Cyclin D1 binds to the Cdk4 and Cdk6 to form a pRB kinase. Upon phosphorylation, pRB loses its repressive activity for the E2F transcription factor, which then activates transcription of several genes required for the transition from the G1-to S-phase and for DNA replication. The cyclin D1 gene is rearranged and overexpressed in centrocytic lymphomas and parathyroid tumors and it is amplified and/or overexpressed in a major fraction of human tumors of various types of cancer. Ectopic overexpression of cyclin D1 in fibroblast cultures shortens the G1 phase of the cell cycle. Furthermore, it has been demonstrated that introduction of an antisense cyclin D1 into a human carcinoma cell line, in which the cyclin D1 gene is amplified and overexpressed, causes reversion of the malignant phenotype. Thus, increased expression of cyclin D1 can play a critical role in tumor development and in maintenance of the malignant phenotype. However, it is insufficient to confer transformed properties on primary or established fibroblasts. In this review, we summarize the role of cyclin D1 on tumor development and malignant transformation. In addition, our chemical biology study to understand the regulatory mechanism of cyclin D1 transcription is also reviewed. (Cancer Sci 2007; 98: 629 -635) I n mammalian cells, the progression of replicating cells through the cell cycle is controlled by the sequential formation, activation, and subsequent inactivation of a series of specific cyclindependent kinase (CDK) complexes. The second regulatory mechanism involves the binding of specific inhibitory proteins (p21WAF1, p27KIP1, p57KIP2) to cyclin-CDK complexes or the binding of specific inhibitory proteins directly to CDK4 (p15, p16INK1, p18, and p19).(1−3) There is now abundant evidence that disturbances in specific cyclins, CDKs, or the above-mentioned inhibitory proteins play an important role in several types of human cancer.(1−3) The most frequent abnormalities relate to cyclin D1. The cyclin D1 gene encodes a regulatory subunit of the cdk4 and cdk6 holoenzyme complex, which phosphorylates and deactivates the tumor suppressor protein pRB.(3) The phophorylation of pRB results in its inactivation and the release of E2Fs that have been sequestered by the unphosphorylated (active) form of pRB. Once liberated by pRB inactivation, E2Fs then proceed to activate genes that are essential for advances into the late G1 and S phases. Consistent with its growth-promoting role, cyclin D1 can act as an oncogene. Indeed, rearrangement, amplification, and/or increased expression of the cyclin D1 gene and overexpression of its mRNA have been reported in several types of human cancer, including human parathyroid adenomas, B cell lymphomas, breast, colon, lung, bladder and liver cancers, and squamous carcinomas of the esophagus, head and neck. (2,5,6) The expression of cyclin D1 mRNA and protein peaks during mid-G1 when growth factor-deprived cells are restimulated to enter the cell cycle. (7,8) Inhibition of cyclin D1 expression either by antisense methodo...

Insulin stimulated translocation of the glucose transporter GLUT4 from the cytosol to the plasma membrane in a concentration (1 nM–1 μM)-dependent manner and increased glucose uptake in 3T3-L1 adipocytes. Insulin-induced GLUT4 translocation to the cell surface was prevented by the phosphoinositide 3 kinase (PI3K) inhibitor wortmannin, the 3-phosphoinositide-dependent protein kinase 1 (PDK1) inhibitor BX912 or the Akt1/2 inhibitor MK2206, and by knocking-down PI3K, PDK1 or Akt1/2. Insulin increased phosphorylation of Akt1/2 at Thr308/309 and Ser473/474, to activate Akt1/2, in the adipocytes. Insulin-induced phosphorylation of Akt1/2 was suppressed by wortmannin and knocking-down PI3K, while no significant inhibition of the phosphorylation was obtained with BX912 or knocking-down PDK1. In the cell-free Akt assay, PI3K phosphorylated Akt1 both at Thr308 and Ser473 and Akt2 at Ser474 alone. In contrast, PDK1 phosphorylates Akt1 at Thr308 and Akt2 at Thr309. The results of this study indicate that PI3K activates Akt1, independently of PDK1, and Akt2 by cooperating with PDK1 in the insulin signal transduction pathway linked to GLUT4 translocation.

To elucidate whether the amount of tissue-deposited amyloid in familial amyloid polyneuropathy (FAP) patients decreases or increases over the long-term course after liver transplantation (LT), we examined changes in histopathological and biochemical characteristics of abdominal fat amyloid in the transplanted patients with a postoperative history of more than 10 years. Using a series of aspirated abdominal fat tissues from 6 FAP patients with transthyretin (TTR) Val30Met variant, the severity of amyloid deposits was examined and the composition ratio of wild type-to-variant TTR in fat amyloid was assayed by liquid chromatography-ion trap mass spectrometry (LC-MS/MS). Histopathological examination of abdominal fat tissues demonstrated a significant decrease or disappearance of amyloid deposits in all 6 patients. On LC-MS/MS analysis, the contribution of wild-type TTR to the composition ratio in amyloid fibrils was markedly increased in all patients after LT. This is the first report showing pathological evidence that deposited amyloid in FAP patients with long posttransplantation courses can gradually regress or disappear.

Glycogen synthase kinase-3β (GSK-3β) is a key element to phosphorylate tau and form neurofibrillary tangles (NFTs) found in tauopathies including Alzheimer's disease (AD). A current topic for AD therapy is focused upon how to prevent tau phosphorylation. In the present study, PKCε activated Akt and inactivated GSK-3β by directly interacting with each protein. Inhibition of protein tyrosine phosphatase 1B (PTP1B), alternatively, caused an enhancement in the tyrosine phosphorylation of insulin receptor substrate 1 (IRS-1), allowing activation of Akt through a pathway along an IRS-1/phosphatidylinositol 3 kinase (PI3K)/3-phosphoinositide-dependent protein kinase-1 (PDK1)/Akt axis, to phosphorylate and inactivate GSK-3β. Combination of PKCε activation and PTP1B inhibition more sufficiently activated Akt and inactivated GSK-3β than each independent treatment, to suppress amyloid β (Aβ)-induced tau phosphorylation and ameliorate spatial learning and memory impairment in 5xFAD transgenic mice, an animal model of AD. This may represent an innovative strategy for AD therapy.

Background/Aims: Our earlier studies suggested crosstalk between IRS/PI3 kinase/PDK1/Akt/Rac1/ROCK and (Shc2/Grb2/SOS)/Ras/Raf/MEK/ERK pathways downstream PDGF-ββ receptor responsible for chemotaxis and proliferation of malignant mesothelioma cells. The present study was conducted to obtain evidence for this. Methods: To assess activation of Akt, MEK, and ERK, Western blotting was carried out on MSTO-211H malignant mesothelioma cells using antibodies against phospho-Thr308-Akt, phopho-Ser473-Akt, Akt, phospho-MEK, MEK, phopho-ERK1/2, and ERK1/2. To knock-down Akt, PI3 kinase, PDK1, and Rac1, siRNAs silencing each-targeted gene were constructed and transfected into cells. To monitor Rac1 activity, FRET monitoring was carried out on living and fixed cells. Results: ERK was activated under the basal conditions in MSTO-211H cells, and the activation was prevented by inhibitors for PI3 kinase, PDK1, Akt, and Rac1 or by knocking-down PI3 kinase, PDK1, Akt, and Rac1. Akt was also activated under the basal conditions, and the activation was suppressed by a MEK inhibitor and an ERK1/2 inhibitor. In the FRET analysis, Rac1 was activated under the basal conditions, and the activation was inhibited by a MEK inhibitor and an ERK1/2 inhibitor. Conclusion: The results of the present study show that ERK could be activated by PI3 kinase, PDK1, Akt, and Rac1 and that alternatively, Akt and Rac1 could be activated by MEK and ERK in MSTO-211H cells.

We report a 67-year-old male patient who suffered from nephrotic syndrome and progressive renal dysfunction with monoclonal gammopathy (IgMkappa). Renal biopsy demonstrated amyloid deposition in glomeruli. Immunohistochemical studies of the renal amyloid using a number of antibodies, including anti-lambda and anti-kappa light chains, AA, beta(2)-microglobulin, and transthyretin, showed negative findings. Biochemical analysis of the deposited amyloid fibrils in gastroduodenal mucosa revealed that the amyloid fibrils were composed of an immunoglobulin heavy chain variable region (VH) fragment belonging to the VH1 subgroup, and a diagnosis of AH amyloidosis was made. In our institute, three patients with AH amyloidosis including the present one have been identified during the past 2 years, so AH amyloidosis seems to be by no means a rare disorder.

A library of oxygenated natural steroids, including physalins, withanolides, and perulactones, coupled with the synthetic cage-shaped right-side structure of type B physalins, was constructed. SAR studies for inhibition of NF-κB activation showed the importance of both the B-ring and the oxygenated right-side partial structure. The 5β,6β-epoxy derivatives of both physalins and withanolides showed similar profiles of inhibition of NF-κB activation and appeared to act on NF-κB signaling via inhibition of phosphorylation and degradation of IκBα. In contrast, type B physalins with C5-C6 olefin functionality inhibited nuclear translocation and DNA binding of RelA/p50 protein dimer, which lie downstream of IκBα degradation, although withanolides having the same AB-ring functionality did not. These results indicated that the right-side partial structure of these steroids influences their mode of action.

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