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Human Cell Jul 2024Hypertrophic cardiomyopathy (HCM) is frequently caused by mutations in the MYPBC3 gene, which encodes the cardiac myosin-binding protein C (cMyBP-C). Most pathogenic...
Hypertrophic cardiomyopathy (HCM) is frequently caused by mutations in the MYPBC3 gene, which encodes the cardiac myosin-binding protein C (cMyBP-C). Most pathogenic variants in MYPBC3 are either nonsense mutations or result in frameshifts, suggesting that the primary disease mechanism involves reduced functional cMyBP-C protein levels within sarcomeres. However, a subset of MYPBC3 variants are missense mutations, and the molecular mechanisms underlying their pathogenicity remain elusive. Upon in vitro differentiation into cardiomyocytes, induced pluripotent stem cells (iPSCs) derived from HCM patients represent a valuable resource for disease modeling. In this study, we generated two iPSC lines from peripheral blood mononuclear cells (PBMCs) of a female with early onset and severe HCM linked to the MYBPC3: c.772G > A variant. Although this variant was initially classified as a missense mutation, recent studies indicate that it interferes with splicing and results in a frameshift. The generated iPSC lines exhibit a normal karyotype and display hallmark characteristics of pluripotency, including the ability to undergo trilineage differentiation. These novel iPSCs expand the existing repertoire of MYPBC3-mutated cell lines, broadening the spectrum of resources for exploring how diverse mutations induce HCM. They additionally offer a platform to study potential secondary genetic elements contributing to the pronounced disease severity observed in this individual.
Topics: Humans; Induced Pluripotent Stem Cells; Cardiomyopathy, Hypertrophic; Female; Carrier Proteins; Cell Differentiation; Myocytes, Cardiac; Mutation, Missense; Severity of Illness Index; Mutation; Cell Line; Frameshift Mutation; Leukocytes, Mononuclear; Cells, Cultured
PubMed: 38762696
DOI: 10.1007/s13577-024-01073-y -
The Journal of Biological Chemistry Jun 2024Triple-negative breast cancer (TNBC) is an aggressive breast cancer sub-type with limited treatment options and poor prognosis. Currently, standard treatments for TNBC...
Triple-negative breast cancer (TNBC) is an aggressive breast cancer sub-type with limited treatment options and poor prognosis. Currently, standard treatments for TNBC include surgery, chemotherapy, and anti-PDL1 therapy. These therapies have limited efficacy in advanced stages. Myeloid-cell leukemia 1 (MCL1) is an anti-apoptotic BCL2 family protein. High expression of MCL1 contributes to chemotherapy resistance and is associated with a worse prognosis in TNBC. MCL1 inhibitors are in clinical trials for TNBC, but response rates to these inhibitors can vary and predictive markers are lacking. Currently, we identified a 4-member (AXL, ETS1, IL6, EFEMP1) gene signature (GS) that predicts MCL1 inhibitor sensitivity in TNBC cells. Factors encoded by these genes regulate signaling pathways to promote MCL1 inhibitor resistance. Small molecule inhibitors of the GS factors can overcome resistance and sensitize otherwise resistant TNBC cells to MCL1 inhibitor treatment. These findings offer insights into potential therapeutic strategies and tumor stratification for MCL1 inhibitor use in TNBC.
Topics: Humans; Myeloid Cell Leukemia Sequence 1 Protein; Triple Negative Breast Neoplasms; Female; Cell Line, Tumor; Drug Resistance, Neoplasm; Biomarkers, Tumor; Gene Expression Regulation, Neoplastic; Antineoplastic Agents; Interleukin-6; Proto-Oncogene Protein c-ets-1
PubMed: 38762181
DOI: 10.1016/j.jbc.2024.107375 -
European Journal of Cell Biology Jun 2024Varicose veins are the most common venous disorder in humans and are characterized by hemodynamic instability due to valvular insufficiency and orthostatic lifestyle...
Varicose veins are the most common venous disorder in humans and are characterized by hemodynamic instability due to valvular insufficiency and orthostatic lifestyle factors. It is unclear how changes in biomechanical signals cause aberrant remodeling of the vein wall. Our previous studies suggest that Notch signaling is implicated in varicose vein arterialization. In the arterial system, mechanoresponsive ETS1 is a transcriptional activator of the endothelial Notch, but its involvement in sensing disrupted venous flow and varicose vein formation has not been investigated. Here, we use human varicose veins and cultured human venous endothelial cells to show that disturbed venous shear stress activates ETS1-NOTCH4/DLL4 signaling. Notch components were highly expressed in the neointima, whereas ETS1 was upregulated in all histological layers of varicose veins. In vitro microfluidic flow-based studies demonstrate that even minute changes in venous flow patterns enhance ETS1-NOTCH4/DLL4 signaling. Uniform venous shear stress, albeit an inherently low-flow system, does not induce ETS1 and Notch proteins. ETS1 activation under altered flow was mediated primarily by MEK1/2 and, to a lesser extent, by MEK5 but was independent of p38 MAP kinase. Endothelial cell-specific ETS1 knockdown prevented disturbed flow-induced NOTCH4/DLL4 expression. TK216, an inhibitor of ETS-family, prevented the acquisition of arterial molecular identity and loss of endothelial integrity in cells exposed to the ensuing altered shear stress. We conclude that ETS1 senses blood flow disturbances and may promote venous remodeling by inducing endothelial dysfunction. Targeting ETS1 rather than downstream Notch proteins could be an effective and safe strategy to develop varicose vein therapies.
Topics: Humans; Signal Transduction; Proto-Oncogene Protein c-ets-1; Receptor, Notch4; Varicose Veins; Calcium-Binding Proteins; Adaptor Proteins, Signal Transducing; Endothelial Cells; Endothelium, Vascular; Male; Stress, Mechanical; Membrane Proteins; Female; Human Umbilical Vein Endothelial Cells
PubMed: 38759515
DOI: 10.1016/j.ejcb.2024.151420 -
Cureus Apr 2024Splenic infarction is a rare and likely underdiagnosed complication of Epstein-Barr virus (EBV)-associated infectious mononucleosis (IM). Here, we describe an...
Splenic infarction is a rare and likely underdiagnosed complication of Epstein-Barr virus (EBV)-associated infectious mononucleosis (IM). Here, we describe an 18-year-old Guyanese male with persistent severe left-sided abdominal pain found to be EBV positive and have a large splenic infarct, along with a transient decrease in protein C, protein S, and antithrombin III activity levels. He was treated with supportive care and anticoagulated with heparin and apixaban. We review prior reports and perspectives on underlying pathophysiology, diagnosis, and the management of these cases, which likely do not require anticoagulation but may be considered on a per-case basis.
PubMed: 38756285
DOI: 10.7759/cureus.58414 -
Free Radical Biology & Medicine Aug 2024Photodynamic therapy is a noninvasive treatment in which specific photosensitizers and light are used to produce high amounts of reactive oxygen species (ROS), which can...
Photodynamic therapy is a noninvasive treatment in which specific photosensitizers and light are used to produce high amounts of reactive oxygen species (ROS), which can be employed for targeted tissue destruction in cancer treatment or antimicrobial therapy. However, it remains unknown whether lower amounts of ROS produced by mild photodynamic therapy increase lifespan and stress resistance at the organism level. Here, we introduce a novel photodynamic treatment (PDTr) that uses 20 μM hypericin, a photosensitizer that originates from Hypericum perforatum, and orange light (590 nm, 5.4 W/m, 1 min) to induce intracellular ROS formation (ROS), thereby resulting in lifespan extension and improved stress resistance in C. elegans. The PDTr-induced increase in longevity was abrogated by N-acetyl cysteine, suggesting the hormetic response was driven by prooxidative mechanisms. PDTr activated the translocation of SKN-1/NRF-2 and DAF-16/FOXO, leading to elevated expression of downstream oxidative stress-responsive genes, including ctl-1, gst-4, and sod-3. In summary, our findings suggest a novel PDTr method that extends the lifespan of C. elegans under both normal and oxidative stress conditions through the activation of SKN-1 and DAF-16 via the involvement of many antioxidant genes.
Topics: Animals; Caenorhabditis elegans; Oxidative Stress; Longevity; Caenorhabditis elegans Proteins; Photochemotherapy; Photosensitizing Agents; Reactive Oxygen Species; Transcription Factors; Perylene; Anthracenes; Forkhead Transcription Factors; DNA-Binding Proteins; Superoxide Dismutase; NF-E2-Related Factor 2; Gene Expression Regulation; Light; Acetylcysteine
PubMed: 38754743
DOI: 10.1016/j.freeradbiomed.2024.05.023 -
BioRxiv : the Preprint Server For... May 2024Lyme disease is a tick-borne, multisystem infection caused by the spirochete, . Although antibodies have been implicated in the resolution of Lyme disease, the specific...
Lyme disease is a tick-borne, multisystem infection caused by the spirochete, . Although antibodies have been implicated in the resolution of Lyme disease, the specific B cell epitopes targeted during human infections remain largely unknown. In this study, we characterized and defined the structural epitope of a patient-derived bactericidal monoclonal IgG ("B11") against Outer surface protein C (OspC), a homodimeric lipoprotein necessary for tick-mediated transmission and early-stage colonization of vertebrate hosts. High-resolution epitope mapping was accomplished through hydrogen deuterium exchange-mass spectrometry (HDX-MS) and X-ray crystallography. Structural analysis of B11 Fab-OspC complexes revealed the B11 Fabs associated in a 1:1 stoichiometry with the lateral faces of OspC homodimers such that the antibodies are essentially positioned perpendicular to the spirochete's outer surface. B11's primary contacts reside within the membrane proximal regions of α-helices 1 and 6 and adjacent loops 5 and 6 in one OspC monomer. In addition, B11 spans the OspC dimer interface, engaging opposing α-helix 1', α-helix 2', and loop 2-3' in the second OspC monomer. The B11-OspC structure is reminiscent of the recently solved mouse transmission blocking monoclonal IgG B5 in complex with OspC , indicating a mode of engagement with OspC that is conserved across species. In conclusion, we provide the first detailed insight into the interaction between a functional human antibody and an immunodominant Lyme disease antigen long considered an important vaccine target.
PubMed: 38746285
DOI: 10.1101/2024.04.29.591597 -
BioRxiv : the Preprint Server For... May 2024During heart failure, gene and protein expression profiles undergo extensive compensatory and pathological remodeling. We previously observed that fast skeletal myosin...
UNLABELLED
During heart failure, gene and protein expression profiles undergo extensive compensatory and pathological remodeling. We previously observed that fast skeletal myosin binding protein-C (fMyBP-C) is upregulated in diseased mouse hearts. While fMyBP-C shares significant homology with its cardiac paralog, cardiac myosin binding protein-C (cMyBP-C), there are key differences that may affect cardiac function. However, it is unknown if the expression of fMyBP-C expression in the heart is a pathological or compensatory response. We aim to elucidate the cardiac consequence of either increased or knockout of fMyBP-C expression. To determine the sufficiency of fMyBP-C to cause cardiac dysfunction, we generated cardiac-specific fMyBP-C over-expression mice. These mice were further crossed into a cMyBP-C null model to assess the effect of fMyBP-C in the heart in the complete absence of cMyBP-C. Finally, fMyBP-C null mice underwent transverse aortic constriction (TAC) to define the requirement of fMyBP-C during heart failure development. We confirmed the upregulation of fMyBP-C in several models of cardiac disease, including the use of lineage tracing. Low levels of fMyBP-C caused mild cardiac remodeling and sarcomere dysfunction. Exclusive expression of fMyBP-C in a heart failure model further exacerbated cardiac pathology. Following 8 weeks of TAC, fMyBP-C null mice demonstrated greater protection against heart failure development. Mechanistically, this may be due to the differential regulation of the myosin super-relaxed state. These findings suggest that the elevated expression of fMyBP-C in diseased hearts is a pathological response. Targeted therapies to prevent upregulation of fMyBP-C may prove beneficial in the treatment of heart failure.
SIGNIFICANCE STATEMENT
Recently, the sarcomere - the machinery that controls heart and muscle contraction - has emerged as a central target for development of cardiac therapeutics. However, there remains much to understand about how the sarcomere is modified in response to disease. We recently discovered that a protein normally expressed in skeletal muscle, is present in the heart in certain settings of heart disease. How this skeletal muscle protein affects the function of the heart remained unknown. Using genetically engineered mouse models to modulate expression of this skeletal muscle protein, we determined that expression of this skeletal muscle protein in the heart negatively affects cardiac performance. Importantly, deletion of this protein from the heart could improve heart function suggesting a possible therapeutic avenue.
PubMed: 38746225
DOI: 10.1101/2024.04.30.591979 -
EMBO Reports Jun 2024Centrioles organize centrosomes, the cell's primary microtubule-organizing centers (MTOCs). Centrioles double in number each cell cycle, and mis-regulation of this...
Centrioles organize centrosomes, the cell's primary microtubule-organizing centers (MTOCs). Centrioles double in number each cell cycle, and mis-regulation of this process is linked to diseases such as cancer and microcephaly. In C. elegans, centriole assembly is controlled by the Plk4 related-kinase ZYG-1, which recruits the SAS-5-SAS-6 complex. While the kinase activity of ZYG-1 is required for centriole assembly, how it functions has not been established. Here we report that ZYG-1 physically interacts with and phosphorylates SAS-5 on 17 conserved serine and threonine residues in vitro. Mutational scanning reveals that serine 10 and serines 331/338/340 are indispensable for proper centriole assembly. Embryos expressing SAS-5 exhibit centriole assembly failure, while those expressing SAS-5 possess extra centrioles. We show that in the absence of serine 10 phosphorylation, the SAS-5-SAS-6 complex is recruited to centrioles, but is not stably incorporated, possibly due to a failure to coordinately recruit the microtubule-binding protein SAS-4. Our work defines the critical role of phosphorylation during centriole assembly and reveals that ZYG-1 might play a role in preventing the formation of excess centrioles.
Topics: Caenorhabditis elegans; Centrioles; Phosphorylation; Caenorhabditis elegans Proteins; Animals; Cell Cycle Proteins; Protein Binding; Protein Serine-Threonine Kinases; Serine; Amino Acid Sequence; Protein Kinases
PubMed: 38744971
DOI: 10.1038/s44319-024-00157-y -
PLoS Genetics May 2024The Integrator is a multi-subunit protein complex that catalyzes the maturation of snRNA transcripts via 3' cleavage, a step required for snRNA incorporation with snRNP...
The Integrator is a multi-subunit protein complex that catalyzes the maturation of snRNA transcripts via 3' cleavage, a step required for snRNA incorporation with snRNP for spliceosome biogenesis. Here we developed a GFP based in vivo snRNA misprocessing reporter as a readout of Integrator function and performed a genome-wide RNAi screen for Integrator regulators. We found that loss of the Argonaute encoding csr-1 gene resulted in widespread 3' misprocessing of snRNA transcripts that is accompanied by a significant increase in alternative splicing. Loss of the csr-1 gene down-regulates the germline expression of Integrator subunits 4 and 6 and is accompanied by a reduced protein translation efficiency of multiple Integrator catalytic and non-catalytic subunits. Through isoform and motif mutant analysis, we determined that CSR-1's effect on snRNA processing is dependent on its catalytic slicer activity but does not involve the CSR-1a isoform. Moreover, mRNA-sequencing revealed high similarity in the transcriptome profile between csr-1 and Integrator subunit knockdown via RNAi. Together, our findings reveal CSR-1 as a new regulator of the Integrator complex and implicate a novel role of this Argonaute protein in snRNA 3' processing.
Topics: Caenorhabditis elegans; Animals; RNA, Small Nuclear; Caenorhabditis elegans Proteins; Argonaute Proteins; Alternative Splicing; RNA Interference; RNA Processing, Post-Transcriptional; Spliceosomes
PubMed: 38743783
DOI: 10.1371/journal.pgen.1011284