Biology subjects

Lai, L. B.

Publications and source records attributed to Lai, L. B..

3 recordsLinked to original sources

bioRxiv2025

Solid-state nanopore sensing reveals conformational changes induced by a mutation in a neuron-specific tRNAArg

We demonstrate that solid-state nanopore sensing is a powerful single-molecule method for analyzing RNA conformational ensembles. As a model, we employed n-Tr20, a neuron-specific cytoplasmic tRNAArgUCU, whose C50U mutation is associated with neurodegeneration in C57BL/6J mice. Maturation of the Tr20C50U precursor is impaired as the mutation stabilizes a conformational ensemble different from the wild-type. To gain insights into how this mutation engenders structural differences, we used solid-state nanopore sensing for the real-time identification of metastable conformers that are not easily observable by ensemble methods. Ion-current traces recorded using an 8-nm nanopore revealed broad contours of the conformational landscape of n-Tr20/n-Tr20C50U {+/-} Mg2+. Additionally, cryo-EM analysis and small-angle X-ray scattering studies revealed structural plasticity even more than predicted from the nanopore-sensing data. Since dynamics undergird RNA (dys)function in cellular physiology and pathology, nanopore sensing to determine RNA conformational sampling is a valuable addition to the growing RNA structural analysis toolkit. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=79 SRC="FIGDIR/small/647894v2_ufig1.gif" ALT="Figure 1"> View larger version (22K): org.highwire.dtl.DTLVardef@45bforg.highwire.dtl.DTLVardef@1e31110org.highwire.dtl.DTLVardef@75388forg.highwire.dtl.DTLVardef@1d45037_HPS_FORMAT_FIGEXP M_FIG C_FIG

biophysics↗

bioRxiv2023

Mapping the MOB proteins' proximity network reveals a unique interaction between human MOB3C and the RNase P complex

Distinct functions mediated by members of the monopolar spindle-one-binder (MOB) family of proteins remain elusive beyond the evolutionary conserved and well-established roles of MOB1A and B in regulating the Hippo pathway. Since MOB proteins are adaptors, understanding how they engage in protein-protein interactions and complexes assembly is essential to define the full scope of their biological functions. To address this, we undertook a proximity-dependent biotin identification (BioID) approach to define the interactomes of all seven human MOB proteins in HeLa and HEK293 cell lines. We uncovered > 200 interactions, of which at least 70% are unreported on BioGrid. The generated dataset reliably recalled the bona fide interactors of the well-studied MOBs. We further defined the common and differential interactome between different MOBs on a subfamily and an individual level. We discovered a unique interaction between MOB3C and 7 out of 10 protein subunits of the RNase P complex, an endonuclease that catalyzes tRNA 5 maturation. As a proof-of-principle for the robustness of the generated dataset, we validated the specific interaction of MOB3C with catalytically active RNase P by using affinity purification-mass spectrometry and pre-tRNA cleavage assays of MOB3C pulldowns. In summary, our data provide novel insights into the biology of MOB proteins and reveal the first interactors of MOB3C, components of the RNase P complex, and hence an exciting nexus with RNA biology.

biochemistry↗

bioRxiv2022

RNAs undergo phase transitions with lower critical solution temperatures

Co-phase separation of RNAs and RNA-binding proteins is thought to drive the biogenesis of ribonucleoprotein granules. RNAs can also undergo phase transitions in the absence of proteins. However, the physicochemical driving forces of protein-free, RNA-driven phase transitions remain unclear. Here, we report that RNAs of various types undergo phase transitions with system-specific lower critical solution temperatures (LCSTs). This entropically-driven phase behavior requires Mg2+ ions and is an intrinsic feature of the phosphate backbone that is modulated by RNA bases. RNA-only condensates can additionally undergo enthalpically favorable percolation transitions within dense phases. This is enabled by a combination of Mg2+-dependent bridging interactions among phosphate groups and RNA base-stacking / base-pairing. Phase separation coupled to percolation can cause dynamical arrest of RNAs within condensates and can suppress the catalytic activity of an RNase P ribozyme. Our work highlights the need to incorporate RNA-driven phase transitions into models for RNP granule biogenesis.

biophysics↗