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Biomedical subjects

Mitsuhiro Okamoto

Publications and source records attributed to Mitsuhiro Okamoto.

At least 19 recordsLinked to original sources
PubMed2007

Dephosphorylation of TORC initiates expression of the StAR gene.

Cyclic AMP responsive element (CRE) binding protein (CREB) is known to activate transcription when its Ser133 is phosphorylated. However, transducer of regulated CREB activity (TORC), a CREB specific co-activator, upregulates CREB activity in a phospho-Ser133-independent manner. Interestingly, TORC is also regulated by phosphorylation; the phospho-form is inactive, and the dephospho-form active. When PKA phosphorylates CREB, it inhibits TORC kinases simultaneously and accelerates dephosphorylation of TORC. We show in this report that staurosporine, a kinase inhibitor, induces the expression of the StAR gene in Y1 adrenocortical cells, possibly a result of an increase in the population of dephospho-TORC. The expression of the StAR gene is known to be regulated by SF-1 and CREB, and the co-activators CBP/p300 may mediate the actions of both factors. Our experiments using KG501, a disruptor of the interaction between phospho-CREB and CBP/p300, also support the importance of TORC in the regulation of StAR gene expression.

Animals↗
PubMed2006

Elevated cardiac tissue level of aldosterone and mineralocorticoid receptor in diastolic heart failure: Beneficial effects of mineralocorticoid receptor blocker.

Cardiac aldosterone levels have not been evaluated in diastolic heart failure (DHF), and its roles in this type of heart failure remain unclear. This study aimed to detect cardiac aldosterone by use of a liquid chromatographic-mass spectrometric method and to assess the effects of mineralocorticoid receptor blockade on hypertensive DHF. Dahl salt-sensitive rats fed 8% NaCl diet from 7 wk (hypertensive DHF model) were divided at 13 wk into three groups: those treated with subdepressor doses of eplerenone (12.5 or 40 mg x kg(-1) x day(-1)) and an untreated group. Dahl salt-sensitive rats fed 0.3% NaCl diet served as controls. Cardiac aldosterone was detected in the DHF rats but not in the control rats, with increased ventricular levels of mineralocorticoid receptor. Cardiac levels of 11-deoxycorticosterone, corticosterone, and 11-dehydrocorticosterone were not different between the control and DHF rats, but the tissue level of corticosterone that has an affinity to mineralocorticoid receptor was 1,000 times as high as that of aldosterone. Aldosterone synthase activity and CYP11B2 mRNA were undetectable in the ventricular tissue of the DHF rats. Administration of eplerenone attenuated ventricular hypertrophy, ventricular fibrosis, myocardial stiffening, and relaxation abnormality, leading to the prevention of overt DHF. In summary, the myocardial aldosterone level increased in the DHF rats. However, its value was extremely low compared with corticosterone, and no evidence for enhancement of intrinsic myocardial aldosterone production was found. The upregulation of mineralocorticoid receptor may play a central role in the pathogenesis of DHF, and blockade of mineralocorticoid receptor is likely an effective therapeutic regimen of DHF.

Aldosterone↗
PubMed2006

Plasma atrial natriuretic Peptide and brain natriuretic Peptide levels after radiofrequency catheter ablation of atrial fibrillation.

It has been reported that plasma atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) levels are elevated in patients with atrial fibrillation (AF). The aim of this study was to investigate the change in these patients after pulmonary vein isolation (PVI). In 66 patients with paroxysmal AF (PAF) and without any structural heart disease, plasma ANP and BNP levels were measured before and 3 months after successful PVI. At baseline, in 14 patients, ANP and BNP levels were elevated, and in 52 patients, only BNP levels were elevated. There were no significant relations between the attack frequency or the duration of PAF episodes and ANP or BNP levels. Neither ANP nor BNP level at baseline was a valid predictor of AF recurrence. Even in 31 patients (47%) with recurrent PAF, attacks of PAF were significantly reduced. In 66 patients with elevated ANP and/or BNP levels at baseline, levels were significantly reduced after PVI independent of PAF recurrence (ANP: 69.0+/-23.0 vs 25.0+/-7.7 pg/ml, p<0.0001; BNP: 58.4+/-50.7 vs 22.5+/-27.1 pg/ml, p<0.0001). In 42 patients without AF recurrences, ANP and BNP levels were reduced to within the normal range. In conclusion, in patients with PAF without any structural heart disease, ANP and/or BNP levels were elevated. In those patients, relief of the AF burden by successful PVI significantly reduced elevated plasma ANP and BNP levels.

Atrial Fibrillation↗
PubMed2006

Usefulness of esophageal leads for determining the strategy of pulmonary vein ablation to avoid complications associated with the esophagus.

To avoid fatal complications after extensive pulmonary vein (PV) ablation, it has been proved important to comprehend the anatomic relation between the PVs and the esophagus. In 42 consecutive patients with atrial fibrillation, PV ostial isolation was performed using a basket catheter. The shortest distance and anatomic relation between the esophageal lead and PV ostium, determined by successful PV ostial isolation, was analyzed in biplane fluoroscopic views. In 18 left superior PVs (LSPVs) (43%), 13 left inferior PVs (32%) (LIPVs), and all the right PVs (group A), the shortest distance was > 10 mm in > or = 1 of the biplane fluoroscopic views. In 4 LSPVs (10%) and 2 LIPVs (5%) (group B), the shortest distance was < or = 5 mm in the fluoroscopic views. In the remaining PVs (group C), the esophagus was situated directly behind 10 LSPVs (24%) and 12 LIPVs (29%) (group C1), posteromedial to 1 LSPV (2%) and 9 LIPVs (22%) (group C2), and medial to 9 LSPVs (21%) and 5 LIPVs (12%) (group C3). The risk of esophagus-associated complications with ablation around the left PV ostia was suggested to be high in group B, very low in group A, and relatively low in group C. In group C3, extensive PV ablation might increase the risk of that complication. In conclusion, esophageal leads are useful for determining strategies for PV ablation to avoid esophagus-associated complications, because they enable comprehension of the anatomic relation between the PVs and the esophagus.

Atrial Fibrillation↗
PubMed2006

Incidence, location, and cause of recovery of electrical connections between the pulmonary veins and the left atrium after pulmonary vein isolation.

AIMS: The aim of this study was to reveal the incidence, location, and cause of recovery of the electrical connections (ECs) between the left atrium and the pulmonary veins (PVs) after the segmental ostial PV isolation (PVI). METHODS AND RESULTS: Pulmonary vein mapping and successful PVI were performed using a computerized three-dimensional mapping system (QMS2trade mark) with a basket catheter in 167 PVs in 53 consecutive patients with atrial fibrillation (AF). In 14 patients with recurrent AF after PVI, the same PV mapping and isolation as in the first procedure were performed, and the PV potential maps constructed by QMS2 in two different procedures were compared. Forty-nine recovered ECs were observed in 27 PVs, and all were eliminated by a few local radiofrequency (RF) applications. Thirty-four (69%) of those ECs recovered at the edge of original ECs, and another 15 (31%) recovered at the mid-portion of the continuous broad original ECs. CONCLUSION: Electrical connection recovery occurred most commonly at the edges of original ECs and occasionally at the mid-portion of continuous broad original ECs after PVI probably due to tissue oedema neighbouring the segmental RF lesions. Further RF lesions at the edge of original ECs and linear ablation to the continuous broad ECs may help reduce AF recurrence.

Atrial Fibrillation↗
PubMed2006

Electrophysiological pulmonary vein antrum isolation with a multielectrode basket catheter is feasible and effective for curing paroxysmal atrial fibrillation: efficacy of minimally extensive pulmonary vein isolation.

BACKGROUND: How extensive should an appropriate pulmonary vein (PV) ablation be is a matter of controversy. OBJECTIVE: The study's aim was to investigate the efficacy of minimally extensive PV ablation for isolating the PV antrum (PVA) with the guidance of electrophysiological parameters. METHODS: Fifty-five consecutive symptomatic paroxysmal atrial fibrillation (PAF) patients underwent PV mapping with a multielectrode basket catheter (MBC). A 31-mm MBC was deployed in 3-4 PVs as proximally as possible without dislodgement, and the longitudinal PV mapping enabled us to recognize single sharp potentials formed by the total fusion of the PV and left atrial potentials around the PV ostium or the transverse activation patterns that were observed. Those potentials were defined as PVA potentials. Radiofrequency ablation was performed circumferentially targeting PVA potentials with the end point being their elimination. RESULTS: After circumferential PVA ablation, electrical disconnection was achieved in 77% and residual PVA conduction gaps were observed in 23% of all targeted PVs. Those residual conduction gaps were mainly located at the border between ipsilateral PVs (42%) and between the left PVs and left atrial appendage (33%) and were eliminated by a mean of 3 +/- 2 minutes of local radiofrequency deliveries. During the follow-up period (11 +/- 5 months), 46 (84%) patients were free of symptomatic PAF without any anti-arrhythmic drugs. No PV stenosis or spontaneous left atrial flutter occurred. CONCLUSIONS: Electrophysiological PVA ablation with an MBC is feasible and effective for curing PAF because this minimally extensive PVA isolation technique targets the optimal sites, achieving both high efficacy and safety.

Aged↗
PubMed2006

Silencing the constitutive active transcription factor CREB by the LKB1-SIK signaling cascade.

Cyclic AMP responsive element (CRE)-binding protein (CREB) is known to activate transcription when its Ser133 is phosphorylated. Two independent investigations have suggested the presence of Ser133-independent activation. One study identified a kinase, salt-inducible kinase (SIK), which repressed CREB; the other isolated a novel CREB-specific coactivator, transducer of regulated CREB activity (TORC), which upregulated CREB activity. These two opposing signals are connected by the fact that SIK phosphorylates TORC and induces its nuclear export. Because LKB1 has been reported to be an upstream kinase of SIK, we used LKB1-defective HeLa cells to further elucidate TORC-dependent CREB activation. In the absence of LKB1, SIK was unable to phosphorylate TORC, which led to constitutive activation of CRE activity. Overexpression of LKB1 in HeLa cells improved the CRE-dependent transcription in a regulated manner. The inactivation of kinase cascades by 10 nm staurosporine in LKB1-positive HEK293 cells also induced unregulated, constitutively activated, CRE activity. Treatment with staurosporine completely inhibited SIK kinase activity without any significant effect on the phosphorylation level at the LKB1-phosphorylatable site in SIK or the activity of AMPK, another target of LKB1. Constitutive activation of CREB in LKB1-defective cells or in staurosporine-treated cells was not accompanied by CREB phosphorylation at Ser133. The results suggest that LKB1 and its downstream SIK play an important role in silencing CREB activity via the phosphorylation of TORC, and such silencing may be indispensable for the regulated activation of CREB.

AMP-Activated Protein Kinases↗
PubMed2006

The distal protection during primary percutaneous coronary intervention alleviates the adverse effects of large thrombus burden on myocardial reperfusion.

BACKGROUND: The presence of intracoronary large thrombus burden (LTB) in the infarct-related artery increases the risk of distal embolization and no-reflow during percutaneous coronary intervention (PCI). Evaluation of whether the distal protection (DP) during primary PCI reduces adverse effects of LTB on myocardial reperfusion and infarct size was investigated. METHODS AND RESULTS: A consecutive series of 88 patients with acute myocardial infarction undergoing primary PCI using DP were compared with 81 consecutive patients treated by primary PCI alone. The DP use showed similar post-procedural myocardial blush grade (MBG)-3 and infarct size, but improved corrected thrombolysis in myocardial infarction frame count (cTFC) (29+/-11 vs 35+/-20, p=0.011) and the incidence of ST-segment resolution (80.7% vs 66.7%, p=0.038). In patients with LTB present, however, the DP use reduced occurrences of no-reflow (0% vs 11.8%, p=0.036) and distal embolization (4.8% vs 17.6%, p=0.129), resulting in higher occurrences of MBG-3 (61.9% vs 35.3%, p=0.021) and ST-segment resolution (78.6% vs 50.0%, p=0.009), lower cTFC values (30+/-8 vs 40+/-22, p=0.012) and smaller infarct size (12.2+/-11.2 vs 18.7+/-11.1, p=0.015). CONCLUSIONS: With an improved myocardial reperfusion and smaller infarct size in patients with LTB, the DP during primary PCI might be a better strategy in this particular setting compared with conventional strategy.

Acute Disease↗
PubMed2006

Can segmental pulmonary vein ablation reduce the recurrence of atrial fibrillation when using a higher RF power, larger tip electrode catheter, and additional RF deliveries?: the limitations of point-by-point RF ablation.

The aim of this study was to investigate whether segmental ostial catheter ablation (SOCA) designed to prevent the electrical connections (ECs) between the left atrium and pulmonary veins (PVs) might help increase the efficacy of SOCA in paroxysmal atrial fibrillation (PAF). PV mapping and successful SOCA were performed with a basket catheter in 108 consecutive patients with PAF. Radiofrequency energy was delivered using a maximum output of 30 W with a 4 mm tip catheter (group I; 47) or 40 W with an 8 mm tip catheter (group II; 61). Only in the group II patients were additional radiofrequency deliveries to the specific sites where the ECs tended to recover performed after successful SOCA. After the first procedure, PAF recurred in 47% of the group I patients and 32% of the group II patients. In all 27 patients who underwent repeat procedures, EC recoveries were observed more frequently in group I than in group II (69% versus 49%; P < 0.05). After multiple procedures, there was more freedom from PAF in group II (84%) than in group I (66%) (P < 0.05). SOCA with a higher RF power, larger tip catheter, and additional RF deliveries could achieve a more effective SOCA.

Aged↗
PubMed2005

Molecular identity and gene expression of aldosterone synthase cytochrome P450.

11Beta-hydroxylase (CYP11B1) of bovine adrenal cortex produced corticosterone as well as aldosterone from 11-deoxycorticosterone in the presence of the mitochondrial P450 electron transport system. CYP11B1s of pig, sheep, and bullfrog, when expressed in COS-7 cells, also performed corticosterone and aldosterone production. Since these CYP11B1s are present in the zonae fasciculata and reticularis as well as in the zona glomerulosa, the zonal differentiation of steroid production may occur by the action of still-unidentified factor(s) on the enzyme-catalyzed successive oxygenations at C11- and C18-positions of steroid. In contrast, two cDNAs, one encoding 11beta-hydroxylase and the other encoding aldosterone synthase (CYP11B2), were isolated from rat, mouse, hamster, guinea pig, and human adrenals. The expression of CYP11B1 gene was regulated by cyclic AMP (cAMP)-dependent signaling, whereas that of CYP11B2 gene by calcium ion-signaling as well as cAMP-signaling. Salt-inducible protein kinase, a cAMP-induced novel protein kinase, was one of the regulators of CYP11B2 gene expression.

Animals↗
PubMed2005

Computerized three-dimensional potential mapping with a multielectrode basket catheter can be useful for pulmonary vein electrical disconnection.

INTRODUCTION: Pulmonary vein (PV) isolation (PVI) has been recently proposed as an effective technique to cure atrial fibrillation (AF). AIMS OF THE STUDY: The aim of this study was to investigate the efficacy of a novel technique utilizing a computerized three-dimensional mapping system (QMS2) with a multielectrode basket catheter (MBC) for PVI and to reveal the relation between the style of breakthrough and the network of the PV musculature. METHODS: Sixty-five consecutive patients with frequent AF attacks underwent PV mapping with a 31-mm MBC, and a three-dimensional color animation of the potential map was constructed by the QMS2. The animation color schema was arranged to minimize the low-amplitude left atrial (LA) potentials and emphasize the high-amplitude PV potentials (PVPs). The longitudinal PVP map enabled us to recognize the true breakthroughs and reveal the network of the PV musculature. RESULTS: A total of 205 PVs (65 left superior PVs, 65 right superior PVs, 57 left inferior PVs and 18 right inferior PVs) were mapped and successful PVI was achieved in all PVs, except one that had no PVPs, with a mean radiofrequency duration of 7 +/- 5 minutes per PV. In about 90% of the PVs, a final radiofrequency application eliminated all the distal PVPs simultaneously because the PVI was performed at the appropriate LA-PV junction. A single segmental breakthrough was detected in 17 PVs, single broad breakthrough in 83 PVs, multiple separate breakthroughs with a distal connection between the PV musculatures extending from each separate breakthrough in 88 PVs and multiple separate breakthroughs without that connection in 16 PVs. During the follow-up period, fifty-one (78%) patients were free of symptomatic AF without any antiarrhythmic drugs after multiple procedures (thirty-three (51%) of those patients after the first procedure) and no PV stenosis was found. CONCLUSIONS: Computerized three-dimensional potential mapping can be useful for PVI because it can not only identify the true breakthrough, but can also confirm the elimination of the breakthroughs by the change in the activation sequence through the network of the PV musculature.

Analysis of Variance↗
PubMed2005

Molecular identification of adrenal inner zone antigen as a heme-binding protein.

The adrenal inner zone antigen (IZA), which reacts specifically with a monoclonal antibody raised against the fasciculata and reticularis zones of the rat adrenal, was previously found to be identical with a protein variously named 25-Dx and membrane-associated progesterone receptor. IZA was purified as a glutathione S-transferase-fused or His(6)-fused protein, and its molecular properties were studied. The UV-visible absorption and EPR spectra of the purified protein showed that IZA bound a heme chromophore in high-spin type. Analysis of the heme indicated that it is of the b type. Site-directed mutagenesis studies were performed to identify the amino-acid residues that bind the heme to the protein. The results suggest that two Tyr residues, Tyr107 and Tyr113, and a peptide stretch, D99-K102, were important for anchoring the heme into a hydrophobic pocket. The effect of IZA on the steroid 21-hydroxylation reaction was investigated in COS-7 cell expression systems. The results suggest that the coexistence of IZA with CYP21 enhances 21-hydroxylase activity.

Adrenal Cortex↗
PubMed2004

The CREB coactivator TORC2 functions as a calcium- and cAMP-sensitive coincidence detector.

Elevations in circulating glucose and gut hormones during feeding promote pancreatic islet cell viability in part via the calcium- and cAMP-dependent activation of the transcription factor CREB. Here, we describe a signaling module that mediates the synergistic effects of these pathways on cellular gene expression by stimulating the dephosphorylation and nuclear entry of TORC2, a CREB coactivator. This module consists of the calcium-regulated phosphatase calcineurin and the Ser/Thr kinase SIK2, both of which associate with TORC2. Under resting conditions, TORC2 is sequestered in the cytoplasm via a phosphorylation-dependent interaction with 14-3-3 proteins. Triggering of the calcium and cAMP second messenger pathways by glucose and gut hormones disrupts TORC2:14-3-3 complexes via complementary effects on TORC2 dephosphorylation; calcium influx increases calcineurin activity, whereas cAMP inhibits SIK2 kinase activity. Our results illustrate how a phosphatase/kinase module connects two signaling pathways in response to nutrient and hormonal cues.

14-3-3 Proteins↗
PubMed2004

Salt-inducible kinase (SIK) isoforms: their involvement in steroidogenesis and adipogenesis.

The cloning of salt-inducible kinase-1 (SIK1) that was specifically expressed in the adrenal glands of high-salt diet-fed rats led to subsequent cloning of adipose-specific SIK2 and rather ubiquitous SIK3. The three enzymes constitute a novel serine/threonine kinase subfamily, a member of AMP-activated protein kinase (PKA) family. Physiological roles of SIK1 and SIK2 have been investigated. The SIK1 transcript was expressed very early in the ACTH-stimulated Y1 cells, even before the expression of transcripts for CYP11A and StAR protein. Forced expression of SIK1 inhibited the ACTH-dependent expression of CYP11A- and StAR protein-genes. Cotransfection assays employing CRE-reporter gene showed that SIK1 could repress the PKA-dependent activation of CRE by acting on the bZIP domain of the CRE-binding protein (CREB), though the target site of SIK1-mediated phosphorylation has yet to be determined. ACTH/PKA-dependent nucleocytoplasmic shuttling of SIK1 took place in Y1 cells, implying that the intracellular movement of SIK1 might be a physiologically important determining factor for regulation of steroidogenic gene expression in the early phase of ACTH-stimulation. The SIK2 gene was expressed in 3T3-L1 cells at a very early stage of adipogenesis. SIK2 could phosphorylate Ser-794 of human insulin-receptor-substrate-1 (IRS-1) in vitro as well as in vivo. In addition, the SIK2 activity in db/db mice adipose tissues was significantly higher than that in wild-type adipose. These results strongly suggest that SIK2 may play important role(s) in modulating the insulin-signaling cascade of adipocytes, and thus, may be involved in the development of insulin resistance. Taken together, these results suggest that the SIK isoforms regulate hormonal signal transduction in both adrenal and adipose tissues.

Adipose Tissue↗
PubMed2004

Simple and accurate catheter mapping technique to predict atrial fibrillation foci in the pulmonary veins or posterior right atrium.

OBJECTIVES: The purpose of this study was to investigate the efficacy of a novel catheter mapping technique for predicting atrial fibrillation (AF) foci. BACKGROUND: Most AF originates from pulmonary veins (PVs), but some originate from the right atrium. METHODS: We developed an algorithm by correlating the cardiac recordings obtained from multielectrode catheters placed in the posterior right atrium (RA) and esophagus during pacing from the PVs and superior vena cava (SVC) or crista terminalis (CT) in 10 AF patients. We tested the algorithm's accuracy prospectively in 46 AF patients. RESULTS: During pacing from the left PVs, the esophageal potentials preceded all other potentials. During pacing from both the right PVs and SVC-CT, the first component (FP) of the double potential (DP) recorded in the posterior RA preceded all other potentials. The amplitude of the FP was higher than that of the second DP component during pacing from the SVC-CT, whereas the reverse occurred from the right PVs. The activation sequence of the FPs and esophageal potentials was from superior to inferior during pacing from the superior PVs, whereas the reverse occurred from the inferior PVs. The accuracy of predicting 34 foci in the right PVs, 28 foci in left PVs, and 6 foci in SVC-CT was 100% for all, respectively. The accuracy of discriminating foci in the superior PVs from those in the inferior PVs was 97% in the right PVs and 96% in the left PVs. CONCLUSIONS: The technique using mapping catheters placed in the posterior RA and esophagus is feasible and effective for mapping and ablating AF.

Algorithms↗
PubMed2004

Electrophysiologic characteristics of atrial tachycardia originating from the right pulmonary veins or posterior right atrium: double potentials obtained from the posterior wall of the right atrium can be useful to predict foci of atrial tachycardia in right pulmonary veins or posterior right atrium.

INTRODUCTION: The right pulmonary veins (RPVs) and posterior wall of the right atrium (PRA) are anatomically located adjacent to each other. The aim of this study was to demonstrate the electrophysiologic characteristics of atrial tachycardia (AT) originating from the PRA or RPVs. METHODS AND RESULTS: A total of 26 consecutive patients with AT originating from the RPVs or PRA underwent detailed atrial endocardial mapping and successful radiofrequency catheter ablation. Eight foci were found in the PRA and 18 foci in the RPVs. Analysis of P wave configuration showed that lead V1 was the most helpful in distinguishing the AT foci between these two sites. In all cases, double potential (DP) configurations were recorded from several electrodes of a multielectrode catheter placed in the PRA, and the first DP component (FP) was the earliest potential recorded from the right atrium during the tachycardia. The amplitude of the FP was higher than that of the second DP component (SP) for AT foci originating in the PRA, whereas the reverse occurred for those in the RPV. The activation sequence of the FP was from superior to inferior for the AT foci in the superior RPV, whereas the reverse occurred for the AT foci in the inferior RPV. CONCLUSION: P wave configuration in lead V1 is helpful in distinguishing AT foci between those originating in the PRA and RPVs. The DPs obtained from the PRA can be useful in predicting whether AT foci originate from the PRA or RPVs.

Adolescent↗
PubMed2004

Salt-inducible kinase-1 represses cAMP response element-binding protein activity both in the nucleus and in the cytoplasm.

Salt-inducible kinase-1 (SIK1) is phosphorylated at Ser577 by protein kinase A in adrenocorticotropic hormone-stimulated Y1 cells, and the phospho-SIK1 translocates from the nucleus to the cytoplasm. The phospho-SIK1 is dephosphorylated in the cytoplasm and re-enters the nucleus several hours later. By using green-fluorescent protein-tagged SIK1 fragments, we found that a peptide region (586-612) was responsible for the nuclear localization of SIK1. The region was named the 'RK-rich region' because of its Arg- and Lys-rich nature. SIK1s mutated in the RK-rich region were localized mainly in the cytoplasm. Because SIK1 represses cAMP-response element (CRE)-mediated transcription of steroidogenic genes, the mutants were examined for their effect on transcription. To our surprise, the cytoplasmic mutants strongly repressed the CRE-binding protein (CREB) activity, the extent of repression being similar to that of SIK1(S577A), a mutant localized exclusively in the nucleus. Several chimeras were constructed from SIK1 and from its isoform SIK2, which was localized mainly in the cytoplasm, and they were examined for intracellular localization as well as CREB-repression activity. A SIK1-derived chimera, where the RK-rich region had been replaced with the corresponding region of SIK2, was found in the cytoplasm, its CREB-modulating activity being similar to that of wild-type SIK1. On the other hand, a SIK2-derived chimera with the RK-rich region of SIK1 was localized in both the nucleus and the cytoplasm, and had a CREB-repressing activity similar to that of the wild-type SIK2. Green fluorescent protein-fused transducer of regulated CREB activity 2 (TORC2), a CREB-specific co-activator, was localized in the cytoplasm and nucleus of Y1 cells, and, after treatment with adrenocorticotropic hormone, cytoplasmic TORC2 entered the nucleus, activating CREB. The SIK1 mutants, having a strong CRE-repressing activity, completely inhibited the adrenocorticotropic hormone-induced nuclear entry of green fluorescent protein-fused TORC2. This suggests that SIK1 may regulate the intracellular movement of TORC2, and as a result modulates the CREB-dependent transcription activity. Together, these results indicate that the RK-rich region of SIK1 is important for determining the nuclear localization and attenuating CREB-repressing activity, but the degree of the nuclear localization of SIK1 itself does not necessarily reflect the degree of SIK1-mediated CREB repression.

3T3-L1 Cells↗