SuppInfo

SuppInfo - Supplementary Information I. Experimental...

Info iconThis preview shows pages 1–3. Sign up to view the full content.

View Full Document Right Arrow Icon
Supplementary Information I.Experimental Methods Wild type yeast S. cerevisiae cells (strain W303) were grown overnight in YPD medium then transferred to YNB+ethanol for 2-4 hours. Cells were then imaged using the microfluidic platform ONIX (CellAsic) and an inverted microscope (Nikon Eclipse Ti-E). We imaged the NADH autofluorescence (excitation 370 nm, emission 460 nm) in the cells as the ethanol medium is switched to a medium containing Yeast Nitrogen Base (YNB), glucose, and potassium cyanide (KCN) to induce anaerobic glycolysis. Both simultaneous and independent addition of glucose and KCN were performed. We chose the highest flow rate which will not dislodge the cells in order to ensure the media is shifted as abruptly as possible. For the ONIX microfluidic pump, this flow rate was at 7 psi. In a separate experiment, cells were harvested and starved by resuspending them in phosphate buffer (PBS) before adding glucose and KCN, which induces oscillations in dense cell suspension [15]. Fluorescence measurement is taken every 3 seconds. Note that while synchronized and sustained oscillation is found in dense whole yeast cell suspension, we cannot achieve this density on a single cell layer using the microfluidic chamber. Time-lapse images show a portion of the cells exhibiting a transient oscillation with a period of about 30 seconds in response to glucose and KCN addition. This period is in good agreement with the 36 s period of NADH oscillation observed in dense yeast cell suspensions [15]. Additionally, when the cells are starved in phosphate buffer before the shifting to glucose and KCN, a larger portion of the cells exhibit transient oscillations. Concentrations of KCN and
Background image of page 1

Info iconThis preview has intentionally blurred sections. Sign up to view the full version.

View Full DocumentRight Arrow Icon
glucose were varied and responses were compared but no sustained oscillation was observed. Further attempt to stress the cells by heat shock (which unfolds enzymes, lowering k , and increases ATP demand) or by amino acid starvation (lowering enzyme levels) still did not induce oscillations. II. Comparison between Power law formalism and Michaelis-Menten form Let f define the function of the Michaelis Menten form:   2 2 1 1 c c c c x f x x x (S.1.1) For the negative feedback case, this becomes   2 2 1 1 h h h h x f x x x (S.1.2) If we apply f(x) to all the reactions in (1.1) of the main paper, we get:             ( ) 1 ( ) g h a h a g y x f y f y kf y f x y qf y y q kf y f x k   (S.1.3) Which linearizes to:       '( ) 2 1 1 '( ) 2 a h g kf x x x q a h q g y y q kf x   (S.1.4) Hence, in the linear version, the contribution of the parameters a, h, g in the linearized Michaelis Menten form is simply scaled by ½ compared to the power-law form. Additionally, if the basal
Background image of page 2
Image of page 3
This is the end of the preview. Sign up to access the rest of the document.

Page1 / 16

SuppInfo - Supplementary Information I. Experimental...

This preview shows document pages 1 - 3. Sign up to view the full document.

View Full Document Right Arrow Icon
Ask a homework question - tutors are online