Differential regulation of p34cdc2 and p33cdk2 by transforming growth factor-β1 in murine mammary epithelial cells

Cyclin-dependent kinases (cdks) are a family of proteins whose function plays a critical role in cell cycle traverse. Transforming growth factor-β₁ (TGF-β₁) is a potent growth inhibitor of epithelial cells. Since cdks have been suggested as possible biochemical markers for TGF-β growth inhibition, we investigated the effect of TGF-β₁ on cdc2 and cdk2 in a normal mouse mammary epithelial cell line (MME) and a TGF-β-resistant MME cell line (BG18.2). TGF-β₁ decreases newly synthesized cdc2 protein levels within 6 h after addition. Coincident with this decrease in newly synthesized cdc2 protein was a marked reduction in its ability to phosphorylate histone H1. This decrease in kinase activity is not due to a change in steady-state levels of cdc2 protein, since mRNA and total protein levels of cdc2 are not reduced until 12 h after TGF-β₁ addition. This suggests that the kinase activity of cdc2 is dependent on newly synthesized cdc2 protien. Moreover, the protein synthesis of another cyclin-dependent kinase, cdk2, is not effected by TGF-β₁ addition, but its kinase activity is substantially reduced. Thus, it appears that TGF-β decreases the kinase activity of both cdc2 and cdk2 by distinct mechanisms.     

Elucidating the functional role of endoreduplication in tomato fruit development

Background

Endoreduplication is the major source of endopolyploidy in higher plants. The process of endoreduplication results from the ability of cells to modify their classical cell cycle into a partial cell cycle where DNA synthesis occurs independently from mitosis. Despite the ubiquitous occurrence of the phenomenon in eukaryotic cells, the physiological meaning of endoreduplication remains vague,although several roles during plant development have been proposed, mostly related to cell differentiation and cell size determination.

Scope

Here recent advances in the knowledge of endoreduplication and fruit organogenesis are reviewed, focusing on tomato (Solanum lycopersicum) as a model, and the functional analyses of endoreduplication-associated regulatory genes in tomato fruit are described.

Conclusions

The cyclin-dependent kinase inhibitory kinase WEE1 and the anaphase promoting complex activator CCS52A both participate in the control of cell size and the endoreduplication process driving cell expansion during early fruit development in tomato. Moreover the fruit-specific functional analysis of the tomato CDK inhibitor KRP1 reveals that cell size and fruit size determination can be uncoupled from DNA ploidy levels, indicating that endoreduplication acts rather as a limiting factor for cell growth. The overall functional data contribute to unravelling the physiological role of endoreduplication in growth induction of fleshy fruits.     

Interferon modulates the messenger RNA of G1-controlling genes to suppress the G1-to-S transition in Daudi cells

Interferon (IFN) is one of the potent antiproliferative cytokines and is used to treat some selected cancers. IFN arrests the growth of Burkitt Iymphoma derived cell line Daudi cells in the G1 phase. G1-to-S progression is controlled by positive and negative regulatory genes. Therefore, we investigated the effects of IFN on G1-controlling genes. Expression of cyclin-dependent kinases (Cdks 2, 3, 4, 5, 6), MO 15/Cdk7, and cyclins E and H was studied to assess positive regulators, while p15^Ink4B, p16^Ink4, p18, p21^CipI, and p27^Kip1 were assessed as negative regulators. Cdks 2, 4, 6 and cyclin E were markedly down-regulated. MO15/Cdk7 expression showed little change, but its regulatory subunit (cyclin H) was down-regulated like cyclin E. Expression of p15^Ink4B and p16^Ink4 was not observed. p18 was induced until 48 h and its expression returned to the initial level at 72 h. In contrast, p21^Cip1 mRNA expression remained at the baseline level throughout IFN treatment, while the expression of p27^Kip1 increased at 48 and 72 h. Taken together, these data indicate that IFN changes the messenger RNA of G1-controlling genes towards the suppression of G1-to-S transition.     

Ectopic expression of p27Kip1 in oligodendrocyte progenitor cells results in cell-cycle growth arrest

Oligodendrocyte differentiation is a complex process believed to be controlled by an intrinsic mechanism associated with cell-cycle arrest. Recently, the cell-cycle inhibitor protein p27^Kip1 has been proposed as a key element in causing growth arrest of oligodendrocyte precursor cells. To investigate the effects of p27 upon oligodendrocyte cell development, we have introduced the p27 cDNA in oligodendrocyte progenitor cells using an adenovirus vector. Progenitor cells normally express low levels of p27. After adenoviral infection and p27 overexpression, progenitor cells were able to undergo cell-cycle arrest, even in the presence of strong mitogens. The effects of p27 were shown to be directly upon cyclin-dependent kinase-2 (CDK2), the protein kinase complex responsible for G1/S transition, as immunodepletion of oligodendrocyte extracts of p27 protein resulted in the activation of CDK2 activity. However, cells that became growth arrested owing to infection with p27 adenovirus did not display conventional oligodendrocyte differentiation markers, such as O4 or O1. Taken together, these data provide mechanistic evidence indicating that p27 is primarily involved in oligodendroglial progenitor proliferation by inhibiting CDK2 activity and inducing oligodendrocyte cell-cycle arrest. © 1998 John Wiley & Sons, Inc. J Neurobiol 36: 431–440, 1998     

Endoreduplication in higher plants

Cell polyploidisation can be achieved by endoreduplication, which consists of one or several rounds of DNA synthesis in the absence of mitosis. As a consequence, chromosomes with 2ⁿ chromatids are produced without change in the chromosome number. Endoreduplication is the most common mode of polyploidisation in plants and can be found in many cell types, especially in those undergoing differentiation and expansion. Although accumulating data reveal that this process is developmentally regulated, it is still poorly understood in plants. At the molecular level, the increasing knowledge on plant cell cycle regulators allows the acquisition of new tools and clues to understand the basis of endoreduplication control and, in particular, the switch between cell proliferation and cell differentiation.     

Leishmania donovani cyclin 1 (LdCyc1) forms a complex with cell cycle kinase subunit CRK3 (LdCRK3) and is possibly involved in S-phase-related activities

Expression of Leishmania donovani cyclin 1 (LdCyc1) mRNA during the cell cycle of promastigotes is S-phase specific. Here, we show that the LdCyc1 protein is periodically expressed and the activity of its associated kinase varies during the cell cycle in line with its expression pattern. In addition, we have shown that LdCRK3, homologous to CRK3 from L. mexicana, is the cognate Cdk partner of LdCyc1 and that the activity of the complex is inhibited specifically by heat stable factor(s) from the parasite.     

Proteasome-dependent degradation of human CDC25B phosphatase

The CDC25 dual specificity phosphatase is a universal cell cycle regulator. The evolutionary conservation of this enzyme from yeast to man bears witness to its major role in the control of cyclin-dependent kinases (CDK) activity that are central regulators of the cell cycle machinery. CDC25 phosphatase both dephosphorylates and activates CDKs. Three human CDC25s have been identified. CDC25A is involved in the control of G1/S, and CDC25C at G2/M throught the activation of CDK1-cyclin B. The exact function of CDC25B however remains elusive. We have found that CDC25B is degraded by the proteasome pathway in vitro and in vivo. This degradation is dependent upon phosphorylation by the CDK1-cyclin A complex, but not by CDK1-cyclin B. Together with the observations of others made in yeast and mammals, our results suggest that CDC25B might act as a mitotic starter triggering the activation of an auto-amplification loop before being degraded.     

Cell cycle function of a rice B2-type cyclin interacting with a B-type cyclin-dependent kinase

Cyclin-dependent kinases (CDKs) are involved in the control of cell cycle progression. Plant A-type CDKs are functional homologs of yeast Cdc2/Cdc28 and are expressed throughout the cell cycle. In contrast, B-type CDK (CDKB) is a family of mitotic CDKs expressed during the S/M phase, and its precise function remains unknown. Here, we identified two B2-type cyclins, CycB2;1 and CycB2;2, as a specific partner of rice CDKB2;1. The CDKB2;1-CycB2 complexes produced in insect cells showed a significant level of kinase activity in vitro, suggesting that CycB2 binds to and activates CDKB2. We then expressed green fluorescent protein (GFP)-fused CDKB2;1 and CycB2;2 in tobacco BY2 cells to investigate their subcellular localization during mitosis. Surprisingly, the fluorescence signal of CDKB2;1-GFP was tightly associated with chromosome alignment as well as with spindle structure during the metaphase. During the telophase, the signal was localized to the spindle midzone and the separating sister chromosomes, and then to the phragmoplast. On the other hand, the CycB2;2-GFP fluorescence signal was detected in nuclei during the interphase and prophase, moved to the metaphase chromosomes, and then disappeared completely after the cells passed through the metaphase. Co-localization of CDKB2;1-GFP and CycB2;2-GFP on chromosomes aligned at the center of the metaphase cells suggests that the CDKB2-CycB2 complex may function in retaining chromosomes at the metaphase plate. Overexpression of CycB2;2 in rice plants resulted in acceleration of root growth without any increase in cell size, indicating that CycB2;2 promoted cell division probably through association with CDKB2 in the root meristem.     

Protein kinase C signaling mediates a program of cell cycle withdrawal in the intestinal epithelium

Members of the protein kinase C (PKC) family of signal transduction molecules have been widely implicated in regulation of cell growth and differentiation, although the underlying molecular mechanisms involved remain poorly defined. Using combined in vitro and in vivo intestinal epithelial model systems, we demonstrate that PKC signaling can trigger a coordinated program of molecular events leading to cell cycle withdrawal into G(0). PKC activation in the IEC-18 intestinal crypt cell line resulted in rapid downregulation of D-type cyclins and differential induction of p21(waf1/cip1) and p27(kip1), thus targeting all of the major G(1)/S cyclin-dependent kinase complexes. These events were associated with coordinated alterations in expression and phosphorylation of the pocket proteins p107, pRb, and p130 that drive cells to exit the cell cycle into G(0) as indicated by concomitant downregulation of the DNA licensing factor cdc6. Manipulation of PKC isozyme levels in IEC-18 cells demonstrated that PKCalpha alone can trigger hallmark events of cell cycle withdrawal in intestinal epithelial cells. Notably, analysis of the developmental control of cell cycle regulatory molecules along the crypt-villus axis revealed that PKCalpha activation is appropriately positioned within intestinal crypts to trigger this program of cell cycle exit-specific events in situ. Together, these data point to PKCalpha as a key regulator of cell cycle withdrawal in the intestinal epithelium.     

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。