Studies of the temperature-sensitive cdc37-1 mutant of Saccharomyces cerevisiae suggest that Cdc37 is required for passage through the G1 phase of the cell cycle, but its precise function is not known. We have investigated the role of Cdc37 in the regulation of the cyclin-dependent protein kinase Cdc28. We find that G1 arrest in the cdc37-1 mutant is accompanied by a decrease in the Cdc28 activity associated with the G1 cyclin Cln2 (3,4). Mutations in CDC36 and CDC39 result in constitutive activation of the mating pheromone pathway (5), whereas CDC28 is more directly involved in cell cycle control (1, 2, 6).The function of CDC37 is unknown.The product of the CDC28 gene is a member of the highly conserved family of cyclin-dependent kinases (CDKs), whose activation at specific cell cycle stages requires association with cyclin regulatory subunits (2,(7)(8)(9). The commitment to a new cell division cycle in G1 is controlled by complexes of Cdc28 and the G1 cyclins Clnl, -2, and -3. G1 arrest by mating pheromone involves inhibition of specific Cdc28-Cln complexes and decreased synthesis of Clnl and Cln2. Cdc28 function is also required later in the cell cycle: progress through S phase and mitosis requires activation of Cdc28 by S-phase cyclins (Clb5 and -6) and mitotic cyclins (Clbl, -2, -3, and -4), respectively (see ref For a-factor arrest, 120-ml cultures were grown at 24°C to an OD600 of 0.5. a-Factor (1.0 tLg/ml) was added for 2 hr. For mitotic arrest, 25-ml cultures were grown at 24°C to an OD600 of 0.3. Cells were transferred to medium containing benomyl (60 ,ug/ml) and nocodazole (20 ,g/ml) for 2 hr at 24°C.A 5.8-kb genomic DNA fragment containing the CDC37 gene was a gift of S. Reed (Scripps Institute, La Jolla, CA). To construct the cdc37A allele, a fragment of CDC37 (coding for aa Figs. 1-3, logarithmic-phase cells were resuspended in 700 ptl of ice-cold 20 mM Tris-HCl, pH 7.4/0.1% Triton X-100/100 mM NaCl/5 mM EDTA/50 mM P-glycerophosphate/50 mM NaF/1 mM phenylmethylsulfonyl fluoride/1 mM dithiothreitol with aprotinin (2 ,tg/ml) and leupeptin (1 tLg/ml). One milliliter of glass beads was added, and cells were lysed at 4°C by two pulses (60 s) in a miniBeadBeater (Biospec Products, Bartlesville, OK). Lysates were clarified by centrifugation at 14,000 x g for 10 min at 4°C. In Fig. 4

Progress through the cell cycle is governed by the cyclin-dependent kinases (CDKs), the activation of which requires phosphorylation by the CDK-activating kinase (CAK). In vertebrates, CAK is a trimeric enzyme containing CDK7, cyclin H, and MAT1. CAK from the budding yeast Saccharomyces cerevisiae was identified as an unusual 44-kilodalton protein kinase, Cak1, that is only distantly related to CDKs. Cak1 accounted for most CAK activity in yeast cell lysates, and its activity was constant throughout the cell cycle. The CAK1 gene was essential for cell viability. Thus, the major CAK in S. cerevisiae is distinct from the vertebrate enzyme, suggesting that budding yeast and vertebrates may have evolved different mechanisms of CDK activation.

), we have determined that the KIN28 gene used in two of our experiments ( Fig. 3 and 6) contains two additional mutations that arose during PCR amplification of the sequence from a cDNA library. These mutations result in two amino acid changes in nonconserved regions of the Kin28 protein kinase sequence (N123D and M273T). Since this double mutant gene allows growth of kin28-3 cells and encodes a Kin28 protein with abundant kinase activity ( Fig. 3 and 6), these two mutations do not abolish Kin28 function. However, the presence of these mutations does appear to have an impact on the requirement for T162 phosphorylation in Kin28 function. We showed in Fig. 3 that the T162A mutation blocks Kin28 function, suggesting that phosphorylation at this site might be required for full function. However, G. Faye (unpublished data) and M. Solomon's group (J. Kimmelman, P. Kaldis, C.

In the budding yeast Saccharomyces cerevisiae, Cdc37 is required for the productive formation of Cdc28-cyclin complexes. The cdc37-1 mutant arrests at Start with low levels of Cdc28 protein, which is predominantly unphosphorylated at Thr169, fails to bind cyclin, and has little protein kinase activity. We show here that Cdc28 and not cyclin is specifically defective in the cdc37-1 mutant and that Cdc37 likely does not act as an assembly factor for Cdc28-cyclin complex formation. We have also found that the levels and activity of the protein kinase Cak1 are significantly reduced in the cdc37-1 mutant. Pulse-chase analysis indicates that Cdc28 and Cak1 proteins are both destabilized when Cdc37 function is absent during but not after translation. In addition, Cdc37 promotes the production of Cak1, but not that of Cdc28, when coexpressed in insect cells. We conclude that budding yeast Cdc37, like its higher eukaryotic homologs, promotes the physical integrity of multiple protein kinases, perhaps by virtue of a cotranslational role in protein folding.

We consider small-cell networks with multipleantenna transceivers and base stations (BSs) cooperating to jointly design linear precoders to maximize the network energy efficiency, subject to a sum power and per-antenna power constraints at individual BSs, as well as user-specific quality of service (QoS) requirements. Assuming zero-forcing precoding, we formulate the problem of interest as a concave-convex fractional program to which we proposed a centralized optimal solution based on the prevailing Dinkelbach algorithm. To facilitate distributed implementations, we transform the design problem into an equivalent convex program using Charnes-Cooper's transformation. Then, based on the framework of alternative direction method of multipliers (ADMM), we develop a decentralized algorithm, which is numerically shown to achieve fast convergence. Since BSs are generally power-hungry, it may be more energy-efficient if some BSs can be shut down, while still satisfying the QoS constraints. Toward this end, we investigate the problem of joint precoder design and BS selection, which is a mixed Boolean nonlinear program, and then provide an optimal solution by customizing the branch-andbound method. For real-time applications, we propose a greedy algorithm which achieves near-optimal performance in polynomial time. Numerical results are provided to demonstrate the effectiveness of the proposed algorithms.Index Terms-Small-cell networks, energy efficiency, MIMO, joint design, ADMM, branch-and-bound.

Complete activation of most cyclin-dependent protein kinases (CDKs) requires phosphorylation by the CDK-activating kinase (CAK). In the budding yeast, Saccharomyces cerevisiae, the major CAK is a 44-kDa protein kinase known as Cak1. Cak1 is required for the phosphorylation and activation of Cdc28, a major CDK involved in cell cycle control. We addressed the possibility that Cak1 is also required for the activation of other yeast CDKs, such as Kin28, Pho85, and Srb10. We generated three new temperature-sensitive cak1 mutant strains, which arrested at the restrictive temperature with nonuniform budding morphology. All three cak1 mutants displayed significant synthetic interactions with loss-of-function mutations in CDC28 and KIN28. Loss of Cak1 function reduced the phosphorylation and activity of both Cdc28 and Kin28 but did not affect the activity of Pho85 or Srb10. In the presence of the Kin28 regulatory subunits Ccl1 and Tfb3, Kin28 was phosphorylated and activated when coexpressed with Cak1 in insect cells. We conclude that Cak1 is required for the activating phosphorylation of Kin28 as well as that of Cdc28.

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