Ingrid Robl

Phosphatidyl ethanolamine is essential for targeting the arginine transporter Can1p to the plasma membrane of yeast.

In continuation of our previous study, we show that phosphatidyl ethanolamine (PE) depletion affects, in addition to amino acid transporters, activities of at least two other proton motive force (pmf)-driven transporters (Ura4p and Mal6p). For Can1p, we demonstrate that the lack of PE results in a failure of the permease targeting to plasma membrane. Despite the pleiotropic effect of PE depletion, a specific role of PE in secretion of a defined group of permeases can be distinguished. Pmf-driven transporters are more sensitive to the lack of PE than other plasma membrane proteins.

Construction of phosphatidylethanolamine‐less strain of Saccharomyces cerevisiae. Effect on amino acid transport

A triple yeast mutant was constructed which lacks BST1, the gene for sphingosine lyase, besides the phosphatidylserine decarboxylases PSD1 and PSD2. In this yeast mutant, which can only be grown in the presence of exogenous ethanolamine, phosphatidylethanolamine can be depleted to very low levels. Under those conditions, respiration as well as glucose and 3‐O‐methylglucose uptake proceed unaffected. Plasma membrane ATPase is as active in these cells as that of control cells grown in the presence of ethanolamine. Drastically decreased, however, are H+/amino acid symporters. The activities of arginine (Can1p), proline (Put4p) and general amino acid permease (Gap1p) are decreased more than 20‐fold. Amino acid transport in yeast is dependent on coupling to the proton motive force. It can be envisaged that phosphatidylethanolamine might play a role in this process or in the early steps of the secretion pathway common for all amino acid permeases or, eventually, it could affect the transport proteins directly at the plasma membrane Transformation of the triple mutant with a CEN plasmid harbouring BST1 wild‐type gene totally reversed its phenotype to that observed in the double mutant. Copyright

The C-terminal tetrapeptide HWFW of the Chlorella HUP1 hexose/H+-symporter is essential for full activity and an α-helical structure of the C-terminus

C‐terminal tails of plant hexose/H+‐symporters of the major facilitator superfamily contain a highly conserved motif of four amino acids: HWFW. A deletion of these four amino acids in the Chlorella HUP1 protein leads to a decrease in transport activity by a factor of 3–4. The mutated tail is highly sensitive to trypsin; it does not show α‐helical conformation in contrast to the wild type C‐terminal peptide with an α‐helical content of at least 15%. The production of monoclonal antibody 416B8 recognizing an epitope within the central loop of HUP1 protein has been a prerequisite for the experiments described.