Cyclin-Specific Docking Motifs Promote Phosphorylation of Yeast Signaling Proteins by G1/S Cdk Complexes

Abstract

SUMMARY Background The eukaryotic cell cycle begins with a burst of Cdk phosphorylation. In budding yeast, several Cdk substrates are preferentially phosphorylated at the G1/S transition rather than later in the cell cycle when Cdk activity levels are high. These early Cdk substrates include signaling proteins in the pheromone response pathway. Two such proteins, Ste5 and Ste20, are phosphorylated only when Cdk is associated with the G1/S cyclins Cln1 and Cln2, and not G1, S, or M cyclins. The basis of this cyclin specificity is unknown. Results Here we show that Ste5 and Ste20 have recognition sequences, or “docking” sites, for the G1/S cyclins. These docking sites, which are distinct from Clb5/cyclin A-binding “RxL” motifs, bind preferentially to Cln2. They strongly enhance Cln2-driven phosphorylation of each substrate in vivo, and function largely independent of position and distance to the Cdk sites. We exploited this functional independence to re-wire a Cdk regulatory circuit in a way that changes the target of Cdk inhibition in the pheromone response pathway. Furthermore, we uncover functionally active Cln2 docking motifs in several other Cdk substrates. The docking motifs drive cyclin-specific phosphorylation, and the cyclin preference can be switched by using a distinct motif. Conclusions Our findings indicate that some Cdk substrates are intrinsically capable of being phosphorylated by several different cyclin-Cdk forms, but they are inefficiently phosphorylated in vivo without a cyclin-specific docking site. Docking interactions may play a prevalent but previously unappreciated role in driving phosphorylation of select Cdk substrates preferentially at the G1/S transition.

Among B-Type Cyclins Only CLB5 and CLB6 Promote Premeiotic S Phase in Saccharomyces cerevisiae

“…Additionally, both twohybrid and proteomic screens have identified distinct arrays of interacting proteins for Clb3 and Clb5 with surprisingly little overlap (Ito et al 2001;Archambault et al 2004). This strong substrate preference is likely determined by docking domains within both the cyclins and the substrates (Bhaduri and Pryciak 2011;Koivomagi et al 2011). Alternatively it is possible that subcellular localization influences the effectiveness of cyclins in activating premeiotic S phase.…”

“…A growing body of evidence supports the contention that cyclins target Cdk activity to the correct substrates and hence the cyclin provides specificity to the Cdk activity (Cross et al 1999;Roberts 1999;Loog and Morgan 2005;Bhaduri and Pryciak 2011;Koivomagi et al 2011;Pagliuca et al 2011). In addition to interaction with protein substrates, the specificity of cyclins for particular functions can be imparted by the timing of accumulation, the activity of cyclin-CDK inhibitors, and by subcellular localization (Draviam et al 2001;Miller and Cross 2001;Moore et al 2003;Carlile and Amon 2008).…”

Millimeter wave treatment promotes chondrocyte proliferation via G1/S cell cycle transition

“…Cyclin D1 forms complexes with CDK4 or CDK6, which play an important role in the G 1 /S transition by phosphorylating Rb (24). As a consequence of Rb phosphorylation, E2F is released from the Rb/E2F complexes and then triggers cell progression from the G 1 to S phase (25). During the progression of the cell cycle, CDK activity can be blocked by binding of CDKIs.…”

Molecular basis of the functional distinction between Cln1 and Cln2 cyclins

“…Very recently, it was described that some Cln targets involved in the response to α mating factor like Ste5 and Ste20 contain cyclin docking domains that are better recognized by Cln2 than by Cln1. 41 It is possible that the 225-299 fragment of Cln2, but not the equivalent region from Cln1, might directly contribute to the recognition of the specific CDK substrates involved in morphogenesis and in the process of budding independently of, or in addition to, its effect in cyclin localization.…”

“…40 More recently, cyclin-specific docking motifs that bind preferentially to Cln2 have been described in signaling proteins, such as Ste5 and Ste20. 41 In this work, we attempt to describe new molecular mechanisms contributing to the functional distinction between Cln1 and Cln2.…”