Super-resolution Imaging Unlocks The Role Of Transporter Nanoclusters In Tumor Metabolism

Serine serves arsenic a metabolic nexus successful tumors, coordinating one-carbon metabolism, nucleotide synthesis, and redox regulation. In caller years, targeting nan serine metabolic pathway, peculiarly nan rate-limiting enzyme PHGDH, has attracted sizeable attraction arsenic a promising anticancer strategy. Nevertheless, tumor cells sphere serine homeostasis done nan coordinated regularisation of endogenous synthesis and exogenous uptake, which often limits nan efficacy of single-enzyme inhibition approaches. Notably, nan usability of serine transporters (SerTs) is intimately linked to their nanoscale spatial statement connected nan plasma membrane. However, owing to nan deficiency of highly circumstantial probes and ultra-resolution imaging techniques,
the mechanisms underlying their assembly and functional relevance stay mostly unexplored.

Researchers from Wuhan University of Science and Technology and nan Changchun Institute of Applied Chemistry, Chinese Academy of Sciences person jointly published a insubstantial successful Research entitled "Mechanistic Insight into Serine Flux Regulation done Nanoscale Organization of Glucose and Serine Transporters by Substrate Probe-based dSTORM Imaging." The study developed a small-molecule substrate-based fluorescent probe (Ser-probe) derived from nan building of serine. This chemic probe not only exhibits precocious binding specificity and a one-to-one binding ratio pinch serine transporters (SerTs), but besides possesses an highly mini molecular size. Compared pinch accepted biologic probes specified arsenic antibodies, it offers important advantages successful position of compactness, enabling highly businesslike labeling of densely distributed target proteins connected nan compartment surface. Leveraging nan superior labeling capacity of this probe mixed pinch nonstop stochastic optical reconstruction microscopy (dSTORM), nan researchers achieved super-resolution imaging and quantitative study of aggregate serine transporters (SerTs). The activity systematically revealed nan nanoscale clustering behaviour and regulatory mechanisms of SerTs successful bosom crab cells.
Through precise chemic modification of nan serine molecule, nan investigation squad preserved its transporter-binding moiety and conjugated it pinch a fluorophore, thereby successfully synthesizing Ser-probe. This probe exhibits precocious affinity and specificity. Competitive research demonstrated that its binding tin beryllium wholly inhibited by free serine, which confirms its expertise to specifically explanation SerTs nether physiological conditions. The improvement of this probe provides a caller instrumentality for super-resolution imaging of membrane transporters.
Using dSTORM imaging, it was observed that from normal bosom epithelial cells (MCF10A) to low-malignant MCF7 cells and highly malignant MDA-MB-231 cells, nan constituent density connected nan compartment membrane, mean cluster area, and cluster sum of SerTs each accrued significantly. In MDA-MB-231 cells, nan mean cluster area of SerTs reached doubly that of MCF10A cells, and nan cluster sum accrued by 4.6-fold. Furthermore, travel cytometry study revealed that cells pinch stronger SerT clustering exhibited higher uptake capacity of fluorescently branded serine, indicating a affirmative relationship betwixt nan clustering inclination of these transporters and their carrier function.
By employing dual-color dSTORM imaging, nan study revealed important colocalization of SerTs pinch glucose transporters (GluTs) connected nan plasma membrane, which was peculiarly pronounced successful MCF7 cells pinch precocious PHGDH expression. The colocalization sum reached 4.23%, markedly higher than nan 1.41% observed successful MDA-MB-231 cells. These findings propose that cells pinch beardown endogenous serine synthesis capacity are much for illustration to forming SerT/GluT functional microdomains, thereby coordinately regulating glucose uptake and serine metabolism.
Cholesterol depletion (MβCD treatment) aliases glycosidase digestion (PNGase F treatment) importantly disrupted nan clustered assembly and colocalization of SerTs and GluTs, indicating that lipid raft microdomains and glycoprotein cross-linking networks represent nan beingness ground for maintaining nan stableness of these transporter nanoclusters. This uncovering provides imaginable targets for nan involution of membrane macromolecule assembly.
Glucose deprivation markedly weakened nan clustering capacity of GluTs, reducing nan SerT/GluT colocalization sum from 4.23% to 0.71%. The PHGDH inhibitor CBR-5884 induced enhanced clustering astatine debased concentrations, whereas astatine precocious concentrations its cytotoxic effects led to cluster disassembly. Combined interventions (inhibitor successful operation pinch debased glucose aliases free sialic acid) synergistically disrupted nan macromolecule cluster structures and importantly enhanced nan anticancer effect. Cell viability decreased from 92.55% nether single-drug curen to 87.71% pinch inhibitor successful operation pinch debased glucose, aliases 78.84% pinch inhibitor successful operation pinch free sialic acid.
This study is nan first to uncover a nonstop relationship betwixt nan spatial statement of SerTs and their usability astatine nan nanoscale. It breaks nan accepted paradigm of metabolic research, which has been mostly constricted to enzyme activity and look levels of cardinal proteins, and offers a caller position for investigating nan domiciled of nutrient transporters successful nan regularisation of tumor metabolism.
Developed nan substrate probe applicable to super-resolution imaging of SerTs;
Revealed nan regulatory axis of "transporter nanocluster distribution-metabolic function-drug response";
Established a exemplary of nan synergistic domiciled of lipid raft and glycosylation successful membrane macromolecule assembly.
The clustering shape of SerTs whitethorn service arsenic a biomarker of tumor metabolic state;
Combined targeting of metabolic enzymes and transporter assembly (e.g., PHGDH inhibitors pinch sialic acid) tin heighten therapeutic efficacy;
Provides a caller strategy for processing therapies aimed astatine regulating nan spatial statement of membrane proteins.

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Journal reference:

Jiang, P., et al. (2025) Mechanistic Insight into Serine Flux Regulation done Nanoscale Organization of Glucose and Serine Transporters by Substrate Probe-Based Direct Stochastic Optical Reconstruction Microscopy Imaging. Research. doi.org/10.34133/research.0805.