Publications by authors named "Zachary T Wilson"

6 Publications

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Delayed KCNQ1/KCNE1 assembly on the cell surface helps I fulfil its function as a repolarization reserve in the heart.

J Physiol 2021 May 7. Epub 2021 May 7.

Department of Physiology & Biophysics, Virginia Commonwealth University, Richmond, VA, USA.

Key Points: In adult ventricular myocytes, the slow delayed rectifier (I ) channels are distributed on the surface sarcolemma, not t-tubules. In adult ventricular myocytes, KCNQ1 and KCNE1 have distinct cell surface and cytoplasmic pools. KCNQ1 and KCNE1 traffic from the endoplasmic reticulum to the plasma membrane by separate routes, and assemble into I channels on the cell surface. Liquid chromatography/tandem mass spectrometry applied to affinity-purified KCNQ1 and KCNE1 interacting proteins reveals novel interactors involved in protein trafficking and assembly. Microtubule plus-end binding protein 1 (EB1) binds KCNQ1 preferentially in its dimer form, and promotes KCNQ1 to reach the cell surface. An LQT1-associated mutation, Y111C, reduces KCNQ1 binding to EB1 dimer.

Abstract: Slow delayed rectifier (I ) channels consist of KCNQ1 and KCNE1. I functions as a 'repolarization reserve' in the heart by providing extra current for ventricular action potential shortening during β-adrenergic stimulation. There has been much debate about how KCNQ1 and KCNE1 traffic in cells, where they associate to form I channels, and the distribution pattern of I channels relative to β-adrenergic signalling complex. We used experimental strategies not previously applied to KCNQ1, KCNE1 or I , to provide new insights into these issues. 'Retention-using-selected-hook' experiments showed that newly translated KCNE1 constitutively trafficked through the conventional secretory path to the cell surface. KCNQ1 largely stayed in the endoplasmic reticulum, although dynamic KCNQ1 vesicles were observed in the submembrane region. Disulphide-bonded KCNQ1/KCNE1 constructs reported preferential association after they had reached cell surface. An in situ proximity ligation assay detected I channels in surface sarcolemma but not t-tubules of ventricular myocytes, similar to the reported location of adenylate cyclase 9/yotiao. Fluorescent protein-tagged KCNQ1 and KCNE1, in conjunction with antibodies targeting their extracellular epitopes, detected distinct cell surface and cytoplasmic pools of both proteins in myocytes. We conclude that, in cardiomyocytes, KCNQ1 and KCNE1 traffic by different routes to surface sarcolemma where they assemble into I channels. This mode of delayed channel assembly helps I fulfil its function of repolarization reserve. Proteomic experiments revealed a novel KCNQ1 interactor, microtubule plus-end binding protein 1 (EB1). EB1 dimer (active form) bound KCNQ1 and increased its surface level. An LQT1 mutation, Y111C, reduced KCNQ1 binding to EB1 dimer.
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http://dx.doi.org/10.1113/JP281773DOI Listing
May 2021

Unicommissural Unicuspid Aortic Valve, Ascending Aortic Aneurysm, and Bovine Arch: A Rare Case Report.

World J Pediatr Congenit Heart Surg 2021 May 6:2150135120965201. Epub 2021 May 6.

Banner University Medical Center, Phoenix, AZ, USA.

A 24-year-old man presented with rapidly progressive dyspnea due to mixed aortic stenosis and insufficiency. Unicommissural unicuspid aortic valve, ascending aortic aneurysm, and a bovine arch were identified on computed tomography angiography. Uncomplicated surgical mechanical valve replacement and ascending aortic graft placement improved his symptoms. Aortopathy is common in unicuspid valve patients.
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http://dx.doi.org/10.1177/2150135120965201DOI Listing
May 2021

Global identification of S-palmitoylated proteins and detection of palmitoylating (DHHC) enzymes in heart.

J Mol Cell Cardiol 2021 Jun 23;155:1-9. Epub 2021 Feb 23.

Department of Physiology & Biophysics, Virginia Commonwealth University, Richmond, VA, United States. Electronic address:

High-throughput experiments suggest that almost 20% of human proteins may be S-palmitoylatable, a post-translational modification (PTM) whereby fatty acyl chains, most commonly palmitoyl chain, are linked to cysteine thiol groups that impact on protein trafficking, distribution and function. In human, protein S-palmitoylation is mediated by a group of 23 palmitoylating 'Asp-His-His-Cys' domain-containing (DHHC) enzymes. There is no information on the scope of protein S-palmitoylation, or the pattern of DHHC enzyme expression, in the heart. We used resin-assisted capture to pull down S-palmitoylated proteins from human, dog, and rat hearts, followed by proteomic search to identify proteins in the pulldowns. We identified 454 proteins present in at least 2 species-specific pulldowns. These proteins are operationally called 'cardiac palmitoylome'. Enrichment analysis based on Gene Ontology terms 'cellular component' indicated that cardiac palmitoylome is involved in cell-cell and cell-substrate junctions, plasma membrane microdomain organization, vesicular trafficking, and mitochondrial enzyme organization. Importantly, cardiac palmitoylome is uniquely enriched in proteins participating in the organization and function of t-tubules, costameres and intercalated discs, three microdomains critical for excitation-contraction coupling and intercellular communication of cardiomyocytes. We validated antibodies targeting DHHC enzymes, and detected eleven of them expressed in hearts across species. In conclusion, we provide resources useful for investigators interested in studying protein S-palmitoylation and its regulation by DHHC enzymes in the heart. We also discuss challenges in these efforts, and suggest methods and tools that should be developed to overcome these challenges.
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http://dx.doi.org/10.1016/j.yjmcc.2021.02.007DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8154712PMC
June 2021

A Rare Case of Coccidioidomycosis Constrictive Pericarditis.

Circ Cardiovasc Imaging 2020 12 3;13(12):e010825. Epub 2020 Dec 3.

Banner University Medical Center, Phoenix, Arizona.

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http://dx.doi.org/10.1161/CIRCIMAGING.120.010825DOI Listing
December 2020

Delta-S-Cys-Albumin: A Lab Test that Quantifies Cumulative Exposure of Archived Human Blood Plasma and Serum Samples to Thawed Conditions.

Mol Cell Proteomics 2019 10 19;18(10):2121-2137. Epub 2019 Jul 19.

School of Molecular Sciences and The Biodesign Institute at Arizona State University, Tempe, AZ 85287. Electronic address:

Exposure of blood plasma/serum (P/S) to thawed conditions (> -30 °C) can produce biomolecular changes that skew measurements of biomarkers within archived patient samples, potentially rendering them unfit for molecular analysis. Because freeze-thaw histories are often poorly documented, objective methods for assessing molecular fitness before analysis are needed. We report a 10-μl, dilute-and-shoot, intact-protein mass spectrometric assay of albumin proteoforms called "ΔS-Cys-Albumin" that quantifies cumulative exposure of archived P/S samples to thawed conditions. The relative abundance of S-cysteinylated (oxidized) albumin in P/S increases inexorably but to a maximum value under 100% when samples are exposed to temperatures > -30 °C. The difference in the relative abundance of S-cysteinylated albumin (S-Cys-Alb) before and after an intentional incubation period that drives this proteoform to its maximum level is denoted as ΔS-Cys-Albumin. ΔS-Cys-Albumin in fully expired samples is zero. The range (mean ± 95% CI) observed for ΔS-Cys-Albumin in fresh cardiac patient P/S ( = 97) was, for plasma 12-29% (20.9 ± 0.75%) and for serum 10-24% (15.5 ± 0.64%). The multireaction rate law that governs S-Cys-Alb formation in P/S was determined and shown to predict the rate of formation of S-Cys-Alb in plasma and serum samples-a step that enables back-calculation of the time at which unknown P/S specimens have been exposed to room temperature. A blind challenge demonstrated that ΔS-Cys-Albumin can detect exposure of groups ( = 6 each) of P/S samples to 23 °C for 2 h, 4 °C for 16 h, or -20 °C for 24 h-and exposure of individual specimens for modestly increased times. An unplanned case study of nominally pristine serum samples collected under NIH-sponsorship demonstrated that empirical evidence is required to ensure accurate knowledge of archived P/S biospecimen storage history.
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http://dx.doi.org/10.1074/mcp.TIR119.001659DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6773563PMC
October 2019

CBD-1 organizes two independent complexes required for eggshell vitelline layer formation and egg activation in C. elegans.

Dev Biol 2018 10 16;442(2):288-300. Epub 2018 Aug 16.

Department of Biology and Program in Molecular Biology, Pomona College, Claremont, CA 91711, USA. Electronic address:

Metazoan eggs have a specialized coat of extracellular matrix that aids in sperm-egg recognition. The coat is rapidly remodeled after fertilization to prevent polyspermy and establish a more permanent barrier to protect the developing embryo. In nematodes, this coat is called the vitelline layer, which is remodeled into the outermost layer of a rigid and impermeable eggshell. We have identified three key components of the vitelline layer structural scaffold - PERM-2, PERM-4 and CBD-1, the first such proteins to be described in the nematode C. elegans. CBD-1 tethered PERM-2 and PERM-4 to the nascent vitelline layer via two N-terminal chitin-binding domains. After fertilization, all three proteins redistributed from the zygote surface to the outer eggshell. Depletion of PERM-2 and PERM-4 from the scaffold led to a porous vitelline layer that permitted soluble factors to leak through the eggshell and resulted in embryonic death. In addition to its role in vitelline layer assembly, CBD-1 is also known to anchor a protein complex required for fertilization and egg activation (EGG-1-5/CHS-1/MBK-2). We found the PERM complex and EGG complex to be functionally independent, and structurally organized through distinct domains of CBD-1. CBD-1 is thus a multifaceted regulator that promotes distinct aspects of vitelline layer assembly and egg activation. In sum, our findings characterize the first vitelline layer components in nematodes, and provide a foundation through which to explore both conserved and species-specific strategies used by animals to build protective barriers following fertilization.
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http://dx.doi.org/10.1016/j.ydbio.2018.08.005DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6143425PMC
October 2018