Determining Pathway Bandwidth

Science Signaling  27 May 2008:
Vol. 1, Issue 21, pp. ec193
DOI: 10.1126/stke.121ec193

Signaling pathways allow cells and organisms to respond to changes in the environment. But how quickly a signaling pathway can be activated and deactivated dictates the pathway’s bandwidth--the maximum frequency with which a cell can faithfully reproduce changes in the stimulus. Hersen et al. developed a microfluidic device to trigger periodic changes in culture medium and then used it to rapidly and periodically increase osmolarity while monitoring, using fluorescent reporters, the activation of the high-osmolarity glycerol (HOG) pathway in Saccharomyces cerevisiae. The HOG pathway consists of a mitogen-activated protein kinase (MAPK) cascade, which can be activated by either a two-component phosphorelay (the Sln1 pathway from Sln1 to Ssk1 to the MAPKKKs Ssk2 and Ssk22 to the MAPK Pbs2 to the MAPK Hog1) or an enzymatic sensor system (the Sho1 pathway from Sho1 to the MAPKKK Ste11 to Pbs2 to Hog1). The authors monitored nuclear translocation of a green fluorescent protein (GFP)-tagged Hog1 or production of Gpd1-GFP (encoded by a Hog1 target gene) and calculated a pathway bandwidth of 0.0046 Hz. When the osmolarity was changed at a frequency less than the pathway bandwidth, Hog1-GFP translocation and Gpd1-GFP production increased and decreased in sync with the input stimulus. When the osmolarity was changed more rapidly than the pathway bandwidth, the pathway integrated the input and Hog1-GFP accumulated in the nucleus without oscillating and then reached a plateau. Gpd1-GFP also accumulated, reaching a plateau, without oscillating when the frequency of the change in osmolarity exceeded the pathway bandwidth. This behavior is consistent with the pathway serving as a low-pass filter. The bandwidth calculated for cell swelling was lower than the pathway bandwidth, which suggests that the mechanical change in cell shape was not the limiting factor. Instead, pathway bandwidth was limited by the activation and deactivation rates of the biochemical reactions that mediate the HOG response. Only mutants of the Sln1 pathway, but not the Sho1 pathway, had a bandwidth less than the bandwidth of the pathway in wild-type cells; thus, this input into the HOG MAPK cascade sets the frequency with which the pathway can respond. Furthermore, only the Sln1 pathway was required for integration of high-frequency stimulation that exceeded the pathway bandwidth. The calculated rate of deactivation of the pathway (0.0041 Hz) derived by monitoring Hog1 translocation in wild-type cells or cells that contained mutations that deactivated either the Sho1 arm or the Sln1 arm matched the pathway bandwidth, which suggests that deactivation of HOG signaling, that is, dephosphorylation, at or downstream of Pbs2 set the pathway bandwidth.

P. Hersen, M. N. McClean, L. Mahadevan, S. Ramanathan, Signal processing by the HOG MAP kinase pathway. Proc. Natl. Acad. Sci. U.S.A. 105, 7165-7170 (2008). [Abstract] [Full Text]