Recent advances on cyclins, CDKs and CDK inhibitors

In eukaryotes, cell division is controlled by cyclin-dependent kinases (CDKs). Here we summarize a few new developments on the regulation of the cell cycle by CDK-cyclin complexes. We have focused on three aspects in which there has been recent progress: the structural analysis of these complexes, the phenotypes of mice carrying knockouts of CDK inhibitors and the role of proteolysis in the regulation of the cell cycle.     

The cell cycle: a review of regulation,deregulation and therapeutic targets in cancer

The cell cycle is controlled by numerous mechanisms ensuring correct cell division. This review will focus on these mechanisms, i.e. regulation of cyclin-dependent kinases (CDK) by cyclins, CDK inhibitors and phosphorylating events. The quality checkpoints activated after DNA damage are also discussed. The complexity of the regulation of the cell cycle is also reflected in the different alterations leading to aberrant cell proliferation and development of cancer. Consequently, targeting the cell cycle in general and CDK in particular presents unique opportunities for drug discovery. This review provides an overview of deregulation of the cell cycle in cancer. Different families of known CDK inhibitors acting by ATP competition are also discussed. Currently, at least three compounds with CDK inhibitory activity (flavopiridol, UCN-01, roscovitine) have entered clinical trials.     

Cyclin-dependent kinase 5 (CDK5) and neuronal cell death

Many neurological disorders like Parkinson's and Alzheimer's disease, amyotrophic lateral sclerosis (ALS) or stroke have in common a definite loss of CNS neurons due to apoptotic or necrotic neuronal cell death. Previous studies suggested that proapoptotic stimuli may trigger an abortive and, therefore, eventually fatal cell cycle reentry in postmitotic neurons. Neuroprotective effects of small molecule inhibitors of cyclin-dependent kinases (CDKs), which are key regulators of cell cycle progression, support the cell cycle theory of neuronal apoptosis. However, growing evidence suggests that deregulated CDK5, which is not involved in cell cycle control, rather than cell cycle relevant members of the CDK family, promotes neuronal cell death. Here we summarize the current knowledge about the involvement of CDK5 in neuronal cell death and discuss possible up- or downstream partners of CDK5. Moreover, we discuss potential therapeutic options that might arise from the identification of CDK5 as an important upstream element of neuronal cell death cascades.     

Immunomodulatory effects of CDK4/6 inhibitors

The dysregulation of the cell cycle is one of the hallmarks of cancer. Cyclin-dependent kinase 4 (CDK4) and CDK6 play crucial roles in regulating cell cycle and other cellular functions. CDK4/6 inhibitors have achieved great success in treating breast cancers and are currently being tested extensively in other tumor types as well. Accumulating evidence suggests that CDK4/6 inhibitors exert antitumor effects through immunomodulation aside from cell cycle arrest. Here we outline the immunomodulatory activities of CDK4/6 inhibitors, discuss the immune mechanisms of drug resistance and explore avenues to harness their immunotherapeutic potential when combined with immune checkpoint inhibitors (ICIs) or chimeric antigen receptor (CAR) T-cell therapy to improve the clinical outcomes.     

“Amyloid‐beta accumulation cycle” as a prevention and/or therapy target for Alzheimer's disease

The cell cycle and its regulators are validated targets for cancer drugs. Reagents that target cells in a specific cell cycle phase (e.g., antimitotics or DNA synthesis inhibitors/replication stress inducers) have demonstrated success as broad‐spectrum anticancer drugs. Cyclin‐dependent kinases (CDKs) are drivers of cell cycle transitions. A CDK inhibitor, flavopiridol/alvocidib, is an FDA‐approved drug for acute myeloid leukemia. Alzheimer's disease (AD) is another serious issue in contemporary medicine. The cause of AD remains elusive, although a critical role of latent amyloid‐beta accumulation has emerged. Existing AD drug research and development targets include amyloid, amyloid metabolism/catabolism, tau, inflammation, cholesterol, the cholinergic system, and other neurotransmitters. However, none have been validated as therapeutically effective targets. Recent reports from AD‐omics and preclinical animal models provided data supporting the long‐standing notion that cell cycle progression and/or mitosis may be a valid target for AD prevention and/or therapy. This review will summarize the recent developments in AD research: (a) Mitotic re‐entry, leading to the “amyloid‐beta accumulation cycle,” may be a prerequisite for amyloid‐beta accumulation and AD pathology development; (b) AD‐associated pathogens can cause cell cycle errors; (c) thirteen among 37 human AD genetic risk genes may be functionally involved in the cell cycle and/or mitosis; and (d) preclinical AD mouse models treated with CDK inhibitor showed improvements in cognitive/behavioral symptoms. If the “amyloid‐beta accumulation cycle is an AD drug target” concept is proven, repurposing of cancer drugs may emerge as a new, fast‐track approach for AD management in the clinic setting.     

Regulation of CDK4

Cyclin-dependent kinase (CDK)4 is a master integrator that couples mitogenic and antimitogenic extracellular signals with the cell cycle. It is also crucial for many oncogenic transformation processes. In this overview, we address various molecular features of CDK4 activation that are critical but remain poorly known or debated, including the regulation of its association with D-type cyclins, its subcellular location, its activating Thr172-phosphorylation and the roles of Cip/Kip CDK "inhibitors" in these processes. We have recently identified the T-loop phosphorylation of CDK4, but not of CDK6, as a determining target for cell cycle control by extracellular factors, indicating that CDK4-activating kinase(s) might have to be reconsidered.     

植物细胞周期调控因子及其功能的研究进展

细胞周期调控因子能通过影响细胞周期对植物细胞的生长、分裂和分化产生作用,进而调节植物的生长发育。本文综述了近几年来植物细胞周期调控因子中细胞周期蛋白（cyclin,CYC）、周期蛋白依赖激酶（cyclin-dependent kinase,CDK）等的作用机理及研究进展,阐述了各调控因子在植物生长发育过程中的作用。     

Regional Expression of Key Cell Cycle Proteins in Brain from Subjects with Amnestic Mild Cognitive Impairment

Mild cognitive impairment (MCI) is regarded as a transition stage between the cognitive changes of normal aging and the more serious problems caused by Alzheimer’s disease (AD). Previous studies had demonstrated increased expression of cell cycle proteins in AD brain. In the present study, we have analyzed the expression of the cell cycle proteins, CDK2, CDK5 and cyclin G1 in hippocampus and inferior parietal lobule (IPL) in subjects with amnestic mild cognitive impairment and control using Western blot analysis. The expression of CDK2, CDK5 and cyclin G1 were found to be significantly increased in MCI hippocampus as well as in IPL compared to control brain. These results suggest that some cells may have re-entered the cell cycle. However, the expression of CDK2 and CDK5 is greater in MCI hippocampus compared to those of MCI IPL, and hippocampus is a region that is severely affected by AD pathology. Since these proteins are involved directly or indirectly in microtubule destabilization and hyperphosphorylation of tau, and also in APP processing we hypothesize that cell cycle disturbance may be important contributor in the pathogenesis of AD. Special issue dedicated to Dr. John P. Blass.     

Palbociclib-mediated cell cycle arrest can occur in the absence of the CDK inhibitors p21 and p27

The use of CDK4/6 inhibitors in the treatment of a wide range of cancers is an area of ongoing investigation. Despite their increasing clinical use, there is limited understanding of the determinants of sensitivity and resistance to these drugs. Recent data have cast doubt on how CDK4/6 inhibitors arrest proliferation, provoking renewed interest in the role(s) of CDK4/6 in driving cell proliferation. As the use of CDK4/6 inhibitors in cancer therapies becomes more prominent, an understanding of their effect on the cell cycle becomes more urgent. Here, we investigate the mechanism of action of CDK4/6 inhibitors in promoting cell cycle arrest. Two main models explain how CDK4/6 inhibitors cause G1 cell cycle arrest, which differ in their dependence on the CDK inhibitor proteins p21 and p27. We have used live and fixed single-cell quantitative imaging, with inducible degradation systems, to address the roles of p21 and p27 in the mechanism of action of CDK4/6 inhibitors. We find that CDK4/6 inhibitors can initiate and maintain a cell cycle arrest without p21 or p27. This work clarifies our current understanding of the mechanism of action of CDK4/6 inhibitors and has implications for cancer treatment and patient stratification.     

Regulation of cyclin-dependent kinase 2 activity by ceramide

Cyclin-dependent kinases have been implicated in the inactivation of retinoblastoma (Rb) protein and cell cycle progression. Recent studies have demonstrated that the lipid molecule ceramide is able to induce Rb hypophosphorylation leading to growth arrest and cellular senescence. In this study, we examined the underlying mechanisms of Rb hypophosphorylation and cell cycle progression utilizing the antiproliferative molecule ceramide. C6-Ceramide induced a G0/G1 arrest of the cell cycle in WI38 human diploid fibroblasts. Employing immunoprecipitation kinase assays, we found that ceramide specifically inhibited cyclin-dependent kinase CDK2, with a mild effect on CDC2 and significantly less effect on CDK4. The effect of ceramide was specific such that C6-dihydroceramide was not effective. Ceramide did not directly inhibit CDK2 in vitro but caused activation of p21, a major class of CDK-inhibitory proteins, and led to a greater association of p21 to CDK2. Using purified protein phosphatases, we showed that ceramide activated both protein phosphatase 1 and protein phosphatase 2A activities specific for CDK2 in vitro. Further, calyculin A and okadaic acid, both potent protein phosphatase inhibitors, together almost completely reversed the effects of ceramide on CDK2 inhibition. Taken together, these results demonstrate a dual mechanism by which ceramide inhibits the cell cycle. Ceramide causes an increase in p21 association with CDK2 and through activation of protein phosphatases selectively regulates CDK2. These events may lead to activation of Rb protein and subsequent cell cycle arrest.     

Identification and characterization of the BmCyclin L1-BmCDK11A/B complex in relation to cell cycle regulation

Cyclin proteins are the key regulatory and activity partner of cyclin-dependent kinases (CDKs), which play pivotal regulatory roles in cell cycle progression. In the present study, we identified a Cyclin L1 and 2 CDK11 2 CDK11 splice variants, CDK11A and CDK11B, from silkworm, Bombyx mori. We determined that both Cyclin L1 and CDK11A/B are nuclear proteins, and further investigations were conducted to elucidate their spatiofunctional features. Cyclin L1 forms a complex with CDK11A/B and were co-localized to the nucleus. Moreover, the dimerization of CDK11A and CDK11B and the effects of Cyclin L1 and CDK11A/B on cell cycle regulation were also investigated. Using overexpression or RNA interference experiments, we demonstrated that the abnormal expression of Cyclin L1 and CDK11A/B leads to cell cycle arrest and cell proliferation suppression. Together, these findings indicate that CDK11A/B interacts with Cyclin L1 to regulate the cell cycle.     

The Rb-CDK4/6 signaling pathway is critical in neural precursor cell cycle regulation

The tumor suppressor, retinoblastoma (Rb), is involved in both terminal mitosis and neuronal differentiation. We hypothesized that activation of the Rb pathway would induce cell cycle arrest in primary neural precursor cells, independent of the proposed function of cyclin-dependent kinases 4/6 (CDK4/6) to sequester the CIP/KIP CDK inhibitors (CKIs) p21 and p27 from CDK2. We expressed dominant negative adenovirus mutants of CDKs 2, 4, and 6 (dnCDK2, dnCDK4, and dnCDK6) in neural progenitor cells derived from E12.5 wild type and Rb-deficient mouse embryos. In contrast to previous studies, our results demonstrate that in addition to dnCDK2, the dnCDK4/6 mutants can induce growth arrest. Moreover, the dnCDK4/6-mediated inhibition is Rb-dependent. The dnCDK2 partially inhibited cell growth in Rb-deficient cells, suggesting that CDK2 may have additional targets. A previously proposed function of CDK4/6 is CKI sequestration, thereby preventing the resulting inhibition of CDK2, believed to be the key regulator of cell cycle. However, our immunoprecipitations revealed that the dominant negative CDK mutants could arrest cell growth despite their interaction with p21 and p27. Taken together, our results demonstrate that both CDK2 and CDK4/6 are crucial for cell cycle regulation. Furthermore, our data underscore the importance of the Rb regulatory pathway in neuronal development and cell cycle regulation, independent of CKI sequestration.     

A dinoflagellate CDK5-like cyclin-dependent kinase

Background information. Mitosis during the dinoflagellate cell cycle is unusual in that the nuclear envelope remains intact and segregation of the permanently condensed chromosomes uses a cytoplasmic mitotic spindle. To examine regulation of the dinoflagellate cell cycle in the context of these unusual nuclear features, it is necessary to isolate and characterize cell cycle regulators such as CDK (cyclin‐dependent kinase). Results. We report the characterization of a CDK from the dinoflagellate Lingulodinium polyedrum. This CDK reacts with an anti‐PSTAIRE antibody and was identified by protein microsequencing after partial purification. The protein microsequence shows homology toward the Pho85/CDK5 clade of CDKs. Neither the amount nor the phosphorylation state changed over the course of the cell cycle, in agreement with results reported for CDK5 family members in other systems. Conclusions. We conclude we have probably isolated a dinoflagellate CDK5‐like protein. The data reported here support the identification of this protein as a CDK5 homologue, and suggest that dinoflagellates may contain several CDK families.     

Identification and characterization of CAC1 as a novel CDK2-associated cullin

Cell cycle progression is tightly controlled by cyclins and cyclin-dependent kinases (CDKs). CDK2 plays a crucial role in regulating cell cycle progression, but how CDK2 is regulated is still incompletely understood. In this study, we report the identification and characterization of a novel gene CAC1 that regulates CDK2 activity. The open reading frame sequence of this gene encodes a protein of 369 amino acids which contains a Cullin domain, and this protein is physically associated with CDK2. As such, we have designated it Cdk-Associated Cullin1, or CAC1. CAC1 is highly expressed in cancer tissues and cancer cell lines. Interestingly, CAC1 is expressed in a cell cycle-dependent manner and its expression is high in late G1 to S phase. Knockdown of CAC1 by RNAi inhibits cell proliferation and induces G1/S arrest. Since CAC1 interacts with CDK2 and promotes the kinase activity of CDK2 protein, we propose that CAC1 is a novel cell cycle associated protein capable of promoting cell proliferation. Our data provide insight into the mechanism by which CDK2 is regulated and the molecular basis of cell cycle progression in cancer.