Pom1 gradient buffering through intermolecular auto‐phosphorylation

Abstract

Concentration gradients provide spatial information for tissue patterning and cell organization, and their robustness under natural fluctuations is an evolutionary advantage. In rod-shaped Schizosaccharomyces pombe cells, the DYRK-family kinase Pom1 gradients control cell division timing and placement. Upon dephosphorylation by a Tea4-phosphatase complex, Pom1 associates with the plasma membrane at cell poles, where it diffuses and detaches upon auto-phosphorylation. Here, we demonstrate that Pom1 auto-phosphorylates intermolecularly, both in vitro and in vivo, which confers robustness to the gradient. Quantitative imaging reveals this robustness through two system’s properties: The Pom1 gradient amplitude is inversely correlated with its decay length and is buffered against fluctuations in Tea4 levels. A theoretical model of Pom1 gradient formation through intermolecular auto-phosphorylation predicts both properties qualitatively and quantitatively. This provides a telling example where gradient robustness through super-linear decay, a principle hypothesized a decade ago, is achieved through autocatalysis. Concentration-dependent autocatalysis may be a widely used simple feedback to buffer biological activities.

Multi-phosphorylation reaction and clustering tune Pom1 gradient mid-cell levels according to cell size

“…Two models have been proposed to explain the source of this correction. One model suggests gradient buffering is the consequence of concentration-dependent inter-molecular phosphorylation, which promotes Pom1 detachment from the membrane (Hersch et al, 2015). This model also explains the correction for the even larger variations in levels of Tea4 concentration at cell 3 poles.…”

“…Thus, these Pom1 mutants poorly correct intrinsic variations of Pom1 concentrations at the cell tips. Another evidence for buffering comes from the steep decrease in the coefficient of variation of Pom1 WT from the cell tip to the cell middle (Hersch et al, 2015). While we confirm this observation with our current data set, the decrease in the coefficient of variation at cell tips and cell sides is much smaller for Pom1 1A , Pom1 2A and Pom1 3A (from 9.6 for Pom1 WT to 6.6 12 for Pom1 1A , 5.8 for Pom1 2A , and 4.2 for Pom1 3A ; Figure 4F, Figure 4 -supplement 1C).…”

“…The Pom1 gradient forms upon local dephosphorylation of Pom1 at the cell pole, promoting Pom1 membrane binding (Hachet et al, 2011). Previous modelling work demonstrated that the multi-phosphorylation reaction Pom1 undergoes prior to its membrane detachment could provide a timer function for Pom1 lateral diffusion and serve as a buffering mechanism for the gradient (Hersch et al, 2015). An alternative model proposed that differential diffusion coefficients of Pom1 clusters of varying sizes underlie the buffering of the gradient with the largest clusters causing a "traffic jam" event at the cell tip, which is relieved by the progressive fractionation and the subsequent increase in diffusion coefficients of smaller clusters away from the gradient source (Saunders et al, 2012).…”

“…Info 1). Briefly, all 240 gradient profiles were aligned to the maximum value at the cell pole and smoothed with a Gaussian filter as previously described (Hersch et al, 2015). The profiles were binned by 5% and the first 0.5μm of the profile values were deleted to avoid the effect of the gradient plateau at the cell pole.…”

“…An interesting feature of Pom1 gradients is a noise correction mechanism that compensates for large variations in protein concentration at the cell poles, which can vary up to 4-fold (Hersch et al, 2015;Saunders et al, 2012). In a simple diffusive gradient, the decay length (the distance at which the concentration is reduced to a certain fraction of its amplitude) is independent of the amplitude at the pole.…”