapy is a major challenge in creating cancer vaccines. Here, using an entirely unbiased approach, we queried all possible neoepitopes in a mouse cancer model and asked which of those are effective in mediating tumor rejection and, independently, in eliciting a measurable CD8 response. This analysis uncovered a large trove of effective anticancer neoepitopes that have strikingly different properties from conventional epitopes and suggested an algorithm to predict them.

Alternative Splicing of FN (Fibronectin) Regulates the Composition of the Arterial Wall Under Low Flow.

Objective: Exposure of the arterial endothelium to low and disturbed flow is a risk factor for the erosion and rupture of atherosclerotic plaques and aneurysms. Circulating and locally produced proteins are known to contribute to an altered composition of the extracellular matrix at the site of lesions, and to contribute to inflammatory processes within the lesions. We have previously shown that alternative splicing of FN (fibronectin) protects against flow-induced hemorrhage.

A novel phosphoglycerol serine-glycine lipodipeptide of is a TLR2 ligand.

is a Gram-negative anaerobic periodontal microorganism strongly associated with tissue-destructive processes in human periodontitis. Following oral infection with the periodontal bone loss in mice is reported to require the engagement of Toll-like receptor 2 (TLR2). Serine-glycine lipodipeptide or glycine aminolipid classes of engage human and mouse TLR2, but a novel lipid class reported here is considerably more potent in engaging TLR2 and the heterodimer receptor TLR2/TLR6.

A novel urinary biomarker protein panel to identify children with ureteropelvic junction obstruction - A pilot study.

Introduction And Objective: Reliable urinary biomarker proteins would be invaluable in identifying children with ureteropelvic junction obstruction (UPJO) as the existing biomarker proteins are inconsistent in their predictive ability. Therefore, the aim of this study was to identify consistent and reliable urinary biomarker proteins in children with UPJO.

Methods: To identify candidate biomarker proteins, total protein from age-restricted (<2 years) and sex-matched (males) control (n = 22) and UPJO (n = 21) urine samples was analyzed by mass spectrometry.

Hydrogen deuterium exchange defines catalytically linked regions of protein flexibility in the catechol -methyltransferase reaction.

Human catechol -methyltransferase (COMT) has emerged as a model for understanding enzyme-catalyzed methyl transfer from -adenosylmethionine (AdoMet) to small-molecule catecholate acceptors. Mutation of a single residue (tyrosine 68) behind the methyl-bearing sulfonium of AdoMet was previously shown to impair COMT activity by interfering with methyl donor-acceptor compaction within the activated ground state of the wild type enzyme [J. Zhang, H.

Characterizing Protein Kinase Substrate Specificity Using the Proteomic Peptide Library (ProPeL) Approach.

Characterizing protein kinase substrate specificity motifs represents a powerful step in elucidating kinase-signaling cascades. The protocol described here uses a bacterial system to evaluate kinase specificity motifs in vivo, without the need for radioactive ATP. The human kinase of interest is cloned into a heterologous bacterial expression vector and allowed to phosphorylate E.

Glycogen synthase kinase-3 (GSK-3) activity regulates mRNA methylation in mouse embryonic stem cells.

Glycogen synthase kinase-3 (GSK-3) activity regulates multiple signal transduction pathways and is also a key component of the network responsible for maintaining stem cell pluripotency. Genetic deletion of α and Gsk-3β or inhibition of GSK-3 activity via small molecules promotes stem cell pluripotency, yet the mechanism underlying the role for GSK-3 in this process remains ambiguous. Another cellular process that has been shown to affect stem cell pluripotency is mRNA methylation (mA).

The Cac2 subunit is essential for productive histone binding and nucleosome assembly in CAF-1.

Nucleosome assembly following DNA replication controls epigenome maintenance and genome integrity. Chromatin assembly factor 1 (CAF-1) is the histone chaperone responsible for histone (H3-H4) deposition following DNA synthesis. Structural and functional details for this chaperone complex and its interaction with histones are slowly emerging.

DNA-mediated association of two histone-bound complexes of yeast Chromatin Assembly Factor-1 (CAF-1) drives tetrasome assembly in the wake of DNA replication.

Nucleosome assembly in the wake of DNA replication is a key process that regulates cell identity and survival. Chromatin assembly factor 1 (CAF-1) is a H3-H4 histone chaperone that associates with the replisome and orchestrates chromatin assembly following DNA synthesis. Little is known about the mechanism and structure of this key complex.

The Cac1 subunit of histone chaperone CAF-1 organizes CAF-1-H3/H4 architecture and tetramerizes histones.

The histone chaperone Chromatin Assembly Factor 1 (CAF-1) deposits tetrameric (H3/H4) histones onto newly-synthesized DNA during DNA replication. To understand the mechanism of the tri-subunit CAF-1 complex in this process, we investigated the protein-protein interactions within the CAF-1-H3/H4 architecture using biophysical and biochemical approaches. Hydrogen/deuterium exchange and chemical cross-linking coupled to mass spectrometry reveal interactions that are essential for CAF-1 function in budding yeast, and importantly indicate that the Cac1 subunit functions as a scaffold within the CAF-1-H3/H4 complex.

Molecular Determinants of Tubulin's C-Terminal Tail Conformational Ensemble.

Tubulin is important for a wide variety of cellular processes including cell division, ciliogenesis, and intracellular trafficking. To perform these diverse functions, tubulin is regulated by post-translational modifications (PTM), primarily at the C-terminal tails of both the α- and β-tubulin heterodimer subunits. The tubulin C-terminal tails are disordered segments that are predicted to extend from the ordered tubulin body and may regulate both intrinsic properties of microtubules and the binding of microtubule associated proteins (MAP).