Biology subjects

Wu, L. F.

Publications and source records attributed to Wu, L. F..

8 recordsLinked to original sources

bioRxiv2026

Branched-chain amino acid fermentation as an alternative mammalian electron sink

Hypoxia disrupts mitochondrial respiration and increases the NADH/NAD+ ratio, causing reductive stress. To maintain redox homeostasis, mammalian cells divert electrons toward fermentation. While fermentation in mammals typically involves lactate production, we identify the fermentation of branched-chain amino acids (BCAAs) as an alternative electron sink activated by hypoxia. The resulting metabolites are excreted in urine as a distinct mechanism for alleviating reductive stress. BCAA fermentation is catalyzed by lactate dehydrogenase (LDH) enzymes and is highly responsive to the NADH/NAD+ ratio. Consequently, BCAA fermentation products are sensitive biomarkers for reductive stress in human contexts ranging from resistance exercise to severe hypoxemia. Furthermore, we find that mouse sperm have evolved highly efficient BCAA fermentation, providing a specific metabolic strategy to support the anaerobic electron flow that facilitates flagellar hypermotility across mammalian sperm. Our work highlights an under-appreciated fate of BCAAs in response to reductive stress.

cell biology↗

bioRxiv2023

Small Molecule in situ Resin Capture - an Organism Independent Strategy for Natural Product Discovery.

Microbial natural products remain an important resource for drug discovery. Yet, commonly employed discovery techniques are plagued by the rediscovery of known compounds, the relatively few microbes that can be cultured, and laboratory growth conditions that do not elicit biosynthetic gene expression among myriad other challenges. Here we introduce a culture independent approach to natural product discovery that we call the Small Molecule In situ Resin Capture (SMIRC) technique. SMIRC exploits in situ environmental conditions to elicit compound production and represents a new approach to access poorly explored chemical space by capturing natural products directly from the environments in which they are produced. In contrast to traditional methods, this compound-first approach can capture structurally complex small molecules across all domains of life in a single deployment while relying on Nature to provide the complex and poorly understood environmental cues needed to elicit biosynthetic gene expression. We illustrate the effectiveness of SMIRC in marine habitats with the discovery of numerous new compounds and demonstrate that sufficient compound yields can be obtained for NMR-based structure assignment. Two new compound classes are reported including one novel carbon skeleton that possesses a functional group not previously observed among natural products and a second that possesses potent biological activity. We introduce expanded deployments, in situ cultivation, and metagenomics as methods to facilitate compound discovery, enhance yields, and link compounds to producing organisms. This compound first approach can provide unprecedented access to new natural product chemotypes with broad implications for drug discovery. Significance StatementPharmaceutically relevant microbial natural products have traditionally been discovered using a microbe-first approach in which bioassays are used to guide the isolation of active compounds from crude culture extracts. While once productive, it is now widely recognized that this approach fails to access the vast chemical space predicted from microbial genomes. Here, we report a new approach to natural product discovery in which compounds are captured directly from the environments in which they are produced. We demonstrate the applications of this technique with the isolation and identification of both known and new compounds including several that possess new carbon skeletons and one with promising biological activity.

biochemistry↗

bioRxiv2022

Dissecting the effects of GTPase and kinase domain mutations on LRRK2 endosomal localization and activity

Parkinsons disease-causing LRRK2 mutations lead to varying degrees of Rab GTPase hyperphosphorylation. Puzzlingly, LRRK2 GTPase-inactivating mutations--which do not affect intrinsic kinase activity--lead to higher levels of cellular Rab phosphorylation than kinase-activating mutations. Here, we investigated whether mutation-dependent differences in LRRK2 cellular localization could explain this discrepancy. We discovered that blocking endosomal maturation leads to the rapid formation of mutant LRRK2+ endosomes on which LRRK2 phosphorylates substrate Rabs. LRRK2+ endosomes are maintained through positive feedback, which mutually reinforces membrane localization of LRRK2 and phosphorylated Rab substrates. Furthermore, across a panel of mutants, cells expressing GTPase-inactivating mutants formed strikingly more LRRK2+ endosomes than cells expressing kinase-activating mutants, resulting in higher total cellular levels of phosphorylated Rabs. Our study suggests that an increased probability of LRRK2 GTPase-inactivating mutants to be retained on intracellular membranes over the kinase-activating mutants leads to higher substrate phosphorylation.

systems biology↗

bioRxiv2022

Commitment in the Pink1/Parkin mitophagy decision circuit

Mechanisms that prevent accidental degradation of healthy mitochondria by the PINK1/Parkin mitophagy pathway are poorly understood. On the surface of damaged mitochondria, PINK1 accumulates and acts as the input signal to a positive feedback loop of Parkin recruitment, which in turn promotes mitochondrial degradation via mitophagy. However, PINK1 is also present on healthy mitochondria where it could errantly recruit Parkin and thereby activate this positive feedback loop. Here, we quantitatively mapped the relationship between PINK1 input levels and Parkin recruitment dynamics using live-cell microscopy and mathematical modeling. We found that Parkin is recruited to the mitochondria only if PINK1 levels exceed a threshold and only after a delay that is inversely proportional to PINK1 levels. The threshold and delay provide a "two-factor authentication" step for PINK1/Parkin activation. These properties arise from the PINK1/Parkin circuit topology and provide a mechanism for cells to assess damage signals before committing to mitophagy.

systems biology↗

bioRxiv2020

On the role of photoreceptor identity in controlling accurate wiring of the Drosophila visual circuit

During development, neurons extend in search of synaptic partners. Precise control of axon extension velocity can therefore be crucial to ensuring proper circuit formation. How velocity is regulated - particularly by the extending axons themselves - remains poorly understood. Here, we investigate this question in the Drosophila visual system, where photoreceptors make precise connections with a specific set of synaptic partners that together create a circuit underpinning neural superposition (NSP). We used a combination of genetic perturbations and quantitative image analysis to investigate the influence of cell identity on growth cone velocity and subsequent spatial-temporal coincidence of presynaptic and postsynaptic neurons. Our study provides a case study of how cell autonomous properties of presynaptic axons play a pivotal role in controlling the dynamics of growing axons and determining the formation of a precise neuronal circuit.

bioinformatics↗

bioRxiv2019

Non-stem progenitors enable coordinated changes in gut epithelial cell-type composition

Renewing tissues have the remarkable ability to continually produce both proliferative progenitor and specialized differentiated cell-types. How are complex milieus of microenvironmental signals interpreted to coordinate tissue cell-type composition? Here, we develop a high-throughput approach that combines organoid technology and quantitative imaging to address this question in the context of the intestinal epithelium. Using this approach, we comprehensively survey enteroid responses to individual and paired perturbations to eight epithelial signaling pathways. We uncover culture conditions that enrich for specific cell-types, including Lgr5+ stem and enteroendocrine cells. We analyze interactions between perturbations and dissect mechanisms underlying an unexpected mutual antagonism between EGFR and IL-4 signals. Finally, we show that, across diverse perturbations, modulating proliferation of transit-amplifying cells also consistently changes the composition of differentiated secretory and absorptive cell-types. This property is conserved in vivo and can arise from differential amplification of secretory and absorptive progenitor cells. Taken together, the observations highlight an underappreciated role for transit-amplifying cells in which proliferation of these short-lived progenitors provides a lineage-based mechanism for tuning differentiated cell-type composition.

cell biology↗

bioRxiv2019

Inhibitory effect of Bacillus subtilis WL-2 and its IturinA lipopeptides against Phytophthora infestans

Potato late blight triggered by Phytophthora infestans ((Mont.) de Bary) represents a great food security threat worldwide and is difficult to control. Currently, Bacillus spp. have been considered biocontrol agents to control many fungal diseases. Here, Bacillus subtilis WL-2 was selected as the antifungal strain with the most potential against P. infestans mycelium growth. Additionally, the functional metabolites extracted from WL-2 were identified as IturinA-family cyclic lipopeptides (CLPs) via high-performance liquid chromatography (HPLC) and electrospray ionization mass spectrometry (ESI-MS). Analyses using scanning and transmission electron microscopy (SEM and TEM) revealed that IturinA caused a change in the mycelial surface and damage to the internal cell structure, including cell membrane disruption and irregular organelle formation. Moreover, propidium iodide staining and nucleic acid and protein release were detected to clarify the cell membrane damage caused by IturinA. Additionally, IturinA triggered reactive oxygen species (ROS) generation and malondialdehyde (MDA) production. Mitochondrial membrane potential (MMP), mitochondrial respiratory chain complexes activity (MRCCA), respiratory control rate (RCR), and oxidative phosphorylation efficiency (P/O) assays indicated that P. infestans mitochondria affected by IturinA were so seriously damaged that the MMP and MRCCA declined remarkably and that mitochondrial ATP production ability was weakened. Therefore, IturinA induces cell membrane damage, oxidative stress, and dysfunction of mitochondria, resulting in P. infestans hyphal cell death. As such, the results highlight that B. subtilis WL-2 and IturinA have great potential as candidates for inhibiting P. infestans mycelium growth and controlling potato late blight.\n\nIMPORTANCEPotato (Solanum tuberosum L.) is the fourth most common global food crop, and its planting area and yield increase yearly. Notably, in 2015, China initiated a potato staple food conversion strategy, and by 2020, approximately 50% of potatoes will be consumed as a staple food. The plant pathogen fungus Phytophthora infestans ((Mont.) de Bary) is the culprit of potato late blight; however, biological agents rather than chemicals are highly necessary to control this threatening disease. In this study, we discovered an antifungal substance, IturinA, a lipopeptide produced by Bacillus subtilis WL-2. Moreover, our research revealed the actual mechanism of IturinA against P. infestans mycelium growth and clarified the potential of B. subtilis WL-2 and IturinA as a biocontrol agent against P. infestans mycelium growth as well as for controlling the development of late blight in potato cultivation.

microbiology↗

bioRxiv2017

Response-Time Behaviors Of Intercellular Communication Network Motifs

Cell-to-cell communication networks have critical roles in diverse organismal processes, such as coordinating tissue development or immune cell response. However, compared to intracellular signal transduction networks, the function and engineering principles of cell-to-cell communication networks are far less understood. Here, we study cell-to-cell communication networks using a framework that models the input-to-output relationship of intracellular signal transduction networks with a single function--the response-time distribution. We identify a prototypic response-time distribution--the gamma distribution--arising in both experimental data sets and mathematical models of signal-transduction pathways. We find that simple cell-to-cell communication circuits can generate bimodal response-time distributions, and can control synchronization and delay of cell-population responses independently. We apply our modeling approach to explain otherwise puzzling data on cytokine secretion onset times in T cells. Our approach can be used to predict communication network structure using experimentally accessible input-to-output measurements and without detailed knowledge of intermediate steps.

biophysics↗