GH induced lipolysis stimulation in 3T3-L1 adipocytes stably expressing hGHR: analysis on signaling pathway and activity of 20K hGH

Asada, Nobuhiko; Takahashi, Yuka; Wada, Mayuko; Naito, N; Uchida, Hiroshi; Ikeda, Miwa; Honjo, Masaru

doi:10.1016/s0303-7207(00)00202-1

Cited by 34 publications

(29 citation statements)

References 36 publications

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“…Although PKC-independent activation of STATs has been reported, 23,24 other studies have demonstrated PKC-dependent activation of STAT proteins, 25,26 indicating that STAT phosphorylation may be either upstream or downstream of PKC activation. To determine whether Bryostatin 1-induced STAT1 activation is dependent on PKC signaling, CLL cells were analyzed for Bryostatin 1-induced STAT1 activation following treatment with selective pharmacologic inhibitors of PKC.…”

Section: Activation Of Stat1 Is Dependent On Pkc Activationmentioning

confidence: 98%

STAT1 mediates differentiation of chronic lymphocytic leukemia cells in response to Bryostatin 1

Battle

Frank

2003

Blood

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Section: Activation Of Stat1 Is Dependent On Pkc Activationmentioning

confidence: 98%

STAT1 mediates differentiation of chronic lymphocytic leukemia cells in response to Bryostatin 1

Battle

Frank

2003

Blood

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“…Aberrant splicing to an in-frame cryptic splice site in exon 3, which occurs 5-10% of the time, produces a transcript lacking the first 45 nucleotides of exon 3 and encodes a 20-kDa protein. The 20-kDa isoform apparently retains full biological activity (11)(12)(13). Complete skipping of exon 3, which occurs 0.1-5% of the time, results in production of a 17.5-kDa protein that acts as a dominant negative by blocking secretion of the full-length protein (14).…”

mentioning

confidence: 99%

Growth Hormone Deficiency and Splicing Fidelity

Solis

Peng

Crawford

et al. 2008

Journal of Biological Chemistry

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The majority of mutations that cause isolated growth hormone deficiency type II are the result of aberrant splicing of transcripts encoding human growth hormone. Such mutations increase skipping of exon 3 and encode a 17.5-kDa protein that acts as a dominant negative to block secretion of full-length protein produced from unaffected alleles. Previously, we identified a splicing regulatory element in exon 3 (exonic splicing enhancer 2 (ESE2)), but we had not determined the molecular mechanism by which this element prevents exon skipping. Here, we show that two members of the serine/arginine-rich (SR) protein superfamily (ASF/SF2 and SC35) act antagonistically to regulate exon 3 splicing. ASF/SF2 activates exon 3 inclusion, but SC35, acting through a region just downstream of ESE2, can block such activation. These findings explain the disease-causing mechanism of a patient mutation in ESE2 that creates a functional SC35-binding site that then acts synergistically with the downstream SC35 site to produce pathological levels of exon 3 skipping. Although the precedent for SR proteins acting as repressors is established, this is the first example of a patient mutation that creates a site through which an SR protein represses splicing.Pre-mRNA splicing is the process of intron removal and exon joining to form mature, protein-coding transcripts. This requires both cis-acting RNA elements and trans-acting factors. The main cis-acting elements are the 5Ј splice site, 3Ј splice site, polypyrimidine tract, and branch point sequence, each of which are defined by short, degenerate consensus sequences in higher eukaryotes (1, 2). Recognition of these elements by trans-acting factors, including five small nuclear ribonucleoproteins, results in assembly of the catalytically active spliceosome in which the two steps of splicing occur. Because of the lack of sequence conservation, the pairing of U1 snRNA with the 5Ј splice site and U2 with the branch point results in a range of splice site strengths. This allows for regulation of splice site selection in cases of alternative splicing but at the same time creates a potential fidelity problem in cases where weak splice sites need to be properly recognized. These weak splice sites often require additional sequences known as enhancer elements that are recognized by regulatory splicing factors, such as SR 3 proteins, which guide the spliceosome to the correct splice sites. Splicing enhancers are typically purinerich and can be found in both exons and introns and are accordingly named exonic (ESEs) and intronic splicing enhancers. SR proteins are a large family of conserved proteins that have N-terminal RNA recognition motifs and C-terminal domains rich in serine-arginine dipeptides (RS domains). The RNA recognition motifs tether the proteins to enhancer elements, and the RS domains assist spliceosome assembly through proteinprotein interactions (3).SR proteins define exon boundaries by two mechanisms that are not mutually exclusive. First, SR proteins enhance splicing in an RS domain-i...

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“…Recently we have revealed that hGH markedly stimulates lipolysis in 3T3-L1-hGHR which can efficiently express human growth hormone receptor (hGHR) after differentiation to adipocyte [5]. Using this cell line, we have also confirmed that the lipolytic action of 22K hGH is reduced in the presence of hGHBP, suggesting that increased hGHBP inhibited 22K hGH-induced lipolysis in humans.…”

Section: Introductionmentioning

confidence: 57%

“…3T3-L1-hGHR cell construction, culture and conversion to adipocytes were described previously [5]. After 18–24 h culture in Dulbecco’s modified Eagle’s medium (DMEM) with 2% bovine serum albumin (BSA), the 3T3-L1-hGHR adipocytes were incubated in 750 µl of the medium for lipolysis (Krebs-Ringer bicarbonate buffer (pH 7.4) with 4% BSA (fatty acid content 0.005%), 2 m M glucose, and 40 n M dexamethasone) containing GHs or without GHs as control for 48 h [5].…”

Section: Methodsmentioning

confidence: 99%

“…After 18–24 h culture in Dulbecco’s modified Eagle’s medium (DMEM) with 2% bovine serum albumin (BSA), the 3T3-L1-hGHR adipocytes were incubated in 750 µl of the medium for lipolysis (Krebs-Ringer bicarbonate buffer (pH 7.4) with 4% BSA (fatty acid content 0.005%), 2 m M glucose, and 40 n M dexamethasone) containing GHs or without GHs as control for 48 h [5]. When effects of hGHBP were studied, hGHs were preincubated with 5 n M hGHBP for 1 h at 25°C.…”

Section: Methodsmentioning

confidence: 99%

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Effects of 22K or 20K Human Growth Hormone on Lipolysis, Leptin Production in Adipocytes in the Presence and Absence of Human Growth Hormone Binding Protein

Asada

Takahashi²,

Honjo³

2000

Horm Res Paediatr

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Objective and Method: We studied the effects of human growth hormone (hGH) on leptin production and lipolysis stimulation in the presence or absence of human growth hormone binding protein (hGHBP) using 3T3- L1-hGHR adipocytes which efficiently express human growth hormone receptor. Results and Conclusion: It was clarified that (1) hGH decreases leptin secretion after hGH-induced lipolysis stimulation, and (2) the reduction of leptin production and lipolysis stimulation by 22K hGH was attenuated with hGHBP, whereas that by 20K hGH, which is a naturally occurring isoform of 22K hGH, was not affected with hGHBP.

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GH induced lipolysis stimulation in 3T3-L1 adipocytes stably expressing hGHR: analysis on signaling pathway and activity of 20K hGH

Cited by 34 publications

References 36 publications

STAT1 mediates differentiation of chronic lymphocytic leukemia cells in response to Bryostatin 1

STAT1 mediates differentiation of chronic lymphocytic leukemia cells in response to Bryostatin 1

Growth Hormone Deficiency and Splicing Fidelity

Effects of 22K or 20K Human Growth Hormone on Lipolysis, Leptin Production in Adipocytes in the Presence and Absence of Human Growth Hormone Binding Protein

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