Mechanisms Implicated in the Response of System A to Hypertonic Stress and Amino Acid Deprivation Still Can Be Different

Alfieri et al. (2004) analyzed the response of system A to hypertonic stress by determining changes in SNAT2 mRNA levels in a panel of cell lines, including the same cell type, CHO-K1, that we routinely used in the past for this kind of studies. It is evident that a discrepancy between their results and ours regarding the effect of hypertonicity on SAT2/SNAT2 mRNA levels exists. Our CHO-K1 clone was originally obtained from E. who developed a panel of somatic mutant cell lines with altered system A activity and regulatory prop-among others). In our experiments, the SAT2/ SNAT2 cDNA probe used for Northern blot analysis was a fragment cloned from that particular cell line (this is the hamster SNAT2 orthologue, accession no. of the partial sequence AF363584). Although we do not believe these slight differences might explain this discrepancy, the study we published in the Journal of General Physiology (López-Fontanals et al., 2003) does not particularly rely upon this observation to conclude that the osmoregulatory and the amino acid–regulated responses of system A are mediated by different signal transduction pathways. In that study, we combined inhibitors of the MAP kinase pathway, as well as negative dominant cells for selected kinases in this transduction pathway, and modulators of the cell cycle machinery, to demonstrate that, at least in CHO-K1 cells, both stimuli trigger independent responses. All these experiments were performed by looking at system A functional activity, which is presumably associated with SNAT2 expression in CHO-K1 cells. This is in agreement with our previous work (Ruiz-Montasell et al., 1994) in which we used a particular somatic cell CHO-K1 mutant (CHO-K1 ala r 4), generated at Ellis Englesberg's laboratory (Moffett and Englesberg, 1984, 1986), that lacked the ability to respond to amino acid starvation but, interestingly, still retained the hyperosmotic response. Although we agree that the existence of the system A activating protein is still an open question, as Alfieri et al. discuss, the conclusion that both pathways must converge at some point cannot be drawn from the mere observation that the two stimuli induce an increase in SAT2/SNAT2 mRNA levels. We respectfully believe that this is a simple interpretation of these data. Most genes can be transcriptionally activated by different agents/stimuli without bearing any common step in their transduction pathways, except that they converge, obviously, at the end somewhere on the gene promoter. Nevertheless, as the authors point out, even for the …