Notch4 is required for tumor onset and perfusion.
2013; 5 (1): 7-?
Circadian Rhythm Gene Period 3 Is an Inhibitor of the Adipocyte Cell Fate
JOURNAL OF BIOLOGICAL CHEMISTRY
2011; 286 (11): 9063-9070
Notch4 is a member of the Notch family of receptors that is primarily expressed in the vascular endothelial cells. Genetic deletion of Notch4 does not result in an overt phenotype in mice, thus the function of Notch4 remains poorly understood.We examined the requirement for Notch4 in the development of breast cancer vasculature. Orthotopic transplantation of mouse mammary tumor cells wild type for Notch4 into Notch4 deficient hosts enabled us to delineate the contribution of host Notch4 independent of its function in the tumor cell compartment.Here, we show that Notch4 expression is required for tumor onset and early tumor perfusion in a mouse model of breast cancer. We found that Notch4 expression is upregulated in mouse and human mammary tumor vasculature. Moreover, host Notch4 deficiency delayed the onset of MMTV-PyMT tumors, wild type for Notch4, after transplantation. Vessel perfusion was decreased in tumors established in Notch4-deficient hosts. Unlike in inhibition of Notch1 or Dll4, vessel density and branching in tumors developed in Notch4-deficient mice were unchanged. However, final tumor size was similar between tumors grown in wild type and Notch4 null hosts.Our results suggest a novel role for Notch4 in the establishment of tumor colonies and vessel perfusion of transplanted mammary tumors.
View details for DOI 10.1186/2045-824X-5-7
View details for PubMedID 23601498
Genetic Dissection of the Oncogenic mTOR Pathway Reveals Druggable Addiction to Translational Control via 4EBP-eIF4E
2010; 17 (3): 249-261
Glucocorticoids rapidly and robustly induce cell fate decisions in various multipotent cells, although the precise mechanisms of these important cellular events are not understood. Here we showed that glucocorticoids repressed Per3 expression and that this repression was critical for advancing mesenchymal stem cells to the adipocyte fate. Exogenous expression of Per3 inhibited adipogenesis, whereas knocking out Per3 enhanced that fate. Moreover, we found that PER3 formed a complex with PPAR? and inhibited PPAR?-mediated transcriptional activation via Ppar? response elements. Consistent with these findings, Per3 knock-out mice displayed alterations in body composition, with both increased adipose and decreased muscle tissue compared with wild-type mice. Our findings identify Per3 as potent mediator of cell fate that functions by altering the transcriptional activity of PPAR?.
View details for DOI 10.1074/jbc.M110.164558
View details for Web of Science ID 000288247700037
View details for PubMedID 21228270
Role of caveolin-1 in thyroid phenotype, cell homeostasis, and hormone synthesis: in vivo study of caveolin-1 knockout mice
AMERICAN JOURNAL OF PHYSIOLOGY-ENDOCRINOLOGY AND METABOLISM
2009; 297 (2): E438-E451
We genetically dissect the contribution of the most prominent downstream translational components of mTOR signaling toward Akt-driven lymphomagenesis. While phosphorylation of rpS6 is dispensable for cancer formation, 4EBP-eIF4E exerts significant control over cap-dependent translation, cell growth, cancer initiation, and progression. This effect is mediated at least in part through 4EBP-dependent control of Mcl-1 expression, a key antiapoptotic protein. By using an active site inhibitor of mTOR, PP242, we show a marked therapeutic response in rapamycin-resistant tumors. The therapeutic benefit of PP242 is mediated through inhibition of mTORC1-dependent 4EBP-eIF4E hyperactivation. Thus, the 4EBP-eIF4E axis downstream of mTOR is a druggable mediator of translational control and Akt-mediated tumorigenesis that has important implications for the treatment of human cancers.
View details for DOI 10.1016/j.ccr.2010.01.021
View details for Web of Science ID 000275804200007
View details for PubMedID 20227039
Suppression of Myc oncogenic activity by ribosomal protein haploinsufficiency
2008; 456 (7224): 971-U79
In human thyroid, caveolin-1 is localized at the apex of thyrocytes, but its role there remains unknown. Using immunohistochemistry, (127)I imaging, transmission electron microscopy, immunogold electron microscopy, and quantification of H(2)O(2), we found that in caveolin-1 knockout mice thyroid cell homeostasis was disrupted, with evidence of oxidative stress, cell damage, and apoptosis. An even more striking phenotype was the absence of thyroglobulin and iodine in one-half of the follicular lumina and their presence in the cytosol, suggesting that the iodide organification and binding to thyroglobulin were intracellular rather than at the apical membrane/extracellular colloid interface. The latter abnormality may be secondary to the observed mislocalization of the thyroid hormone synthesis machinery (dual oxidases, thyroperoxidase) in the cytosol. Nevertheless, the overall uptake of radioiodide, its organification, and secretion as thyroid hormones were comparable to those of wild-type mice, suggesting adequate compensation by the normal TSH retrocontrol. Accordingly, the levels of free thyroxine and TSH were normal. Only the levels of free triiodothyronine showed a slight decrease in caveolin-1 knockout mice. However, when TSH levels were increased through low-iodine chow and sodium perchlorate, the induced goiter was more prominent in caveolin-1 knockout mice. We conclude that caveolin-1 plays a role in proper thyroid hormone synthesis as well as in cell number homeostasis. Our study demonstrates for the first time a physiological function of caveolin-1 in the thyroid gland. Because the expression and subcellular localization of caveolin-1 were similar between normal human and murine thyroids, our findings in caveolin-1 knockout mice may have direct relevance to the human counterpart.
View details for DOI 10.1152/ajpendo.90784.2008
View details for Web of Science ID 000268252700019
View details for PubMedID 19435853
Roles of hydrogen peroxide in thyroid physiology and disease
JOURNAL OF CLINICAL ENDOCRINOLOGY & METABOLISM
2007; 92 (10): 3764-3773
The Myc oncogene regulates the expression of several components of the protein synthetic machinery, including ribosomal proteins, initiation factors of translation, RNA polymerase III and ribosomal DNA. Whether and how increasing the cellular protein synthesis capacity affects the multistep process leading to cancer remains to be addressed. Here we use ribosomal protein heterozygote mice as a genetic tool to restore increased protein synthesis in Emu-Myc/+ transgenic mice to normal levels, and show that the oncogenic potential of Myc in this context is suppressed. Our findings demonstrate that the ability of Myc to increase protein synthesis directly augments cell size and is sufficient to accelerate cell cycle progression independently of known cell cycle targets transcriptionally regulated by Myc. In addition, when protein synthesis is restored to normal levels, Myc-overexpressing precancerous cells are more efficiently eliminated by programmed cell death. Our findings reveal a new mechanism that links increases in general protein synthesis rates downstream of an oncogenic signal to a specific molecular impairment in the modality of translation initiation used to regulate the expression of selective messenger RNAs. We show that an aberrant increase in cap-dependent translation downstream of Myc hyperactivation specifically impairs the translational switch to internal ribosomal entry site (IRES)-dependent translation that is required for accurate mitotic progression. Failure of this translational switch results in reduced mitotic-specific expression of the endogenous IRES-dependent form of Cdk11 (also known as Cdc2l and PITSLRE), which leads to cytokinesis defects and is associated with increased centrosome numbers and genome instability in Emu-Myc/+ mice. When accurate translational control is re-established in Emu-Myc/+ mice, genome instability is suppressed. Our findings demonstrate how perturbations in translational control provide a highly specific outcome for gene expression, genome stability and cancer initiation that have important implications for understanding the molecular mechanism of cancer formation at the post-genomic level.
View details for DOI 10.1038/nature07449
View details for Web of Science ID 000261768300050
View details for PubMedID 19011615
Reciprocal negative regulation between thyrotropin/3 ',5 '-cyclic adenosine monophosphate-mediated proliferation and caveolin-1 expression in human and murine thyrocytes
2007; 21 (4): 921-932
The long-lived thyroid cell generates, for the synthesis of thyroid hormones, important amounts of H2O2 that are toxic in other cell types. This review analyzes the protection mechanisms of the cell and the pathological consequences of disorders of this system.The literature on H2O2 generation and disposal, thyroid hormone synthesis, and their control in the human thyroid is analyzed.In humans, H2O2 production by dual-oxidases and consequently thyroid hormone synthesis by thyroperoxidase are controlled by the phospholipase C-Ca2+-diacylglycerol arm of TSH receptor action. H2O2 in various cell types, and presumably in thyroid cells, is a signal, a mitogen, a mutagen, a carcinogen, and a killer. The various protection mechanisms of the thyroid cell against H2O2 are analyzed. They include the separation of the generating enzymes (dual-oxidases), their coupling to thyroperoxidase in a proposed complex, the thyroxisome, and H2O2 degradation systems.It is proposed that various pathologies can be explained, at least in part, by overproduction and lack of degradation of H2O2 (tumorigenesis, myxedematous cretinism, and thyroiditis) and by failure of the H2O2 generation or its positive control system (congenital hypothyroidism).
View details for DOI 10.1210/jc.2007-0660
View details for Web of Science ID 000250148400003
View details for PubMedID 17666482
Mutated E-cadherin: Genomic and functional characterization in thyroid cells from the KAT family
2004; 14 (11): 902-909
The expression of caveolins is down-regulated in tissue samples of human thyroid autonomous adenomas and in the animal model of this disease. Because several cell types present in thyroid express caveolins, it remained unclear if this down-regulation occurs in thyrocytes and which are the mechanism and role of this down-regulation in the tumor context. Here we show that prolonged stimulation of isolated human thyrocytes by TSH/cAMP/cAMP-dependent protein kinase inhibits caveolins' expression. The expression of caveolins is not down-regulated by activators of other signaling pathways relevant to thyroid growth/function. Therefore, the down-regulation of caveolins' expression in autonomous adenomas is a direct consequence of the chronic activation of the TSH/cAMP pathway in thyrocytes. The down-regulation of caveolin-1 occurs at the mRNA level, with a consequent protein decrease. TSH/cAMP induces a transcription-dependent, translation-independent destabilization of the caveolin-1 mRNA. This effect is correlated to the known proliferative role of that cascade in thyrocytes. In vivo, thyrocytes of caveolin-1 knockout mice display enhanced proliferation. This demonstrates, for the first time, the in vivo significance of the specific caveolin-1 down-regulation by one mitogenic cascade and its relation to a human disease.
View details for DOI 10.1210/me.2006-0328
View details for Web of Science ID 000245227100011
View details for PubMedID 17202321
Sphingolipid-cholesterol domains (lipid rafts) in normal human and dog thyroid follicular cells are not involved in thyrotropin receptor signaling
2004; 145 (3): 1464-1472
Members of a family of thyroid cell lines (KAT) were analyzed because they expressed a higher molecular weight (135 kd) form of E-cadherin at their surface. We found that this aberrant E-cadherin is the result of a point mutation in the exon 9 donor splice site causing a skipping of exon 9 with consequent deletion of the corresponding aminoacids on E-cadherin protein. As a spin-off, we report that the various members of the KAT family share this mutation as well as the genetic background. Furthermore we found that this mutated protein leads to disturbed cell-cell adhesion although E-cadherin is still able to mediate the formation of the cadherin/ catenin complex. We also demonstrate the presence of another cell-cell adhesion complex, formed by Pcadherin and the catenins. The latter is also not able to mediate cell-cell adhesion. Although these cells lack cell-cell adhesion they are not invasive without exogenous stimulus.
View details for Web of Science ID 000225651100005
View details for PubMedID 15671768
Kinetics of thyrotropin-stimulating hormone (TSH) and thyroid-stimulating antibody binding and action on the TSH receptor in intact TSH receptor-expressing CHO cells
JOURNAL OF CLINICAL ENDOCRINOLOGY & METABOLISM
2003; 88 (11): 5366-5374
Partition of signaling molecules in sphingolipid-cholesterol-enriched membrane domains, among which are the caveolae, may contribute to signal transduction efficiency. In normal thyroid, nothing is known about a putative TSH/cAMP cascade compartmentation in caveolae or other sphingolipid-cholesterol-enriched membrane domains. In this study we show for the first time that caveolae are present in the apical membrane of dog and human thyrocytes: caveolin-1 mRNA presence is demonstrated by Northern blotting in primary cultures and that of the caveolin-1 protein by immunohistochemistry performed on human thyroid tissue. The TSH receptor located in the basal membrane can therefore not be located in caveolae. We demonstrate for the first time by biochemical methods the existence of sphingolipid-cholesterol-enriched domains in human and dog thyroid follicular cells that contain caveolin, flotillin-2, and the insulin receptor. We assessed a possible sphingolipid-cholesterol-enriched domains compartmentation of the TSH receptor and the alpha- subunit of the heterotrimeric G(s) and G(q) proteins using two approaches: Western blotting on detergent-resistant membranes isolated from thyrocytes in primary cultures and the influence of 10 mm methyl-beta-cyclodextrin, a cholesterol chelator, on basal and stimulated cAMP accumulation in intact thyrocytes. The results from both types of experiments strongly suggest that the TSH/cAMP cascade in thyroid cells is not associated with sphingolipid-cholesterol-enriched membrane domains.
View details for DOI 10.1210/en.2003-1432
View details for Web of Science ID 000189035500058
View details for PubMedID 14670987
Immunohistochemical study of heat shock proteins 27, 60 and 70 in the normal human adrenal and in adrenal tumors with suppressed ACTH production
MICROSCOPY RESEARCH AND TECHNIQUE
2003; 61 (3): 315-323
The kinetics of TSH binding and the effects of TSH and thyroid-stimulating antibody (TSAb) on cAMP accumulation have been measured in TSH receptor-expressing CHO cells (CHO-TSHR cells). The parallel kinetics of TSH binding to its receptor and of cell cAMP concentration after the addition and withdrawal of TSH show that in the case of this receptor, signal generation and concentration are at all times proportional to occupancy. In physiological ionic medium, TSAb, but not TSH, action is slowed and in some cases almost nonexistent. The kinetics of cAMP disappearance after washout of TSAb is also slower. cAMP accumulation is faster for Fabs than for the TSAb from which they derive. Analysis of the data suggest that 1) serum TSAb are oligoclonal antibodies sets, at low concentrations, with a high affinity for the TSH receptor; 2) ionic interactions are involved in the action of TSAb on the TSH receptor; and 3) TSAb activation of the TSH receptor is at least a two-step process. Among others, a possible explanation is that the full activation of the receptor requires the binding of two or more different antibody molecules on different sites of the same TSH receptor. This analysis provides a benchmark for studies of experimentally induced monoclonal antibodies activating the TSH receptor.
View details for DOI 10.1210/jc.2003-030664
View details for Web of Science ID 000186393900048
View details for PubMedID 14602775
Oncogenic mutations in the thyrotropin receptor of autonomously functioning thyroid nodules in the Japanese population
EUROPEAN JOURNAL OF ENDOCRINOLOGY
2002; 147 (3): 287-291
Heat shock proteins (HSPs) are known to protect cells against various aggressions and to assist in the correct folding of nascent proteins as well as in the recovery of denatured ones. HSP70 increases its levels in the cell in response to any stress and is induced by ACTH in the adrenal gland. HSP60 is located in the mitochondria and assists in the folding of mitochondrial peptides. HSP27 is the only small HSP that is stress-induced. HSP27 and HSP70 are known to protect cells against apoptosis while, on the contrary, HSP60 is proapoptotic, increasing caspases maturation. We studied the expression of these HSPs in human adrenal tissue both in the normal glands (12 cases) and in tumoral tissue from cortisol producing adrenal adenomas (6 cases). Besides being neoplastic, these cells live in a particular ambience of lack of ACTH due to the suppression of the hypothalamic-pituitary ACTH secretion induced by the elevated levels of cortisol. HSP27 is highly expressed in the normal adrenal and shows a marked reduction of expression in Cushing's adrenal tissue. Although with overall lower levels of expression in the normal adrenal, HSP70 exhibited a similar pattern of reduction in tumoral tissue. HSP60, on the other hand, increased significantly and consistently in adrenal Cushing tumors. Besides the possible consequences of incorrect folding of nascent peptides, the alterations observed in tumoral tissue seem to act in an apoptotic direction. The only factor that we observed that could be contributing to these changes was the lack of plasma ACTH.
View details for DOI 10.1002/jemt.10341
View details for Web of Science ID 000183120200012
View details for PubMedID 12768547
Constitutively activating mutations of the thyrotropin receptor (TSHR) have been found in the majority of autonomously functioning thyroid nodules (AFTNs) in European patients. The reported frequency of these mutations varies among reports but amounts to 50-80%. To date, only one such mutation responsible for AFTNs has been identified in the Japanese population and the pathogenic role of such mutations in Japanese AFTNs has been questioned. In the present study, we evaluated the frequency of activating mutations in the TSHR and G(alpha)s in 10 Japanese AFTNs.Genomic DNA was extracted from fresh frozen tissue. The TSHR and the almost entire sequence of the gene coding for the alpha subunit of Gs have been amplified and sequenced.In sequence analysis, four mutations in the TSHR (T632A, I486M, M453T and L512R) were found. To complete our analysis, we searched mutations in the gene coding for the alpha subunit of Gs, in the samples negative for TSHR mutations. In one case a mutation (R201H) affecting GTPase activity was found.If we focus on the solitary nodules, we obtain the same mutation proportion as in European patients (70%). The absence of TSHR and G(alpha)s mutations in a significant proportion of autonomous adenomas in multinodular goiters suggests that other causes may also play a role in the genesis of these lesions.
View details for Web of Science ID 000178743900002
View details for PubMedID 12213664