Biology subjects

Jang, H. S.

Publications and source records attributed to Jang, H. S..

4 recordsLinked to original sources

bioRxiv2025

Interprotomer Communication and Functional Asymmetry in H/ACA snoRNPs

H/ACA small nucleolar ribonucleoproteins (H/ACA snoRNPs) facilitate essential cellular processes such as RNA modification, folding, and stability. Here, we present multiple cryo-EM structures of endogenous, catalytically active insect H/ACA snoRNPs containing two protomers assembled on a two-hairpin H/ACA snoRNA. By characterizing key protein-protein and protein-RNA interactions, we reveal the coordination of pseudouridylation activity across the two protomers which explains the predominance of two-hairpin structures in eukaryotic H/ACA snoRNAs. Moreover, we discovered that several mutations in H/ACA proteins associated with Dyskeratosis congenita directly impair pseudouridine formation suggesting how these mutations disrupt RNA modification and ribosome biogenesis in this disease. Additionally, we uncover coordinated structural changes between Nop10, Nhp2 and the N-terminal extensions of Cbf5 in the 3' protomer that resemble active and inactive conformations and may regulate H/ACA snoRNP activity. In summary, this study provides detailed insight into the structure and function of RNA modification-competent, asymmetric H/ACA snoRNPs, which play pivotal roles in cellular processes including ribosome biogenesis, rRNA folding, (m)RNA modification, and telomere maintenance. Significance statementH/ACA snoRNPs are critical for numerous RNA processes as they utilize different H/ACA snoRNAs to introduce pseudouridines in ribosomal and other non-coding RNAs as well as mRNAs. In addition, H/ACA snoRNPs stabilize and fold RNA thereby contributing to ribosome biogenesis and telomere maintenance. Mutations in H/ACA snoRNP subunits such as dyskerin (pseudouridine synthase), Nhp2, and Nop10 cause the genetic disorder Dyskeratosis congenita. Here, we present cryo-EM structures of active eukaryotic H/ACA snoRNPs, revealing an asymmetric, dimeric complex of two protomers. We identify the key inter-protomer contact hotspots and biochemically characterize mutations at these interfaces. Our findings emphasize the importance of inter-subunit interactions for snoRNP stability and activity, providing mechanistic insight into H/ACA snoRNP function and Dyskeratosis congenita pathogenesis.

biochemistry↗

bioRxiv2023

Ultra-Fast Bioorthogonal Spin-Labeling and Distance Measurements in Mammalian Cells Using Small, Genetically Encoded Tetrazine Amino Acids

Studying protein structures and dynamics directly in the cellular environments in which they function is essential to fully understand the molecular mechanisms underlying cellular processes. Site-directed spin-labeling (SDSL)--in combination with double electron-electron resonance (DEER) spectroscopy--has emerged as a powerful technique for determining both the structural states and the conformational equilibria of biomacromolecules. In-cell DEER spectroscopy on proteins in mammalian cells has thus far not been possible due to the notable challenges of spin-labeling in live cells. In-cell SDSL requires exquisite biorthogonality, high labeling reaction rates and low background signal from unreacted residual spin label. While the bioorthogonal reaction must be highly specific and proceed under physiological conditions, many spin labels display time-dependent instability in the reducing cellular environment. Additionally, high concentrations of spin label can be toxic. Thus, an exceptionally fast bioorthogonal reaction is required that can allow for complete labeling with low concentrations of spin-label prior to loss of signal. Here we utilized genetic code expansion to site-specifically encode a novel family of small, tetrazine-bearing non-canonical amino acids (Tet-v4.0) at multiple sites in green fluorescent protein (GFP) and maltose binding protein (MBP) expressed both in E. coli and in human HEK293T cells. We achieved specific and quantitative spin-labeling of Tet-v4.0-containing proteins by developing a series of strained trans-cyclooctene (sTCO)-functionalized nitroxides--including a gem-diethyl-substituted nitroxide with enhanced stability in cells--with rate constants that can exceed 106 M-1 s-1. The remarkable speed of the Tet-v4.0/sTCO reaction allowed efficient spin-labeling of proteins in live HEK293T cells within minutes, requiring only sub-micromolar concentrations of sTCO-nitroxide added directly to the culture medium. DEER recorded from intact cells revealed distance distributions in good agreement with those measured from proteins purified and labeled in vitro. Furthermore, DEER was able to resolve the maltose-dependent conformational change of Tet-v4.0-incorporated and spin-labeled MBP in vitro and successfully discerned the conformational state of MBP within HEK293T cells. We anticipate the exceptional reaction rates of this system, combined with the relatively short and rigid side chains of the resulting spin labels, will enable structure/function studies of proteins directly in cells, without any requirements for protein purification. TOC O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=77 SRC="FIGDIR/small/525763v2_ufig1.gif" ALT="Figure 1"> View larger version (21K): org.highwire.dtl.DTLVardef@1251523org.highwire.dtl.DTLVardef@18ee6ddorg.highwire.dtl.DTLVardef@d6d4b0org.highwire.dtl.DTLVardef@259a53_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗

bioRxiv2021

ChexMix: A Literature Content Extraction Tool for Bioentities

Biomedical databases grow by more than a thousand new publications every day. The large volume of biomedical literature that is being published at an unprecedented rate hinders the discovery of relevant knowledge from keywords of interest to gather new insights and form hypotheses. A text-mining tool, PubTator, helps to automatically annotate bioentities, such as species, chemicals, genes, and diseases, from PubMed abstracts and full-text articles. However, the manual re-organization and analysis of bioentities is a non-trivial and highly time-consuming task. ChexMix was designed to extract the unique identifiers of bioentities from query results. Herein, ChexMix was used to construct a taxonomic tree with allied species among Korean native plants and to extract the medical subject headings unique identifier of the bioentities, which co-occurred with the keywords in the same literature. ChexMix discovered the allied species related to a keyword of interest and experimentally proved its usefulness for multi-species analysis.

bioinformatics↗

bioRxiv2018

Co-opted transposons help perpetuate conserved higher-order chromosomal structures

Transposable elements (TEs) make up half of mammalian genomes and shape genome regulation by harboring binding sites for regulatory factors. These include architectural proteins--such as CTCF, RAD21 and SMC3--that are involved in tethering chromatin loops and marking domain boundaries. The 3D organization of the mammalian genome is intimately linked to its function and is remarkably conserved. However, the mechanisms by which these structural intricacies emerge and evolve have not been thoroughly probed. Here we show that TEs contribute extensively to both the formation of species-specific loops in humans and mice via deposition of novel anchoring motifs, as well as to the maintenance of conserved loops across both species via CTCF binding site turnover. The latter function demonstrates the ability of TEs to contribute to genome plasticity and reinforce conserved genome architecture as redundant loop anchors. Deleting such candidate TEs in human cells leads to a collapse of such conserved loop and domain structures. These TEs are also marked by reduced DNA methylation and bear mutational signatures of hypomethylation through evolutionary time. TEs have long been considered a source of genetic innovation; by examining their contribution to genome topology, we show that TEs can contribute to regulatory plasticity by inducing redundancy and potentiating genetic drift locally while conserving genome architecture globally, revealing a paradigm for defining regulatory conservation in the noncoding genome beyond classic sequence-level conservation.\n\nOne-sentence summaryCo-option of transposable elements maintains conserved 3D genome structures via CTCF binding site turnover in human and mouse.

genomics↗