Biology subjects

Kang, H.-S.

Publications and source records attributed to Kang, H.-S..

6 recordsLinked to original sources

bioRxiv2026

Toward understanding drivers of specialized metabolism in Actinomycetota: insights from 1432 transcriptomics datasets for 132 strains

Actinomycetota are major sources of specialized metabolites with applications in drug discovery and agriculture, yet much of their biosynthetic potential remains silent under standard laboratory conditions. Limited understanding of the mechanisms controlling biosynthetic gene cluster (BGC) activation constrains metabolite discovery and production. Here, we generated 1432 RNA-seq datasets from 132 Actinomycetota strains grown in eight media to identify patterns associated with BGC expression. On average, strains expressed 44% of their encoded BGCs across the tested conditions, with more expression observed among known BGCs (61%) compared to uncharacterized BGCs (35%). Co-expression analyses revealed frequent associations between BGCs and transporters, transcriptional regulators, and proteins containing DUF397 and DUF742 domains. Targeted overexpression of candidate genes selected from BGC-associated co-expression modules increased metabolite production, with DUF397- and DUF742-containing operons showing the broadest effects by boosting the levels of several different specialized metabolites. Other genes boosted levels in a metabolite-specific manner. Together, our results support a multilayered model of BGC regulation in Actinomycetota in which BGC expression is shaped by medium composition, BGC-specific regulators, and integration of BGCs into broader transcriptional network modules. By connecting BGC expression to specific media and co-expressed genes, this study also provides a resource for selecting growth conditions and engineering specialized metabolism.

microbiology↗

bioRxiv2026

Molecular basis of SPRTN activation by DNA and ubiquitin

The promiscuous metalloprotease SPRTN is the key enzyme for proteolytic repair of DNA-protein crosslinks (DPCs). To prevent uncontrolled SPRTN activity, its activation must be tightly regulated. Using NMR spectroscopy and in vitro reconstitution, we elucidate the molecular basis of SPRTNs activation by DNA and ubiquitin. We identify an autoinhibitory mechanism governed by intramolecular electrostatic interactions between a negatively charged linker helix and SPRTNs positively charged DNA-binding domains. DNA relieves this autoinhibition by competitively displacing the linker from the DNA-binding domains, thereby inducing a conformational shift to an open, catalytically active state. This open state enables ubiquitin binding to SPRTNs protease domain, further stabilizing the active conformation. Disruption of the interaction between the autoinhibitory linker and the DNA-binding domains locks SPRTN in a constitutively open state, resulting in enhanced protease activity. Collectively, our data reveal how DNA and ubiquitin cooperate to convert SPRTN from an autoinhibited conformation into its active state.

biochemistry↗

bioRxiv2024

Allosteric activation of the SPRTN protease by ubiquitin maintains genome stability

The DNA-dependent protease SPRTN maintains genome stability by degrading toxic DNA-protein crosslinks (DPCs). To understand how SPRTNs promiscuous protease activity is confined to the cleavage of crosslinked proteins, we reconstitute the repair of DPCs including their modification with SUMO and ubiquitin chains, using recombinant human proteins. We discover that DPC ubiquitylation strongly activates SPRTN independently of SPRTNs known ubiquitin-binding domains. Using protein structure prediction, MD simulations and NMR spectroscopy we reveal that ubiquitin binds to an interface at the back of SPRTNs protease domain, promoting an active conformation. Replacing key interfacial residues prevents ubiquitin-dependent activation of SPRTN, which leads to genomic instability and cell cycle defects in cells expressing hypomorphic SPRTN variants that cause premature aging and liver cancer in Ruijs-Aalfs syndrome patients. Collectively, our results demonstrate that SPRTN activation is coupled to the modification of the crosslinked protein, explaining how specificity is achieved during DPC repair.

biochemistry↗

bioRxiv2023

PURA Syndrome-causing mutations impair PUR-domain integrity and affect P-body association

Mutations in the human PURA gene cause the neuro-developmental PURA syndrome. In contrast to several other mono-genetic disorders, almost all reported mutations in this nucleic acid binding protein result in the full disease penetrance. In this study, we observed that patient mutations across PURA impair its previously reported co-localization with processing bodies. These mutations either destroyed the folding integrity, RNA binding or dimerization of PURA. We also solved the crystal structures of the N- and C-terminal PUR domains of human PURA and combined them with molecular dynamics simulations and NMR measurements. The observed unusually high dynamics and structural promiscuity of PURA indicated that this protein is particularly susceptible to mutations impairing its structural integrity. It offers an explanation why even conservative mutations across PURA result in the full penetrance of symptoms in patients with PURA syndrome.

molecular biology↗

bioRxiv2022

The RBM17/SPF45-SAP30BP interaction is essential for splicing in a subset of human short introns

Human pre-mRNA splicing requires the removal of introns with highly variable lengths, from tens to over a million nucleotides. Therefore, mechanisms of intron recognition and splicing are likely not universal. Recently, we reported that splicing in a subset of human short introns with truncated polypyrimidine tracts depends on RBM17 (SPF45), instead of the canonical splicing factor U2AF heterodimer. Here, we demonstrate that SAP30BP, a factor previously implicated in transcriptional control, is an essential splicing cofactor for RBM17. In vitro binding and NMR analyses demonstrate that a U2AF-homology motif (UHM) in RBM17 binds directly to a newly identified UHM-ligand motif (ULM) in SAP30BP. We show that this RBM17-SAP30BP interaction is required to specifically recruit RBM17 to phosphorylated SF3B1 (SF3b155), a U2 snRNP component in active spliceosomes. We propose a unique mechanism for splicing in a subset of short introns, in which SAP30BP guides RBM17 in the assembly of active spliceosomes. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=193 SRC="FIGDIR/small/522300v4_ufig1.gif" ALT="Figure 1"> View larger version (32K): org.highwire.dtl.DTLVardef@12da1c3org.highwire.dtl.DTLVardef@d71e53org.highwire.dtl.DTLVardef@108160corg.highwire.dtl.DTLVardef@1df7275_HPS_FORMAT_FIGEXP M_FIG C_FIG In briefFukumura et al. discover a general splicing mechanism in a subset of human short introns with truncated polypyrimidine tracts. This splicing reaction is mediated by intermediary RBM17-SAP30BP complex, instead of the known U2AF heterodimer. SAP30BP binding to RBM17 may support RBM17 association with active phosphorylated SF3B1 in U2 snRNP. HighlightsO_LIRBM17 (SPF45) is a splicing factor required for a subset of human short introns C_LIO_LISAP30BP is an essential cofactor, which interacts with RBM17 via UHM-ULM binding C_LIO_LIRBM17 forms a weak complex with SAP30BP before its binding with SF3B1 in U2 snRNP C_LIO_LIRBM17-SAP30BP complex supports RBM17 to be recruited to active phosphorylated SF3B1 C_LI

molecular biology↗

bioRxiv2020

Structural evolution of the tissue-specific U2AF2 paralog and alternative splicing factor LS2

The Drosophila melanogaster LS2 protein is a tissue-specific paralog of U2AF2 that mediates testis-specific alternative splicing. In order to understand the structural mechanisms underlying the distinct RNA binding specificity we determined the solution structures of the LS2 RNA recognition motif (RRM) domains and characterized their interaction with cis-regulatory guanosine-rich RNA motifs found in intron regions upstream of alternatively spliced exons. We show that the guanosine-rich RNA adopts a G quadruplex (G4) fold in vitro. The LS2 tandem RRMs adopt canonical RRM folds that are connected by a 38-residue linker that harbors a small helical motif 0. The LS2 RRM2 domain and the 0 helix in the interdomain linker mediate interactions with the G4 RNA. The functional importance of these unique molecular features in LS2 is validated by mutational analysis in vitro and RNA splicing assays in vivo. RNA sequencing data confirm the enrichment of G4-forming LS2 target motifs near LS2-affected exons. Our data indicate a role of G quadruplex structures as cis-regulatory motifs in introns for the regulation of alternative splicing, that engage non-canonical interactions with a tandem RRM protein. These results highlight the intriguing molecular evolution of a tissue-specific splicing factor from its conserved U2AF2 paralog as a result of (retro-) gene duplication in D. melanogaster.

biochemistry↗