Docking to a Basic Helix Promotes Specific Phosphorylation by G1-Cdk1

Cyclins are the activators of cyclin-dependent kinase (CDK) complex, but they also act as docking scaffolds for different short linear motifs (SLiMs) in CDK substrates and inhibitors. According to the unified model of CDK function, the cell cycle is coordinated by CDK both via general CDK activity thresholds and cyclin-specific substrate docking. Recently, it was found that the G1-cyclins of S. cerevisiae have a specific function in promoting polarization and growth of the buds, making the G1 cyclins essential for cell survival. Thus, while a uniform CDK specificity of a single cyclin can be sufficient to drive the cell cycle in some cells, such as in fission yeast, cyclin specificity can be essential in other organisms. However, the known G1-CDK specific LP docking motif, was not responsible for this essential function, indicating that G1-CDKs use yet other unknown docking mechanisms. Here we report a discovery of a G1 cyclin-specific (Cln1,2) lysine-arginine-rich helical docking motif (the K/R motif) in G1-CDK targets involved in the mating pathway (Ste7), transcription (Xbp1), bud morphogenesis (Bud2) and spindle pole body (Spc29, Spc42, Spc110, Sli15) function of S. cerevisiae. We also show that the docking efficiency of K/R motif can be regulated by basophilic kinases such as protein kinase A. Our results further widen the list of cyclin specificity mechanisms and may explain the recently demonstrated unique essential function of G1 cyclins in budding yeast.

Proteomics Analysis Identified Hub Genes Associated With RNAi-Induced Silencing of G1 Cyclins in MDA-MB-231 Cells

Background: The G1 cyclins are the most potent candidates in the pathogenesis of breast cancer. This study was designed to analyze the synergistic effect of G1 cyclins silencing on the proliferation of breast cancer cells and to identify G1 cyclins-molecular targets by proteomics approach.Methods: The MDA-MB-231 cells were transfected by a dual shRNA vector targeting G1 cyclins through a bidirectional survivin promoter. Silencing efficacy and cell proliferation were evaluated by real-time PCR, Western blot, and MTS assays, respectively. The protein expression profile was evaluated by 2D gel electrophoresis and mass spectroscopy. Further, bioinformatics tools were applied to identify the molecular targets by G1 cyclins and their possible biological consequences.Results: In response to G1 cyclins silencing, the proliferation of cells exposed to the dual shRNA vector decreased significantly at 72 h post-transfection. The reduction of G1 cyclins proteins was following their mRNA expression level as well. Protein signature of cells was altered in response to the silencing of G1 cyclins, 13 up-regulated, and seven down-regulated. Network analysis of G1 cyclins-regulated proteins identified ACTB, HSP90AA1, ALB, and HSPA5 as the hub genes according to the degree method.The regulated proteins by G1 cyclins participate in cancer-related pathways such as PI3K-Akt signaling pathway and pathways in cancer (HSP90AA1; HSP90AB1; HSP9B1), HIF-1 signaling pathway (LDHA; ALDOA; PGK1), apoptosis and proteoglycans in cancer (ACTB).Conclusions: We identified the G1 cyclins-regulated proteins and their mode of action in the pathogenesis of breast cancer. Our findings suggested that G1 cyclins participate in various biological functions. Meanwhile, it offers a new perception of the interactions among G1 cyclins-regulated proteins to identify targeted treatment. It follows-up research in the field of breast cancer.

Roles of G1 cyclins in the temporal organization of yeast cell cycle - a transcriptome-wide analysis

Oscillating gene expression is crucial for correct timing and progression through cell cycle. In Saccharomyces cerevisiae, G1 cyclins Cln1-3 are essential drivers of the cell cycle and have an important role for temporal fine-tuning. We measured time-resolved transcriptome-wide gene expression for wild type and cyclin single and double knockouts over cell cycle with and without osmotic stress. Clustering of expression profiles, peak-time detection of oscillating genes, integration with transcription factor network dynamics, and assignment to cell cycle phases allowed us to quantify the effect of genetic or stress perturbations on the duration of cell cycle phases. Cln1 and Cln2 showed functional differences, especially affecting later phases. Deletion of Cln3 led to a delay of START followed by normal progression through later phases. Our data and network analysis suggest mutual effects of cyclins with the transcriptional regulators SBF and MBF.