| 1. |
- Axelsson, Erik, et al.
(författare)
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The genetic consequences of dog breed formation-Accumulation of deleterious genetic variation and fixation of mutations associated with myxomatous mitral valve disease in cavalier King Charles spaniels
- 2021
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Ingår i: PLOS Genetics. - : Public Library of Science (PLoS). - 1553-7390 .- 1553-7404. ; 17:9
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Tidskriftsartikel (refereegranskat)abstract
- Selective breeding for desirable traits in strictly controlled populations has generated an extraordinary diversity in canine morphology and behaviour, but has also led to loss of genetic variation and random entrapment of disease alleles. As a consequence, specific diseases are now prevalent in certain breeds, but whether the recent breeding practice led to an overall increase in genetic load remains unclear. Here we generate whole genome sequencing (WGS) data from 20 dogs per breed from eight breeds and document a similar to 10% rise in the number of derived alleles per genome at evolutionarily conserved sites in the heavily bottlenecked cavalier King Charles spaniel breed (cKCs) relative to in most breeds studied here. Our finding represents the first clear indication of a relative increase in levels of deleterious genetic variation in a specific breed, arguing that recent breeding practices probably were associated with an accumulation of genetic load in dogs. We then use the WGS data to identify candidate risk alleles for the most common cause for veterinary care in cKCs-the heart disease myxomatous mitral valve disease (MMVD). We verify a potential link to MMVD for candidate variants near the heart specific NEBL gene in a dachshund population and show that two of the NEBL candidate variants have regulatory potential in heartderived cell lines and are associated with reduced NEBL isoform nebulette expression in papillary muscle (but not in mitral valve, nor in left ventricular wall). Alleles linked to reduced nebulette expression may hence predispose cKCs and other breeds to MMVD via loss of papillary muscle integrity.
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| 2. |
- Fabini, Edoardo, et al.
(författare)
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Unveiling the Biochemistry of the Epigenetic Regulator SMYD3
- 2019
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Ingår i: Biochemistry. - : AMER CHEMICAL SOC. - 0006-2960 .- 1520-4995. ; 58:35, s. 3634-3645
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Tidskriftsartikel (refereegranskat)abstract
- SET and MYND domain-containing protein 3 (SMYD3) is a lysine methyltransferase that plays a central role in a variety of cancer diseases, exerting its pro-oncogenic activity by methylation of key proteins, of both nuclear and cytoplasmic nature. However, the role of SMYD3 in the initiation and progression of cancer is not yet fully understood and further biochemical characterization is required to support the discovery of therapeutics targeting this enzyme. We have therefore developed robust protocols for production, handling, and crystallization of SMYD3 and biophysical and biochemical assays for clarification of SMYD3 biochemistry and identification of useful lead compounds. Specifically, a time-resolved biosensor assay was developed for kinetic characterization of SMYD3 interactions. Functional differences in SMYD3 interactions with its natural small molecule ligands SAM and SAH were revealed, with SAM forming a very stable complex. A variety of peptides mimicking putative substrates of SMYD3 were explored in order to expose structural features important for recognition. The interaction between SMYD3 and some peptides was influenced by SAM. A nonradioactive SMYD3 activity assay using liquid chromatography-mass spectrometry (LC-MS) analysis explored substrate features of importance also for methylation. Methylation was notable only toward MAP kinase kinase kinase 2 (MAP3K2_K-260)-mimicking peptides, although binary and tertiary complexes were detected also with other peptides. The analysis supported a random bi-bi mechanistic model for SMYD3 methyltransferase catalysis. Our work unveiled complexities in SMYD3 biochemistry and resulted in procedures suitable for further studies and identification of novel starting points for design of effective and specific leads for this potential oncology target.
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| 3. |
- Parenti, Marco Daniele, et al.
(författare)
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Discovery of the 4-aminopiperidine-based compound EM127 for the site-specific covalent inhibition of SMYD3
- 2022
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Ingår i: European Journal of Medicinal Chemistry. - : Elsevier. - 0223-5234 .- 1768-3254. ; 243
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Tidskriftsartikel (refereegranskat)abstract
- Recent findings support the hypothesis that inhibition of SMYD3 methyltransferase may be a therapeutic avenue for some of the deadliest cancer types. Herein, active site-selective covalent SMYD3 inhibitors were designed by introducing an appropriate reactive cysteine trap into reversible first-generation SMYD3 inhibitors. The 4-amino-piperidine derivative EM127 (11C) bearing a 2-chloroethanoyl group as reactive warhead showed selectivity for Cys186, located in the substrate/histone binding pocket. Selectivity towards Cys186 was retained even at high inhibitor/enzyme ratio, as shown by mass spectrometry. The mode of interaction with the SMYD3 substrate/ histone binding pocket was revealed by crystallographic studies. In enzymatic assays, 11C showed a stronger SMYD3 inhibitory effect compared to the reference inhibitor EPZ031686. Remarkably, 11C attenuated the proliferation of MDA-MB-231 breast cancer cell line at the same low micromolar range of concentrations that reduced SMYD3 mediated ERK signaling in HCT116 colorectal cancer and MDA-MB-231 breast cancer cells. Furthermore, 11C (5 mu M) strongly decreased the steady-state mRNA levels of genes important for tumor biology such as cyclin dependent kinase 2, c-MET, N-cadherin and fibronectin 1, all known to be regulated, at least in part, by SMYD3. Thus, 11C is as a first example of second generation SMYD3 inhibitors; this agent represents a covalent and a site specific SMYD3 binder capable of potent and prolonged attenuation of methyltransferase activity.
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| 4. |
- Talibov, Vladimir O., 1991-, et al.
(författare)
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Biophysical approach reveals a novel allosteric ligand binding site of SMYD3 histone methyltransferase
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- SET-and MYND-domain containing protein 3 (SMYD3) is a lysine methyltransferase that plays a role in epigenetic regulation. The protein was shown to have cancerogenic activities and is considered to be a perspective drug target. Here, we propose a Surface Plasmon Resonance-based (SPR) biophysical platform to aid SMYD3 drug discovery. The SPR screening assay was validated with a small subset of drug-like compounds, and resulted in an assay hit. The hit compound-SMYD3 complex structure was solved, and a new allosteric ligand binding site of the protein was revealed. The interaction was found localized within the previously reported SMYD3-heat shock protein 90 (HSP90) recognition site, thereby rendering the hit compound as a perspective candidate for a development of a protein-protein interface inhibitor.
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| 5. |
- Talibov, Vladimir O, 1991-, et al.
(författare)
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Discovery of an allosteric ligand binding site in SMYD3 lysine methyltransferase
- 2021
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Ingår i: ChemBioChem. - : Wiley. - 1439-4227 .- 1439-7633. ; 22:9, s. 1597-1608
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Tidskriftsartikel (refereegranskat)abstract
- SMYD3 is a multifunctional epigenetic enzyme with lysine methyl transferase activity and various interaction partners. It is implicated in the pathophysiology of cancers but with an unclear mechanism. To discover tool compounds for clarifying its biochemistry and potential as a therapeutic target, a set of drug-like compounds was screened using a biosensor-based competition assay. Diperodon was identified as an allosteric ligand. The ( R )-and ( S )-enantiomers of the racemic drug were isolated and their affinities determined ( K D > = 42 and 84 ÎŒM). Co-crystallization revealed that both enantiomers bind to a previously unidentified allosteric site in the C-terminal protein binding domain, consistent with its weak inhibitory effect. No competition between diperodon and HSP90 (a known SMYD3 interaction partner) was observed although HSP90-SMYD3 binding was confirmed ( K D = 13 ÎŒM). The allosteric site appears to be druggable and suitable for exploration of non-catalytic SMYD3 functions and therapeutics with new mechanisms of action.
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