Biology subjects

Kim, N. H.

Publications and source records attributed to Kim, N. H..

7 recordsLinked to original sources

bioRxiv2023

Bimodality in Ras signaling originates from processivity of the Ras activator SOS without classic kinetic bistability

Ras is a small GTPase that is central to important functional decisions in diverse cell types. An important aspect of Ras signaling is its ability to exhibit bimodal, or switch-like activity. We describe the total reconstitution of a receptor-mediated Ras activation-deactivation reaction catalyzed by SOS and p120-RasGAP on supported lipid membrane microarrays. The results reveal a bimodal Ras activation response, which is not a result of classic kinetic bistability, but is rather driven by the distinct processivity of the Ras activator, SOS. Furthermore, the bimodal response is controlled by the condensation state of the scaffold protein, LAT, to which SOS is recruited. Processivity-driven bimodality leads to stochastic bursts of Ras activation even under strongly deactivating conditions. This behavior contrasts classic kinetic bistability and is distinctly more resistant to pharmacological inhibition.

biophysics↗

bioRxiv2023

Plasma membrane association and resistosome formation of plant helper immune receptors

Intracellular plant immune receptors, termed NLRs, respond to pathogen effectors delivered into plant cells. Activation of NLRs typically confers immunity. Sensor NLRs, involved in effector recognition, are either TIR-NLRs (TNLs) or CC-NLRs (CNLs). Helper NLRs, required for sensor NLR signaling, include CCR-NLRs (RNLs) and a special class of CNLs known as NRCs. Activated TNLs produce small molecules that induce an association between the EDS1/SAG101 heterodimer and the NRG1s helper RNLs. Auto active NRG1s oligomerize and form calcium signaling channels largely localized at the plasma membrane (PM). The molecular mechanisms of helper NLR PM association and effector induced NRG1 oligomerization are not well characterized. We find that both RNLs and NRCs require positively charged residues in the second and fourth helices of their CCR or CC domain for phospholipid binding and PM association before and after activation, despite conformational changes that accompany activation. We demonstrate that effector activation of TNLs induces NRG1 oligomerization at the PM and that the cytoplasmic pool of EDS1/SAG101 is critical for cell death function. EDS1/SAG101 cannot be detected in the oligomerized NRG1 resistosome, suggesting that additional unknown triggers might be required to induce the dissociation of EDS1/SAG101 from the previously described NRG1/EDS1/SAG101 heterotrimer before subsequent NRG1 oligomerization, or that the conformational changes resulting from NRG1 oligomerization abrogate the interface for EDS1/SAG101 association. Our data provide new observations regarding dynamic PM association during helper NLR activation and underpin an updated model for effector induced NRG1 resistosome formation.

plant biology↗

bioRxiv2022

The metabolite alpha-ketobutyrate increases health and life spans by activating AMPK

Aging is a complex process that is directly related to human health and disease. The extraordinary finding that aging is malleable, as shown in model organisms whose life and health spans are extended by specific gene mutations or dietary or pharmacological perturbations 1-3, has offered enormous hope for our understanding and treatment of aging and related diseases. Although many molecules have been identified that can extend the lifespan of model organisms, few have been shown to alleviate age-related symptoms or illness in mammals 4. Here we show that supplementation with the endogenous metabolite -ketobutyrate (-KB) increases the lifespan of adult C. elegans. Using Gelfree DARTS-PROTOMAP, we identified microtubule-actin cross-linking factor (MACF1) that was protected against proteolysis in the presence of -KB. MACF1 belongs to the spectraplakin family of giant, evolutionarily conserved proteins with versatile functions 5, but their link to longevity regulation has not been explored. -KBs longevity effect in C. elegans is abrogated by loss-of-function mutation in vab-10, encoding the worm ortholog of mammalian MACF1 6. Like -KB treatment, vab-10 knockdown activates AMP-activated protein kinase (AMPK), and AMPK is required for -KB effects on longevity. The findings suggest a model in which -KB increases longevity by activating AMPK via VAB-10/MACF1 modulation. -KB also delays aging in mammals, increasing the lifespan of aged male mice and the healthspan of both male and female animals. Targeting of broadly expressed scaffolding proteins in connection to cellular energy homeostasis seems to be a clever way that nature has devised for metabolite signals to impinge upon multiple organ and tissue systems, which may have utility for controlling aging and related diseases.

physiology↗

bioRxiv2022

Allelic variation in the Arabidopsis TNL CHS3/CSA1 immune receptor pair reveals two functional regulatory modes

Some plant NLR immune receptors are encoded in head-to-head pairs that function together. Alleles of the NLR pair CHS3/CSA1 form three clades. The clade 1 sensor CHS3 contains an integrated domain (ID) with homology to regulatory domains, which is lacking in clades 2 and 3. We defined two regulatory modes for CHS3/CSA1 pairs. One is likely mediated by effector binding to the clade 1 ID of CHS3 and the other relies on CHS3/CSA1 pairs from all clades detecting effector modification of an associated pattern recognition receptor. We suggest that an ancestral Arabidopsis CHS3/CSA1 pair gained a second recognition specificity and regulatory mechanism through ID acquisition, while retaining its original specificity as a Guard against perturbation of pattern recognition receptor targeting by a pathogen effector. This likely comes with a cost, since both ID and non-ID alleles of the pair persist in diverse Arabidopsis populations through balancing selection. SummaryWe dissect a novel case where two regulatory modes emerged across three clades of the co-evolved CHS3/CSA1 plant immune receptor pairs, which features recruitment of an integrated domain (ID) into the clade 1 CHS3 alleles. Pre- and post-ID integration alleles maintain functionality; balancing selection maintains both in the Arabidopsis pan-genome.

plant biology↗

bioRxiv2022

Emergence of glycogen synthase kinase-3 interaction domain enhances phosphorylation of SARS-CoV-2 nucleocapsid protein

A structural protein of SARS-CoV-2, nucleocapsid (N) protein is abundantly expressed during viral replication. The N protein is phosphorylated by glycogen synthase kinase (GSK)-3 on the serine/arginine (SR) rich motif located in disordered regions. Although phosphorylation by GSK-3{beta} constitutes a critical event for viral replication, the molecular mechanism underlying N phosphorylation is not well understood. In this study, we found the putative alpha-helix L/FxxxL/AxxRL motif known as the GSK-3 interacting domain (GID), commonly found in many endogenous GSK-3{beta} binding proteins, such as Axins, FRATs, WWOX and GSKIP. Indeed, N interacts with GSK-3{beta} similarly to Axin, and Leu to Glu substitution of the GID abolished the interaction, with loss of N phosphorylation. Unlike with endogenous GID proteins, the N interaction neither disturbs endogenous GSK-3 activity nor regulates subsequent canonical Wnt activity and the Snail-EMT program. Notably, N abundance in SARS-CoV-2 is incomparably high compared to other coronaviruses, such as 229E, OC43 and HKU1. Compared to other coronaviruses, N harbors a CDK1 primed phosphorylation site and Gly-rich linker for enhanced phosphorylation by GSK-3{beta}. Furthermore, we found that the S202R mutant found in Delta and R203K/G204R mutant found in the Omicron variant allows increased abundance and hyper-phosphorylation of N. Our observations suggest that the emergence of GID and mutations for increased phosphorylation in N may have contributed to the emergence of SARS-CoV-2 and evolution of variants, respectively. Further study, especially in a BSL3-equipped facility, is required to elucidate the functional importance of GID and N phosphorylation in SARS-CoV-2 and variants.

cell biology↗

bioRxiv2021

The plant immune receptors NRG1.1 and ADR1 are calcium influx channels

Plant nucleotide-binding leucine-rich repeat receptors (NLRs) regulate immunity and cell death. RPW8 domain-containing "helper" NLRs (RNLs) are required by many "sensor" NLRs. Our crystal structure of the RNL N REQUIREMENT GENE 1.1 (NRG1.1) N-terminal signaling domain resembled that of the resting state plant resistosome-forming HOPZ-ACTIVATED RESISTANCE 1 (ZAR1) and the animal MIXED-LINEAGE KINASE-LIKE (MLKL) cation channel. Active NRG1.1 oligomerized, was enriched in plasma membrane puncta and conferred cytoplasmic Ca2+ influx in plant and human HeLa cells. NRG1.1-dependent Ca2+ influx and cell death were sensitive to Ca2+ channel blockers. Ca2+ influx and cell death mediated by NRG1.1 and ACTIVATED DISEASE RESISTANCE 1 (ADR1), another RNL, required conserved negatively charged N-terminal residues. Thus, RNLs apparently form influx channels to directly regulate cytoplasmic [Ca2+] and consequent cell death. One Sentence SummaryA specific class of plant immune receptors function as calcium-permeable channels upon activation to induce cell death.

plant biology↗

bioRxiv2020

Size-dependent protein segregation creates a spatial switch for Notch and APP signaling

Aberrant cleavage of Notch by {gamma}-secretase is implicated in numerous diseases, but how cleavage is regulated in space and time is unclear. Here, we report that cadherin-based adherens junctions (cadAJs) are sites of high cell-surface {gamma}-secretase activity, as well as sites of constrained physical space that excludes {gamma}-secretase substrates having large extracellular domains (ECDs) like Notch. ECD shedding initiates drastic spatial relocalization of Notch to cadAJs, allowing enzyme-substrate interactions and downstream signaling. Spatial mutations by adjusting the ECD size or the physical constraint alter signaling. Dysregulation of this spatial switch promotes precocious differentiation of ventricular zone neural progenitor cells in vivo. We show the generality of this spatial switch for amyloid precursor protein proteolysis. Thus, cadAJs create spatially distinct biochemical compartments regulating cleavage events involving {gamma}-secretase and preventing aberrant activation of receptors. One Sentence SummaryNotch cleavage by {gamma}-secretase is regulated through dynamic spatial control of receptors, adhesion molecules, and activating proteases

cell biology↗