Ste5 protein controls a switch-like mating decision

Budding yeast of the species Saccharomyces cerevisiae (Left)


Beer Yeast (Saccharomyces cerevisiae) Plush (Down)

Introduction

Budding yeast of the species Saccharomyces cerevisiae is the most useful yeast especially in brewing and baking. The haploid yeast cell Saccharomyces cerevisiae choose mate partners. MATa and MATα are the two haploid forms of accharomyces cerevisiae. Yeasts have sexual and asexual reproductive processes. The most common is the asexual reproduction by budding in yeast. Haploid (represent as n in the number of chromosomes in individual gamete) cell usually die however, diploid cell goes under the process or sporulation by entering sexual reproduction and produce haploid spores which can mate to diploid (represent as 2n that have two homologous copies of each chromosome).

Figure 1

Figure 1: Life cycle of yeast cell. Diagram showed budding in 1, conjugation in 2 and spore in 3.

Yeast serves as model organisms; they have been used often as a representation of how other cells work and develop. Yeast cells usually produce asexually through a process of budding, which produces a new cell close to original cell by pinching off a portion of the original cell. There are cases of sexual reproduction that occur sometimes; yeast cells reproduce sexually by a mating process in which DNA gets mixed and split from two cells into two cells. To have this mating process, each cell has to produce nodule which one can grouped together is called shmoo. The shmooing process runs around two hours.

In a recent study, scaffold proteins play an essential role in controlling the processes of many signaling cascades. With two or more components of a signaling pathway, scaffold proteins can help to localize signaling molecules to a cell’s specific parts or elevate the signaling pathway’s effectiveness. The other effect of scaffold proteins is the thresholds and dynamics of signaling reactions by positive and negative feedback signals. The function of scaffold protein from Figure 2 is recent progress through understanding of immune cells through mathematical modeling and engineered scaffold proteins. They showed 4 ways that scaffold proteins can function.(Shaw and Filbert 2009)

Figure 2

Figure 2: The function of scaffold proteins. Four functions of scaffold protein have been shown in this diagram. In part a, four diagram showed assembly in a signaling pathway, localization in a signaling pathway to specific intracellular location or compartment, controlled positive and negative feedback signals, and protection of phosphatase deactivation. Part b displays the function curves that were generated.(Shaw and Filbert 2009)

Figure 3

Figure 3 : Distinct morphologies in yeast during mating response : Axial budding, bipolar budding, cell cycle arrested, and shmooing.

Movies

Three movies showed different development over-time morphologies during pheromone response.

Axial budding

Axial budding is in the no α-factor stimulus.

Bipolar budding

Bipolar budding is in the 0.1μM of α-factor stimulus.

Shmooing

Shmooing is in the 0.1μM of α-factor stimulus.

Why is this research, switch-like mating decision in yeast, important?

Yeasts are single-celled microbes which often use as model organisms that help understand how cells work. The switch-like mating decision is poorly understood in molecular mechanism which research team showed the switching mechanism cause MAPK Fus3 and a phosphotase Ptc1 to control protein Ste5. The competition result of Fus3 from Ste5 switch-like dissociation is important to produce the switch-like response. Mating decision is made at the early stage of the pheromone pathway. Architecture of the Fus3-Ste5-Ptc1 circuit produce a novel ultrasensitivity mechanisms which is robust to these proteins. This robustness helps to determine mating in stochastic or genetic variation between each individual. Scaffold proteins are found in numerous eukaryotic signaling pathways. Scaffolded MAPKs are major disease including cancer, diabetes, obesity, etc. For prediction of a future studies, if the similar mechanism occur in mammalian signaling then they can confirm the important goal for therapeutic intervention.

In yeast, how does the scaffold protein Ste5 control a switch-like mating decision?

–      Ste5 protein as a control of cell-fate decision

–      Competition between the MAPK Fus3 and a phosphotase Ptc1

–      Fus3-Ste5 interaction

The role of the Ste5 protein

The haploid yeast organism being studied is Saccharomyces cerevisiae S288c. This particular organism secretes pheromones when it is ready to mate. Ste5 protein is a pheromone-response scaffold protein. This protein is used for coding and is a modulator concerning the decision to mate sexually in yeast cells. Yeast, at cellular and molecular levels, has shown itself to have a lot in common with people when it comes to the mechanism behind the decision to mate.

There is a switching mechanism in yeast cells that causes their decision to mate in as little time as two minutes. The switch mechanism is based upon and controlled by a chemical change on a signal protein. The cause of this mating process is determined through a network of signaling proteins known as mitogen-activated protein kinase (MAPK). (Malleshaiah MK, et al. 2010) When the pheromone concentration is high in the area surrounding the cells the MAPK reacts with four possible sites on the ste5 which in turn causes the mating response. The cell’s decision to mate occurs very quickly in order to prevent the loss of a potential mate to a competing cell. Ste5 needs to direct signaling through mating pathway to the mitogen-activated protein kinase (MAPK), Fus3. (Good et al. 2009)

How does Ste5 affect reproduction?

Interaction between Fus3-Ste5 using an Ste5ND mutant unexpectedly reduces and restricts the mating response and adequately devastates the switch-like shmooing. They reproduce to change the activity in this interaction. Ste5ND has Fus3-docking motif (FDM) which is disrupted so that it does not group with Fus3. When reacting with Ste5ND, Fus3 undergoes the schmoozing process and fits the Hill function. When Shmooing happen, steady-state at Fus3-Ste5 complex determined switch-like dissociation over α-factor. Levels of the Fus3-Ste5 complex, protein-fragment complementation assay on Renilla reniformis (Rluc) signifies kinetics. (Malleshaiah MK, et al. 2010)

Figure 4

Figure 4 : Switch-like shmooing in yeast requires the Fus3–Ste5 interaction. In (a), in MATa cells, α-factor pheromone activates a MAPK cascade that produces phosphorylated, active Fus3. Then, Fus3 dissociates from Ste5 and phosphorylates downstream targets to help mating.
Bottom panel:. Around MATa cells in red show different morphologies as identified by the α -factor concentration sensed. As the MATa cell senses a critical concentration of a-factor in green, it ‘shmoos’ and mates with the MATa cell.
In (b) and (c), The fraction of different morphologies observed in MATa ste5Δ cells expressing either wild-type Ste5, Ste5WT. (b) or the Ste5ND mutant (c). Morphologies: axial in green or bipolar in blue budding, arrested in black and shmooing in red.

Figure 5

Figure 5 :A novel form of ultrasensitivity explains the switch-like mating
decision. In (a), schematic of two-stage binding: Fus3 or Ptc1 first bind to their Ste5 docking sites in green and then group with individual phosphosites in red and grey enzyme
domains. In (b), steady-state Ste5 phosphorylation in open circles versus
a-factor for Ste5 with four in solid or one in dashed phosphosites. Grey bar:
threshold concentration of α-factor. At the alpha-factor threshold, the amount of Ste5 phosphorylation “switches” off. In (c), Fus3-Ste5 complex with non-phosphorylatable mutants. Model predictions for red and blue circles. In (d), Fus3-Ste5 interaction with constitutively phosphorylated mutants. Model fits for red and blue circles.  And in (e), Fus3-Ste5, ptc1Δ cells, and Ste5-Ptc1, wild type cells, interact with α-factor. The Hill coefficient(nH) and errors are calculated.

In their study of the prediction model, they identified robustness of switching to changing the concentration of Ptc1 for both the grouping Fus3 to Ste5. The small amount of cells in shmoo interacts in Fus3-Ste5 and shmoo responded with modulation by phosphosites that are active. They suggested that possible causes include how phosphorylation-dependent change in affinity of Ste5 for Fus3 appears: either the negative charge of Ste5 phosphate group or Ste5 directly affects the Fus3. The important thing is to understand the improvement of α-mediated group with Ptc1 to Ste5 phosphosites. There are two possibilities which Ste5 forms to change for allowing phosphosites to Ptc1 or Ptc1 concentration at Ste5 that increases at the membrane. Ste5ND individual cells always found in one of the morphological states and there are other switches downstream of Fus3-Ste5 switch in population cells.(Malleshaiah MK, et al. 2010)

Figure 6

Figure 6 : Experiment model prediction. In part a, Changes of Fus3-Ste5 with many Ptc1 concentrations in vivo: wild type, knockout(ptc1Δ) and +Ptc1. In part b, steady-state Fus3-Ste5 complex as α-factor using single (-1PS:AbCD), double (-2PS:abCD), triple (-3PS:Abcd), or quadruple (-4PS:abcd) Ste5 non-phosphorylatable mutant. The Hill coefficient(nH) and errors are calculated.

Conclusions

Research from Imperial College London says that the new findings on yeast cells mating habits could help current and future researchers who are looking into the development of cancer cells and stem cells. The researchers used a complex mathematical model involving the concentration of both pheromones around the cell and proteins related to the mating process in order to determine whether mating will occur or not. Because yeast cells and mammalian cells have many of the same proteins, they think that the mathematical model can be used to find out the causes of changes in other non-yeast cells. Hopefully, this model will be able to help in developing new therapies and drugs.

They stated that molecular mechanism in switch-like mating decision is poorly understood. And their graphs have too much jargon so that supplementary reading was helpful and other sources were needed. There is a lot much information about materials and methods and results; however, there needs to be more discussion about Ptc1. This research definitely helps for biotechnology research for future.

References

1. Good, M., et al., The Ste5 scaffold directs mating signaling by catalytically unlocking the Fus3 MAP kinase for activation. Cell, 2009. 136(6): p. 1085-97.

2. Malleshaiah MK, et al.  (2010) The scaffold protein Ste5 directly controls a switch-like mating decision in yeast. Nature 465(7294):101-5

3. Shaw, A.S. and E.L. Filbert, Scaffold proteins and immune-cell signalling. Nat Rev Immunol, 2009. 9(1): p. 47-56.

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