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Results for “ectopic gene conversion”

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PubMed2005

1alpha,25-Dihydroxyvitamin D3 regulates the expression of Id1 and Id2 genes and the angiogenic phenotype of human colon carcinoma cells.

1alpha,25-Dihydroxyvitamin D3 (1alpha,25(OH)2D3) has antitumor activity in addition to its classical action on calcium metabolism and bone tissue biology. It is thought to regulate the expression of multiple target genes and thus modulate processes critical for tumor growth and metastases. Here we show that 1alpha,25(OH)2D3 differentially regulates the expression of Id1 and Id2 genes, members of a family of transcriptional regulators of cell proliferation and differentiation. 1alpha,25(OH)2D3 induced epithelial differentiation in SW480-ADH human colon carcinoma cell line by promoting expression of the proteins implicated in adherent junction formation, including E-cadherin, and by inhibiting beta-catenin transcriptional activity. 1alpha,25(OH)2D3 activated the human Id1 gene promoter and rapidly induced Id1 RNA and protein. Ectopic overexpression of Id1 was not sufficient to induce E-cadherin, which was critical for the morphological changes induced by 1alpha,25(OH)2D3 in SW480-ADH cells. Conversely, Id2 transcription rate, RNA and protein levels were decreased by 1alpha,25(OH)2D3. Id2 downregulation by 1alpha,25(OH)2D3 mediated the antiproliferative effect of 1alpha,25(OH)2D3 on SW480-ADH cells. In addition, we showed that 1alpha,25(OH)2D3 changed the levels of the inducer of angiogenesis, vascular endothelial growth factor and the potent antiangiogenic factor thrombospondin-1, leading to a balanced change in the angiogenic potential of SW480-ADH human colon carcinoma cells.

Animals↗

PubMed2001

The Caenorhabditis elegans gene lin-26 can trigger epithelial differentiation without conferring tissue specificity.

How epithelial cell fates become specified is poorly understood. We have previously shown that the putative C2H2 zinc-finger transcription factor LIN-26 is required for the differentiation of ectodermal and mesodermal epithelial cells in Caenorhabditis elegans. Here, we report that ectopic LIN-26 expression during early gastrulation transforms most blastomeres into epithelial-like cells. Specifically, LIN-26 induced the expression of three epithelial markers: the adherens junction protein JAM-1; DLG-1, which is essential for the assembly of JAM-1 at junctions; and CHE-14, which is involved in apical trafficking. Furthermore, ultrastructural studies revealed that ectopic LIN-26 expression induced the formation of adherens-like junctions. However, ectopic lin-26 expression did not confer any tissue-specific cell fate, such as the epidermal cell fate, as evidenced from the observation that several epidermal-specific genes were not induced. Conversely, we show that epidermal cells displayed some polarity defects in lin-26 mutants. We conclude that lin-26 can induce epithelial differentiation and that epitheliogenesis is not a default pathway in C. elegans.

Animals↗

PubMed1996

Regulation and function of the terminal gap gene huckebein in the Drosophila blastoderm.

Pattern formation in Drosophila involves a cascade of maternal and zygotic factors which are spatially restricted in the blastoderm embryo. Here we show that the Drosophila gene huckebein (hkb), a member of the gap gene class of segmentation genes, is not only required for suppression of segmentation in the terminal regions of the embryo but also to spatially restrict germ layer formation at the beginning of gastrulation. hkb encodes a Sp1/egr-like zinc finger protein, likely to be a transcription factor. Its absence in hkb mutants causes the ectodermal and mesodermal primordia to expand at the expense of endoderm anlagen, which are completely absent in null alleles of hkb. Conversely, ectopic expression of hkb inhibits the formation of the major gastrulation fold which gives rise to the mesoderm and prevents normal segmentation in the ectoderm of the trunk region.

Animals↗

PubMed2000

Prolactin enhances CCAAT enhancer-binding protein-beta (C/EBP beta) and peroxisome proliferator-activated receptor gamma (PPAR gamma) messenger RNA expression and stimulates adipogenic conversion of NIH-3T3 cells.

Extracellular stimuli trigger adipocyte differentiation by inducing the complex cascades of transcription. Transcription factors CCAAT enhancer-binding proteins (C/EBPs) and peroxisome proliferator-activated receptor gamma (PPARgamma) play crucial roles in this process. Although ectopic expression of these factors in NIH-3T3 cells, a multipotential mesenchymal stem cell line, results in adipogenic conversion, little is known as to hormonal factors that regulate adipogenesis in these cells. In this report we demonstrate that PRL, a lactogenic hormone, enhances C/EBPbeta and PPARbeta mRNA expression and augments adipogenic conversion of NIH-3T3 cells. Moreover, we show that ectopic expression of the PRL receptor in NIH-3T3 cells results in efficient adipocyte conversion when stimulated with PRL and a PPARgamma ligand, as evidenced by expression of the adipocyte differentiation-specific genes as well as the presence of fat-laden cells. We further demonstrate that signal transducer and activator of transcription 5 (Stat5), a PRL signal transducer, activates aP2 promoter in a PRL-dependent manner. These results suggest that PRL acts as an adipogenesis-enhancing hormone in NIH-3T3 cells.

3T3 Cells↗

PubMed1999

PPARgamma ligand-dependent induction of STAT1, STAT5A, and STAT5B during adipogenesis.

We have recently demonstrated that STAT1, STAT5A, and STAT5B are induced during adipogenesis of cultured preadipocytes in a differentiation-dependent manner. Members of the C/EBP and PPAR families of transcription factors have also been shown to be induced during adipocyte differentiation and to play a significant role in the regulation of fat-specific genes. In this investigation, we have examined the ability of C/EBPs and PPARs to contribute to STAT protein expression during conversion of non-precursor fibroblasts to functionally mature adipocytes. For this study, NIH-3T3 fibroblasts engineered to ectopically co-express C/EBPbeta and C/EBPdelta under the control of a tetracycline-responsive, inducible expression system were utilized to assess STAT expression during controlled adipogenesis. Data presented here demonstrate that STAT1, STAT5A, and STAT5B, but not STAT3 and STAT6, were induced in a tetracycline-responsive manner during the differentiation of these engineered fibroblasts. The STAT protein accumulation resulting from C/EBP expression was tightly coupled to the morphological conversion of fibroblasts to adipocytes and represents an expression profile identical to that reported for mature adipocytes in vivo. Data are also presented demonstrating that STAT protein accumulation and adipocyte conversion occurred only during controlled conditions leading to the expression of PPARgamma and that the expression of these three STATs was tightly regulated in a PPARgamma ligand dose-response fashion. These data illustrate that the cascade of transcriptional events leading to adipogenesis regulate the STAT family of transcription factors and that the differentiation-dependent upregulation of STAT protein expression is regulated downstream of PPARgamma in a ligand-dependent manner.

3T3 Cells↗

PubMed2005

FGF19-FGFR4 signaling elaborates lens induction with the FGF8-L-Maf cascade in the chick embryo.

The fibroblast growth factor (FGF) family is known to be involved in vertebrate eye development. However, distinct roles of individual FGF members during eye development remain largely elusive. Here, we show a detailed expression pattern of Fgf19 in chick lens development. Fgf19 expression initiated in the forebrain, and then became restricted to the distal portion of the optic vesicle abutting the future lens placode, where FGF receptor 4 (Fgfr4), a receptor for FGF19, was expressed. Fgf8, a positive regulator for L-Maf, was expressed in a portion of the optic vesicle. To examine the role of FGF19 signaling during early eye development, Fgf19 was misexpressed near the presumptive lens ectoderm; however, no alteration in the expression of lens marker genes was observed. Conversely, a secreted form of FGFR4 was misexpressed to inhibit an FGF19 signal, resulting in the induction of L-Maf expression. To further define the relationship between L-Maf and Fgf19, L-Maf misexpression was performed, resulting in ectopic induction of Fgf19 expression by Hamburger and Hamilton's stage 12/13. Furthermore, misexpression of Fgf8 induced Fgf19 expression in addition to L-Maf. These results suggest that FGF19-FGFR4 signaling plays a role in early lens development in collaboration with FGF8 signaling and L-Maf transcriptional system.

Animals↗

PubMed2000

RNA interference demonstrates a role for nautilus in the myogenic conversion of Schneider cells by daughterless.

Schneider SL2 cells activate the myogenic program in response to the ectopic expression of daughterless alone, as indicated by exit from the cell cycle, syncytia formation, and the presence of muscle myosin fibrils. Myogenic conversion can be potentiated by the coexpression of DMEF2 and nautilus with daughterless. In RT-PCR assays Schneider cells express two mesodermal markers, nautilus and DMEF2 mRNAs, as well as very low levels of daughterless mRNA but no twist. Full-length RT-PCR products for nautilus and DMEF2 encode immunoprecipitable proteins. We used RNA-i to demonstrate that both endogenous nautilus expression and DMEF2 expression are required for the myogenic conversion of Schneider cells by daughterless. Coexpression of twist blocks conversion by daughterless but twist dsRNA has no effect. Our results indicate that Schneider cells are of mesodermal origin and that myogenic conversion with ectopic expression of daughterless occurs by raising the levels of daughterless protein sufficiently to allow the formation of nautilus/daughterless heterodimers. The effectiveness of RNA-i is dependent upon protein half-life. Genes encoding proteins with relatively short half-lives (10 h), such as nautilus or HSF, are efficiently silenced, whereas more stable proteins, such as cytoplasmic actin or beta-galactosidase, are less amenable to the application of RNA-i. These results support the conclusion that nautilus is a myogenic factor in Drosophila tissue culture cells with a functional role similar to that of vertebrate MyoD. This is discussed with regard to the in vivo functions of nautilus.

Animals↗

PubMed2005

An Arabidopsis aspartic protease functions as an anti-cell-death component in reproduction and embryogenesis.

The components and pathways that regulate and execute developmental cell death programmes in plants remain largely unknown. We have found that the PROMOTION OF CELL SURVIVAL 1 (PCS1) gene in Arabidopsis, which encodes an aspartic protease, has an important role in determining the fate of cells in embryonic development and in reproduction processes. The loss-of-function mutation of PCS1 causes degeneration of both male and female gametophytes and excessive cell death of developing embryos. Conversely, ectopic expression of PCS1 causes the septum and stomium cells that normally die in the anther wall to survive instead, leading to a failure in anther dehiscence and male sterility. PCS1 provides a new avenue for understanding the mechanisms of the programmed cell death processes that are associated with developmental pathways in plants and makes available a useful tool for engineering the male sterility trait for hybrid seed production.

Arabidopsis↗

PubMed1998

XCoe2, a transcription factor of the Col/Olf-1/EBF family involved in the specification of primary neurons in Xenopus.

BACKGROUND: Primary neurogenesis in Xenopus is a model for studying the control of neural cell fate decisions. The specification of primary neurons appears to be driven by transcription factors containing a basic region and a helix-loop-helix (HLH) motif: expression of Xenopus neurogenin-related-1 (X-ngnr-1) defines the three prospective domains of primary neurogenesis, and expression of XNeuroD coincides with neuronal differentiation. The transition between neuronal competence and stable commitment to a neuronal fate remains poorly characterised, however. RESULTS: Drosophila Collier and rodent early B-cell factor/olfactory-1 define a family of HLH transcription factors containing a previously unknown type of DNA-binding domain. We isolated an orthologous gene from Xenopus, Xcoe2, which is expressed in precursors of primary neurons. Xcoe2 is transcribed after X-ngnr-1 and before XNeuroD. Overexpression of a dominant-negative mutant of XCoe2 prevented neuronal differentiation. Conversely, overexpressed wild-type Xcoe2 could promote ectopic differentiation of neurons, in both the neural plate and the epidermis. In contrast to studies with X-ngnr-1 or XNeuroD, the supernumerary neurons induced by Xcoe2 appeared in a 'salt-and-pepper' pattern, resulting from the activation of X-Delta1 expression and feedback regulation by lateral inhibition. CONCLUSIONS: XCoe2 may play a pivotal role in the transcriptional cascade that specifies primary neurons in Xenopus embryos: by maintaining Delta-Notch signalling, XCoe2 stabilises the higher neural potential of selected progenitor cells that express X-ngnr-1, ensuring the transition between neural competence and irreversible commitment to a neural fate; and it promotes neuronal differentiation by activating XNeuroD expression, directly or indirectly.

Amino Acid Sequence↗

PubMed1994

Sp1/egr-like zinc-finger protein required for endoderm specification and germ-layer formation in Drosophila.

Much of our present knowledge of the biological processes involved in pattern formation in Drosophila is derived from segmentation analysis. Comparatively little is known about the genetic requirement and mechanisms underlying the formation and separation of germ layers by morphogenetic movements during gastrulation. Here we show that the Drosophila gene huckebein (hkb), a member of the gap-gene class of segmentation genes, is required for germ-layer formation at blastoderm. Absence of the hkb product, an Sp1/egr-like zinc-finger protein, causes the ectodermal and mesodermal primordia to expand at the expense of endoderm anlagen. Conversely, ectopic expression of hkb inhibits the formation of the major gastrulation fold which gives rise to the mesoderm and prevents normal segmentation in the ectoderm. Thus, hkb is necessary for endoderm development and its activity defines spatial limits within the blastoderm embryo in which the germ layers are established.

Amino Acid Sequence↗

PubMed2003

Troglitazone antagonizes tumor necrosis factor-alpha-induced reprogramming of adipocyte gene expression by inhibiting the transcriptional regulatory functions of NF-kappaB.

Troglitazone (TGZ), a member of the thiazolidinedione class of anti-diabetic compounds and a peroxisome proliferator activator receptor-gamma (PPAR-gamma) agonist, restores systemic insulin sensitivity and improves the full insulin resistance syndrome in vivo. The mechanisms underlying its in vivo function are not understood. Here we investigated the potential functional interaction between PPAR-gamma and NF-kappaB in adipocytes. We show that TGZ selectively blocked tumor necrosis factor-alpha-induced and NF-kappaB-dependent repression of multiple adipocyte-specific genes and induction of growth phase and other genes. This occurs without interfering with NF-kappaB expression, activation, nuclear translocation, or DNA binding and without suppressing NF-kappaB-dependent survival signals. Notably, the expressions of some tumor necrosis factor-alpha-induced genes in adipocytes were unaffected by PPAR-gamma activation. In reporter gene assays in HeLa cells, ectopic expression of PPAR-gamma abolished induction of a NF-kappaB-responsive reporter gene by the p65 subunit (RelA) of NF-kappaB, and the inhibition was further enhanced in the presence of TGZ. Conversely, overexpression of p65 inhibited induction of a PPAR-gamma-responsive reporter gene by activated PPAR-gamma in a dose-dependent manner. The inhibitory effect was independent of the presence of NF-kappaB-binding sites in the promoter region. Other NF-kappaB family members, p50 and c-Rel as well as the S276A mutant of p65, blocked PPAR-gamma-mediated gene transcription less effectively. Thus, p65 antagonizes the transcriptional regulatory activity of PPAR-gamma in adipocytes, and PPAR-gamma activation can at least partially override the inhibitory effects of p65 on the expression of key adipocyte genes. Our data suggest that inhibition of NF-kappaB activity is a mechanism by which PPAR-gamma agonists improve insulin sensitivity in vivo and that adipocyte NF-kappaB is a potential therapeutic target for obesity-linked type 2 diabetes.

3T3 Cells↗

PubMed2006

Characterization of liver function in transdifferentiated hepatocytes.

We previously demonstrated that dexamethasone (Dex) induces the transdifferentiation (or conversion) of the pancreatic progenitor cell line AR42J-B13 (B13) to hepatocytes based on the expression of liver proteins. We have extended our original observations to determine: (1) the effects of Dex on pancreatic gene expression; (2) the time course of expression of liver enriched transcription factors during conversion from pancreatic to hepatic phenotype; (3) the functional potential of transdifferentiated hepatocytes; (4) the proliferative capacity of transdifferentiated hepatocytes; and (5) whether ectopic expression of transcription factors can induce the hepatic phenotype in pancreatic B13 cells. The results were as follows. The B13 cell markers amylase, synaptophysin, and neurofilament were lost in transdifferentiated hepatocytes compared to control cells and the liver enriched transcription factors C/EBPbeta and C/EBPalpha were induced first, followed by HNF4alpha and then RXRalpha. Using RT-PCR analysis and immunolocalisation studies, we detected hepatic markers (e.g., apolipoprotein B) in Dex-treated cells. In transdifferentiated hepatocytes albumin was secreted, insulin stimulated lipid deposition and ciprofibrate enhanced the expression of catalase. Proliferation of transdifferentiated hepatocytes is promoted in the presence of HGF and NEAA as indicated by the co-expression of the cell cycle markers cyclin D and phosphohistone H3 with liver proteins. Lastly, ectopic expression of C/EBPalpha or C/EBPbeta in AR42J-B13 cells was sufficient to induce transdifferentiation, based on nuclear localization of HNF4alpha and induction of UDP-glucuronosyltransferase expression. These results indicate that the B13 progenitor cell model is suitable for studying liver function and for understanding the molecular and cellular events that occur during transdifferentiation.

Albumins↗

PubMed2006

Antiretroviral potential of human tripartite motif-5 and related proteins.

TRIM5alpha is a potent inhibitor of infection by diverse retroviruses and is encoded by one of a large family of TRIM genes. We found that several TRIM motifs among a panel of selected human TRIM proteins (TRIM1, 5, 6, 18, 19, 21 22, 34) could inhibit infection when artificially targeted to an incoming HIV-1 capsid. Conversely, when ectopically expressed as authentic full-length proteins, most lacked activity against a panel of retroviruses. The exceptions were TRIM1, TRIM5 and TRIM34 proteins. Weak but specific inhibition of HIV-2/SIV(MAC) and EIAV by TRIM34 was noted, and human TRIM5alpha modestly, but specifically, inhibited an HIV-1 strain carrying a mutation in the cyclophilin binding loop (G89V). Restriction activity observed in ectopic expression assays was sometimes not detectable in corresponding RNAi-based knockdown experiments. However, endogenous owl monkey TRIMCyp potently inhibited an SIV(AGM) strain. Overall, sporadic examples of intrinsic antiretroviral activity exist in this panel of TRIM proteins.

Adaptor Proteins, Signal Transducing↗

PubMed2006

Depletion of cAMP-response element-binding protein/ATF1 inhibits adipogenic conversion of 3T3-L1 cells ectopically expressing CCAAT/enhancer-binding protein (C/EBP) alpha, C/EBP beta, or PPAR gamma 2.

The differentiation of preadipocytes to adipocytes is orchestrated by the expression of the "master adipogenic regulators," CCAAT/enhancer-binding protein (C/EBP) beta, peroxisome proliferator-activated receptor gamma (PPARgamma), and C/EBP alpha. In addition, activation of the cAMP-response element-binding protein (CREB) is necessary and sufficient to promote adipogenic conversion and prevent apoptosis of mature adipocytes. In this report we used small interfering RNA to deplete CREB and the closely related factor ATF1 to explore the ability of the master adipogenic regulators to promote adipogenesis in the absence of CREB and probe the function of CREB in late stages of adipogenesis. Loss of CREB/ATF1 blocked adipogenic conversion of 3T3-L1 cells in culture or 3T3-F442A cells implanted into athymic mice. Loss of CREB/ATF1 prevented the expression of PPARgamma, C/EBP alpha, and adiponectin and inhibited the loss of Pref-1. Loss of CREB/ATF1 inhibited adipogenic conversion even in cells ectopically expressing C/EBP alpha, C/EBP beta, or PPARgamma2 individually. CREB/ATF1 depletion did not attenuate lipid accumulation in cells expressing both PPARgamma2 and C/EBP alpha, but adiponectin expression was severely diminished. Conversely ectopic expression of constitutively active CREB overcame the blockade of adipogenesis due to depletion of C/EBP beta but not due to loss of PPARgamma2 or C/EBP alpha. Depletion of CREB/ATF1 did not suppress the expression of C/EBP beta as we had previously observed using dominant negative forms of CREB. Finally results are presented showing that CREB promotes PPARgamma2 gene transcription. The results indicate that CREB and ATF1 play a central role in adipogenesis because expression of individual master adipogenic regulators is unable to compensate for their loss. The data also indicate that CREB not only functions during the initiation of adipogenic conversion but also at later stages.

3T3-L1 Cells↗

PubMed2002

Drosophila HB9 is expressed in a subset of motoneurons and interneurons, where it regulates gene expression and axon pathfinding.

Motoneurons are an essential component of all metazoan nervous systems, but it is unknown whether there is an evolutionarily conserved mechanism for generating motoneurons during neurogenesis. In the vertebrate CNS, HB9/MNR2 transcription factors are specifically expressed in all somatic motoneurons and are necessary to distinguish motoneurons from interneurons, in part by repressing interneuron-specific gene expression. Here, we identify and characterize the single Drosophila ortholog of the HB9/MNR2 gene family. Drosophila HB9 is detected in a subset of motoneurons with ventral muscle targets and in a small group of interneurons, including the well characterized serotonergic interneurons. RNA interference knockdown of HB9 levels leads to defects in motoneuron ventral muscle target recognition, ectopic expression of a marker for dorsally projecting motoneurons (Even-skipped), and defects in serotonergic interneuronal projections. Conversely, ectopic HB9 expression causes an expansion of ventral motoneuron projections and repression of Even-skipped. Thus, Drosophila HB9 is required in a subset of motoneurons and interneurons for establishing proper axon projections but does not have a general role in distinguishing motoneuron and interneuron cell types.

Animals↗

PubMed2001

A MADS box gene from lily (Lilium Longiflorum) is sufficient to generate dominant negative mutation by interacting with PISTILLATA (PI) in Arabidopsis thaliana.

Lily MADS box gene 1 (LMADS1), with sequence homology to the AP3 family of genes, was cloned and characterized from lily (Lilium longiflorum). LMADS1 protein contains almost complete consensus sequence of the PISTILLATA (PI)-derived motif (YEFRVQPSQPNLH) found in the AP3 family of genes and paleoAP3 motif (YGSHDLRLA) found in the AP3 family of genes from the low eudicot, magnolid dicot and monocot species. LMADS1 mRNA was expressed in all four whorls of the flower and absent in the vegetative leaves. The LMADS1 protein was only detected in the petals and stamens, indicating that LMADS1 is possibly post-transcriptionally regulated in lily. Arabidopsis plants transformed with 35S::LMADS1 produced flowers with short petals and stamens, however, no floral organ conversion was observed. Ectopic expression of LMADS1 cDNA truncated with the MADS box domain in Arabidopsis generated the ap3-like dominant negative mutation in which the petals were converted into sepal-like structures and the stamens were converted into carpel-like structures. Yeast two-hybrid analysis indicated that LMADS1 truncated with the MADS box domain is able to sufficiently interact with the Arabidopsis PI protein. This result supports that LMADS1 is the functional counterpart of the AP3 gene in lily. Interestingly, in contrast to other B functional genes, LMADS1 truncated with the MADS box domain is able to strongly form homodimers. LMADS1 may represent an ancestral form of the B function gene, which retains the ability to form homodimers in regulating petal and stamen development in lily.

Amino Acid Sequence↗

PubMed2026

Long noncoding RNA GAS5 disrupts intestinal epithelial barrier function by increasing small vault RNA levels.

Disruptions in the integrity of the intestinal epithelium occur commonly in inflammatory bowel disease (IBD) and critical surgical disorders, but the underlying mechanisms remain largely unknown. Here we identified long noncoding RNA GAS5 as a repressor of intestinal mucosal growth and the function of the gut epithelial barrier. The levels of tissue GAS5/Gas5 increased in mouse intestinal mucosa after colitis and septic stress, as well as in human intestinal mucosa from patients with IBD. Transient and tissue-specific knockdown of Gas5 in mice using CRISPR/Cas9 enhanced the renewal of the mucosa of the small intestine, increased the levels of tight junction (TJ) proteins ZO-1, ZO-2, claudin-1, and claudin-2, and improved gut barrier function. Conversely, ectopic overexpression of GAS5 in intestinal organoids and in cultured intestinal epithelium cells decreased the levels of these TJ proteins and caused epithelial barrier dysfunction. Mechanistic studies revealed that GAS5 acted as a transcriptional enhancer of the gene (2. AUTHOR: Do you mean "genes"?) encoding small noncoding vault RNAs (vtRNAs) and that GAS5 repressed TJ expression by increasing the levels of vtRNAs. Together, our results indicate that GAS5 disrupts the integrity of the intestinal epithelium by impairing mucosal growth and epithelial barrier function and that it represses TJ expression, at least in part, via vtRNAs.

Animals↗

PubMed1999

Role of PPAR gamma in regulating adipocyte differentiation and insulin-responsive glucose uptake.

Adipocyte differentiation is regulated by at least two families of transcription factors, CCAAT/enhancer binding proteins (C/EBPs) and peroxisome proliferator-activated receptors (PPARs). Induction of PPAR gamma gene transcription during the differentiation of preadipocytes into adipocytes in vitro occurs following an initial phase of cell proliferation and requires a direct involvement of C/EBP beta, C/EBP delta, and glucocorticoids. Ectopic expression of PPAR gamma in non-adipogenic, Swiss 3T3 fibroblasts promotes their conversion into adipocytes as indicated by the accumulation of lipid droplets and the induction of C/EBP alpha, aP2, insulin-responsive aminopeptidase (IRAP), and glucose transporter 4 (GLUT4) expression. These PPAR gamma-expressing Swiss cells also exhibit a high level of insulin-responsive glucose uptake that is comparable to that expressed in 3T3-L1 adipocytes. In contrast, PPAR gamma-expressing NIH-3T3 fibroblasts, despite similar lipid accumulation, adipocyte morphology, and aP2 expression, do not synthesize C/EBP alpha and fail to acquire insulin sensitivity. In Swiss 3T3 cells ectopically expressing PPAR gamma, the development of insulin-responsive glucose uptake correlates with C/EBP alpha expression. Furthermore, ectopic expression of C/EBP alpha in NIH-3T3 cells induces PPAR gamma expression and adipogenesis, but also restores insulin-sensitive glucose transport. These results suggest that although PPAR gamma is sufficient to trigger the adipogenic program, C/EBP alpha is required for establishment of insulin-sensitive glucose transport in adipocytes.

3T3 Cells↗