Biology subjects

Veverka, V.

Publications and source records attributed to Veverka, V..

8 recordsLinked to original sources

bioRxiv2026

Regulation of the histone H3K36 methyltransferase Set2 by the histone chaperone Spt6

Histone H3 lysine 36 methylation is a conserved histone modification that is critical for maintaining eukaryotic transcriptional fidelity and genomic stability. In Saccharomyces cerevisiae, this modification is catalyzed by Set2, an ortholog of the mammalian H3K36 methyltransferase SETD2. Previous genetic, biochemical, and structural studies showed that Set2 activity is repressed by a Set2 autoinhibitory domain (AID) and that activation requires the direct binding of the histone chaperone Spt6. To study the role of Spt6 and Set2 autoinhibition in vivo, we have isolated and analyzed multiple classes of Spt6 and Set2 mutants. Our results suggest an autoinhibited form of Set2 in which the catalytic domain is bound by the AID. In strong agreement with our genetic results, biophysical experiments demonstrate that the catalytic domain and AID physically interact, and that the autoinhibition mutants disrupt this interaction. Finally, RNA-seq and ChIP-seq studies show the importance of the Set2-Spt6 interaction for transcription and H3K36 methylation genome-wide. Taken together, our results support a model in which Set2 exists in an inactive, autoinhibited state in vivo through direct catalytic domain-AID interactions, with binding by Spt6 required to release this autoinhibited state.

molecular biology↗

bioRxiv2026

Xeno amino acid alphabets form peptides with familiar secondary structure

Summary paragraphThe central dogma of molecular biology describes how genetic information stored in nucleic acids guides the formation of structured proteins from a single molecular alphabet of twenty amino acids (C20)1,2. The extent to which C20 is uniquely capable of forming structural polymers remains a fundamental open question. Here we demonstrate that peptides built from other, "xeno" amino acid alphabets can adopt protein-like secondary structural motifs. Having designed two different xeno alphabets, with them we constructed both combinatorial random sequence libraries and specific, designed 25-mer sequences. We report sequence-dependent structural motifs that result according to circular dichroism, infrared spectroscopy and nuclear magnetic resonance interpreted by molecular dynamics simulations, supported by extensive quantum mechanical calculations. This evidence, including a solution-phase NMR-resolved helical motif, demonstrates that the potential for amino acids to form structure bearing sequences is not unique to lifes alphabet, and thereby reveals a previously unexplored sequence-structure space. Results inform the search for extraterrestrial life, lay foundations for incorporating novel synthetic functional groups and heteroatoms into structure-bearing alphabets, and provide the first truly independent data with which to test and improve all that has been learned about protein folding from the study of lifes 20 side chains.

biochemistry↗

bioRxiv2025

Ancient amino acid sets enable stable protein folds

Early proteins likely arose from a chemically limited set of amino acids available through prebiotic chemistry, raising a central question in molecular evolution: could such primitive compositions yield stable, functional folds? Using de novo design, we constructed three ancient protein architectures using a reduced, evolution-inspired alphabet of ten amino acids, e.g. lacking all basic and aromatic residues. The resulting structures adopted their intended topologies and showed exceptional resistance to thermal and chemical denaturation. Computational simulations further revealed that proteins built from this restricted alphabet were as mutation-resilient as those using all twenty canonical residues. Besides their evolutionary implications, our results provide a foundation for minimalist protein design and generation of simplified, robust systems in minimal cell engineering.

biochemistry↗

bioRxiv2024

The histone chaperone Spt6 controls chromatin structure through its conserved N-terminal domain

The disassembly and reassembly of nucleosomes by histone chaperones is an essential activity during eukaryotic transcription elongation. This highly conserved process maintains chromatin integrity by transiently removing nucleosomes as barriers and then restoring them in the wake of transcription. While transcription elongation requires multiple histone chaperones, there is little understanding of how most of them function and why so many are required. Here, we show that the histone chaperone Spt6 acts through its acidic, intrinsically disordered N-terminal domain (NTD) to bind histones and control chromatin structure. The Spt6 NTD is essential for viability and its histone binding activity is conserved between yeast and humans. The essential nature of the Spt6 NTD can be bypassed by changes in another histone chaperone, FACT, revealing a close functional connection between the two. Our results have led to a mechanistic model for dynamic cooperation between multiple histone chaperones during transcription elongation.

molecular biology↗

bioRxiv2024

Genomic context-dependent histone H3K36 methylation by three Drosophila methyltransferases and implications for dedicated chromatin readers

Methylation of histone H3 at lysine 36 (H3K36me3) marks active chromatin. The mark is interpreted by epigenetic readers that assist transcription and safeguard the integrity of the chromatin fiber. The chromodomain protein MSL3 binds H3K36me3 to target X-chromosomal genes in male Drosophila for dosage compensation. The PWWP-domain protein JASPer recruits the JIL1 kinase to active chromatin on all chromosomes. Unexpectedly, depletion of K36me3 had variable, locus-specific effects on the interactions of those readers. This observation motivated a systematic and comprehensive study of K36 methylation in a defined cellular model. Contrasting prevailing models, we found that K36me1, K36me2 and K36me3 each represent independent chromatin states. A gene-centric view of the changing K36 methylation landscape upon depletion of the three methyltransferases Set2, NSD and Ash1 revealed local, context-specific methylation signatures. Set2 catalyzes K36me3 predominantly at transcriptionally active euchromatin. NSD places K36me2/3 at defined loci within pericentric heterochromatin and on weakly transcribed euchromatic genes. Ash1 deposits K36me1 at regions with enhancer signatures. The genome-wide mapping of MSL3 and JASPer suggested that they bind K36me2 in addition to K36me3, which was confirmed by direct affinity measurement. This dual specificity attracts the readers to a broader range of chromosomal locations and increases the robustness of their actions.

molecular biology↗

bioRxiv2020

Structural basis of RNA recognition by the SARS-CoV-2 nucleocapsid phosphoprotein

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of the Coronavirus disease 2019 (COVID-19) which is currently negatively affecting the population and disrupting the global economy. SARS-CoV-2 belongs to the +RNA virus family that utilize single-stranded positive-sense RNA molecules as genomes. SARS-CoV-2, like other coronaviruses, has an unusually large genome for a +RNA virus that encodes four structural proteins - the matrix (M), small envelope (E), spike (S) and nucleocapsid phosphoprotein (N) - and sixteen nonstructural proteins (nsp1-16) that together ensure replication of the virus in the host cell. The nucleocapsid phosphoprotein N is essential for linking the viral genome to the viral membrane. Its N-terminal RNA binding domain (N-NTD) captures the RNA genome while the C-terminal domain anchors the ribonucleoprotein complex to the viral membrane via its interaction with the M protein. Here, we characterized the structure of the N-NTD and its interaction with RNA using NMR spectroscopy. We observed a positively charged canyon on the surface of the N-NTD lined with arginine residues suggesting a putative RNA binding site. Next, we performed an NMR titration experiment using an RNA duplex. The observed changes in positions of signals in the N-NTD NMR spectra allowed us to construct a model of the N-NTD in complex with RNA.

biochemistry↗

bioRxiv2020

N-glycosylation blocks and simultaneously fosters different receptor-ligand binding sites: the chameleonic CD44-hyaluronan interaction

While DNA encodes protein structure, glycans provide a complementary layer of information to protein function. As a prime example of the significance of glycans, the ability of the cell surface receptor CD44 to bind its ligand, hyaluronan, is modulated by N-glycosylation. However, the details of this modulation remain unclear. Based on atomistic simulations and NMR, we provide evidence that CD44 has multiple distinct binding sites for hyaluronan, and that N-glycosylation modulates their respective roles. We find that non-glycosylated CD44 favors the canonical sub-micromolar binding site, while glycosylated CD44 binds hyaluronan with an entirely different micromolar binding site. Our findings show (for the first time) how glycosylation can alter receptor affinity by shielding specific regions of the host protein, thereby promoting weaker binding modes. The mechanism revealed in this work emphasizes the importance of glycosylation in protein function and poses a challenge for protein structure determination where glycosylation is usually neglected.

molecular biology↗

bioRxiv2019

Dynamics of transcription-dependent H3K36me3 marking by the SETD2:IWS1:SPT6 ternary complex

SETD2 contributes to gene expression by marking gene bodies with H3K36me3, which is thought to assist in the concentration of transcription machinery at the small portion of the coding genome. Despite extensive genome-wide data revealing the precise localization of H3K36me3 over gene bodies, the physical basis for the accumulation, maintenance, and sharp borders of H3K36me3 over these sites remains rudimentary. Here we propose a model of H3K36me3 marking based on stochastic transcription-dependent placement and transcription-independent spreading. Our analysis of the spatial distributions and dynamic features of these marks indicates that transcription-dependent placement dominates the establishment of H3K36me3 domains compared to transcription-independent spreading processes, and that turnover of H3K36me3 limits its capacity for epigenetic memory. By adding additional terms for asymmetric histone turnover occurring at transcription start sites, our model provides a remarkably accurate representation of H3K36me3 levels and dynamics over gene bodies. Furthermore, we validate our findings by revealing that loss of SPT6 impairs the transcription-coupled activity of the SETD2:IWS1:SPT6 ternary complex, thereby reducing the tight correlation between transcription and H3K36me3 levels at gene bodies.

genomics↗