Jorge, A. C. | bioRxiv Science

bioRxiv2026

Low-nanogram Fourier Transform Isotopic Ratio Mass Spectrometry of Proteins

Stable carbon and nitrogen isotope ratios are widely used in the life sciences to investigate diet, trophic interactions, and metabolic fluxes, but conventional isotope ratio mass spectrometry requires milligram-scale samples, limiting its applicability to small or rare biological specimens. Fourier Transform Isotopic Ratio Mass Spectrometry (FT IsoR MS) enables amino acid-resolved isotope analysis in a proteomics-compatible workflow and has previously been demonstrated at the microgram scale. Here, we assess the lower sample limit of FT IsoR MS by integrating it with single-cell proteomics-style sample preparation. Using human HeLa cells cultured in 13C-glucose-enriched and control media, we show that reliable relative {delta}13C measurements can be obtained from as few as 50 cells, corresponding to <10 ng of total protein, with a precision of approximately {+/-}9{per thousand}. The observed amino acid-specific labeling patterns are metabolically coherent and consistent with bulk measurements, while smaller cell numbers ([≤]10 cells) do not yield statistically robust results. These findings establish the practical sensitivity threshold of FT IsoR MS at the low-nanogram level and demonstrate its suitability for isotope-resolved analyses of small cell populations, micro-organoids, and other low-input biological samples, thereby extending stable isotope analysis toward single-cell-scale applications.

biochemistry↗

bioRxiv2025

Sweet taste receptor regulates proliferation and glucose uptake in glioblastoma cells

Glioblastoma is a brain tumour classified by the World Health Organization as grade 4 due to its aggressiveness, invasiveness, and poor differentiation. Current standard therapies remain largely ineffective, reinforcing the need for novel targets to improve glioblastoma prognosis. Reprogramming of cellular metabolism is an important hallmark of cancer, marked by a shift from oxidative phosphorylation to glycolysis as the primary energy source. This metabolic switch, known as the Warburg effect, is exacerbated by the reduced oxygen and glucose availability in the tumour microenvironment. The sweet taste receptor (STR) is an important glucosensor with recognised role in the regulation of glucose uptake in several organs and in astrocytes, the glioblastoma precursor cells. We hypothesised that STR may regulate glucose uptake and metabolism in glioblastoma, representing a potential anticancer target. To test this, we investigated the effects of STR inhibition with lactisole, a specific TAS1R3 inhibitor, in three glioblastoma cell lines (U-87MG, SNB-19 and U-373MG) and in the UPCI-SCC-154 tongue cancer cell line. We provided evidence that STR inhibition consistently reduced cell viability and migration, without inducing apoptosis or necrosis, and impaired glucose uptake and L-lactate production, particularly in tongue cancer UPCI-SCC-154, and glioblastoma U-87MG and SNB-19 cells. Moreover, in a 3D spheroid model, lactisole reduced the invasion capacity of U-87MG glioblastoma spheroids. These results suggest that STR contributes to glioblastoma cell metabolism and behaviour and may represent a promising therapeutic target.

cancer biology↗