Grobner, G. | Search bioRxiv

bioRxiv2024

Metabolic readouts of tumor instructed normal tissues (TINT) identify aggressive prostate cancer subgroups for tailored therapy

BackgroundProstate cancer (PC) diagnosis relies on histopathological examination of prostate biopsies, which is restricted by insufficient sampling of all tumors present. Including samples from non-PC but tumor instructed normal tissues (TINT) may increase the diagnostic power by exploring the adaptive responses in benign tissues near tumors. MethodsHere, we applied high-resolution magic angle spinning nuclear magnetic resonance (HR MAS NMR) to identify metabolomic biomarkers with high diagnostic value in benign prostate tissues near low/high-grade tumors. ResultsBenign samples near high-grade tumors (B ISUP 3+4) exhibit altered metabolic profiles compared to those close to low-grade tumors (B ISUP 1+2). The levels of six metabolites were significantly different between the two groups; myo-inositol, lysine, serine and combined signal of lysine/leucine/arginine were increased in benign samples near high-grade tumors (B ISUP 3+4) compared to near low-grade tumors (B ISUP 1+2), while levels of ethanolamine and lactate decreased. Additionally, we revealed metabolic differences in non-cancer tissues as a function of their distance to the nearest tumor. Eight metabolites (glutathione, glutamate, combined signal of glutamate/glutamine - glx, glycerol, inosine, ethanolamine, serine and arginine) significantly differentiated between benign tissue located close to the tumor (d [≤] 5 mm) compared to those far away (d [≥] 1 cm). ConclusionsOur HR MAS NMR-based approach identified metabolic signatures in prostate biopsies that reflect the response of benign tissues to the presence of nearby located tumors in the same prostate and confirmed the power of the TINT concept for improved PC diagnostics and understanding of tumor-tissue interactions.

cancer biology↗

bioRxiv2022

Bax Forms a Membrane Surface Protein-Lipid Complex as it Initiates Apoptosis

Cellular clearance by apoptosis is essential in life. In its intrinsic (mitochondrial) pathway apoptotic members of the Bcl-2 (B cell CLL/lymphoma-2) protein family, such as Bax (Bcl-2-associated X) protein, perforate the mitochondrial outer membrane (MOM), which causes release of apoptotic factors and final cell death. How those apoptotic proteins mechanistically exert their action at the membrane level still however remains elusive. Upon internal stress signals Bax is massively recruited to the MOM, where it oligomerizes and partially penetrates into the membrane. Using neutron reflectometry (NR) and attenuated total reflection Fourier-Transform infrared spectroscopy (ATR-FTIR) spectroscopy we unraveled key molecular steps of this membrane-affiliation process of Bax on a spatial and temporal scale. By titrating intact human Bax to MOM-like bilayers containing cardiolipin, essential for protein recruitment, we could identify different functional phases. Initially, there is a fast adsorption event to the membrane surface with high affinity. Thereafter, a kinetically slower (minutes to hours) event occurs with Bax penetration, thereby triggering a major reorganizing of the mitochondrial bilayer. Finally, a membrane-Bax complex is generated, with a minor Bax population remaining membrane-inserted, while the main population is relocated to the membrane surface upon lipid redistribution into a complex with Bax; a process enabling membrane perforation. We propose a comprehensive molecular model of mitochondrial membrane penetration by formation of complex Bax/lipid clusters; a concept which provides a new foundation to understand the cell-killing activity of Bax and its apoptotic relatives in human cells. Significance StatementThe apoptotic Bax protein is a key player in the mitochondrial apoptotic pathway. Here, neutron reflectometry (NR) unravels the mechanism by which Bax is targeting and perforating mitochondria to release apoptotic factors for final cell death. We found that this cardiolipin driven process of the outer mitochondrial membrane system has two main phases. Upon a fast (10-20 min) phase of membrane association Bax initiates the formation of pores by removing lipids and depositing them as Bax/lipid complexes on top of the bilayer on a time scale of several hours similar to in vivo apoptotic cell death. Our results provide a mechanistic rationale for cell-killing processes driven by apoptotic Bcl-2 proteins; and their molecular inhibition in many cancers.

biophysics↗