Sugar beet PMT5a and STP13 carriers suitable for proton-driven plasma membrane sucrose and glucose import in taproots.

Sugar beet (Beta vulgaris) is the major sugar-producing crop in Europe and Northern America, as the taproot stores sucrose at a concentration of around 20%. Genome sequence analysis together with biochemical and electrophysiological approaches led to the identification and characterization of the TST sucrose transporter driving vacuolar sugar accumulation in the taproot. However, the sugar transporters mediating sucrose uptake across the plasma membrane of taproot parenchyma cells remained unknown. As with glucose, sucrose stimulation of taproot parenchyma cells caused inward proton fluxes and plasma membrane depolarization, indicating a sugar/proton symport mechanism. To decipher the nature of the corresponding proton-driven sugar transporters, we performed taproot transcriptomic profiling and identified the cold-induced PMT5a and STP13 transporters. When expressed in Xenopus laevis oocytes, BvPMT5a was characterized as a voltage- and H+-driven low-affinity glucose transporter, which does not transport sucrose. In contrast, BvSTP13 operated as a high-affinity H+/sugar symporter, transporting glucose better than sucrose, and being more cold-tolerant than BvPMT5a. Modeling of the BvSTP13 structure with bound mono- and disaccharides suggests plasticity of the binding cleft to accommodate the different saccharides. The identification of BvPMT5a and BvSTP13 as taproot sugar transporters could improve breeding of sugar beet to provide a sustainable energy crop.

Unlocking roles of cationic and aromatic residues in peptide amphiphiles in treating drug-resistant gram-positive pathogens.

Multidrug resistance (MDR) is a rising threat to global health because the number of essential antibiotics used for treating MDR infections is increasingly compromised. In this work we report a group of new amphiphilic peptides (AMPs) derived from the well-studied G3 (G(IIKK)3I-NH2) to fight infections from Gram-positive bacteria including susceptible Staphylococcus aureus and methicillin-resistant Staphylococcus aureus (MRSA), focusing on membrane interactions. Time-dependent killing experiments revealed that substitutions of II by WW (GWK), II by FF (GFK) and KK by RR (GIR) resulted in improved bactericidal efficiencies compared to G3 (GIK) on both S. aureus and MRSA, with the order of GWK > GIR > GFK > GIK. Electronic microscopy imaging revealed structural disruptions of AMP binding to bacterial cell walls. Fluorescence assays including AMP binding to anionic lipoteichoic acids (LTA) in cell-free and cell systems indicated concentration and time-dependent membrane destabilization associated with bacterial killing. Furthermore, AMP's binding to anionic plasma membrane via similar fluorescence assays revealed a different extent of membrane depolarization and leakage. These observations were supported by the penetration of AMPs into the LTA barrier and the subsequent structural compromise to the cytoplasmic membrane as revealed from SANS (small angle neutron scattering). Both experiments and molecular dynamics (MD) simulations revealed that GWK and GIR could make the membrane more rigid but less effective in diffusive efficiency than GIK and GFK through forming intramembrane peptide nanoaggregates associated with hydrophobic mismatch and formation of fluidic and rigid patches. The reported peptide-aggregate-induced phase-separation emerged as a crucial factor in accelerated membrane disintegration and fast bacterial killing. This work has demonstrated the importance of membrane interactions to the development of more effective AMPs and the relevance of the approaches as reported in assisting this area of research.