Details of a Researcher - Yoshimura, Akihiko

Papers - Yoshimura, Akihiko

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The adapter protein APS associates with the multifunctional docking sites Tyr-568 and Tyr-936 in c-Kit

Wollberg Patrik, Lennartsson Johan, Gottfridsson Eva, Yoshimura Akihiko, Rönnstrand Lars

Biochemical Journal 370 ( 3 ) 1033 - 1038 2003.03

ISSN 0264-6021

The adapter protein APS has previously been shown to be involved in recruiting the ubiquitin E3 ligase c-Cbl to the insulin receptor, the platelet-derived growth factor β-receptor and the erythropoietin receptor, leading to increased degradation of the receptors and inhibition of mitogenesis. Here we demonstrate, by use of immobilized synthetic phosphopeptides corresponding to various autophosphorylated tyrosine residues in the receptor for stem-cell factor (c-Kit), that APS preferentially associates with phosphorylated Tyr-568 and Tyr-936. Tyr-568 has previously been identified as the binding site of the Src family of tyrosine kinases, the Csk-homologous kinase CHK, and the protein tyrosine phosphatase SHP-2. We have recently demonstrated that Tyr-936 is an autophosphorylation site involved in binding the adapter proteins Grb2 and Grb7. We could further demonstrate that the critical determinant for binding of APS is the presence of either a leucine or an isoleucine residue in the position +3 to the phosphorylated tyrosine. This allowed us to design mutants that selectively failed to associate with APS, while still associating with Src family members, SHP-2 and Grb2, respectively.

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Interaction of the growth hormone receptor with cytokine-induced Src homology domain 2 protein in rat adipocytes

Du Ling, Frick G. Peter, Tai Lih Ruey, Yoshimura Akihiko, Goodman H. Maurice

Endocrinology 144 ( 3 ) 868 - 876 2003.03

ISSN 0013-7227

GH stimulates the phosphorylation of tyrosine residues in the GH receptor (GHR), Janus kinase 2 (JAK2), and other signaling proteins in a transient manner that subsides within 1 h. To assess the possible roles of cytokine-induced Src homology domain 2 (SH2) (CIS/SOCS) proteins in these transient responses, we studied the expression and disposition of CIS/SOCS proteins in rat adipocytes, a physiological target of GH action. A tyrosine-phosphorylated protein that appears to be the GHR was coprecipitated from extracts of GH-treated adipocytes with α-CIS. In contrast, no tyrosine-phosphorylated adipocyte proteins were recovered after immunoprecipitation with α-SOCS3, although coprecipitation of GHR with SOCS3 was readily detected in extracts of 3T3-F442A fibroblasts. Interaction of GHR with CIS peaked between 2 and 10 min after adipocytes were treated with GH, when tyrosine phosphorylation of the GHR was maximal. By 60 min after GH, tyrosine phosphorylation of the GHR declined to very low levels, and its interaction with CIS was reduced correspondingly. Proteasome inhibitors prevented the decline in tyrosine-phosphorylated GHR and prolonged interaction of GHR and CIS for at least 1 h. These findings demonstrate the interaction of CIS with the GHR in vivo and suggest that CIS may enhance degradation of the receptor by a proteasomal pathway.

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Differential control of the NIMA-related kinases, Nek6 and Nek7, by serum stimulation

Minoguchi Shigeru, Minoguchi Mayu, Yoshimura Akihiko

Biochemical and Biophysical Research Communications 301 ( 4 ) 899 - 906 2003.02

ISSN 0006-291X

Neks (NIMA-related kinases) are mammalian serine/threonine (Ser/Thr) protein kinases structurally related to Aspergillus NIMA (Never in Mitosis, gene A), which plays essential roles in mitotic signaling. Among these kinases, Nek6 and Nek7 are structurally related and constitute a subfamily in the NIMA/Nek family, although their functions still remain almost elusive. In this report, we studied the enzymatic regulation of Nek6 and Nek7 to gain an insight into their cellular functions. Recombinant Nek7 produced in bacteria was active comparably to Nek6; however, the Nek7 activity in mammalian cells was found to be significantly lower than Nek6. Since Nek6 previously has been reported to in vitro phosphorylate p70 ribosomal S6 kinase at Thr412, we examined if Nek6 and Nek7 activities were controlled by the amino acid supplement, which is known to affect the phosphorylation at Thr412, and did not observe any significant effect. However, we unexpectedly found that Nek7 kinase activity was rapidly and efficiently increased by serum deprivation, while Nek6 activity was decreased. This is well consistent with the lower activity of Nek7 in cells under normal growth conditions. In addition, it was suggested that Nek7 activity would be regulated in a cell cycle-dependent manner, although Nek6 was not. These clear differences in enzymatic control between the highly similar kinases, Nek6 and Nek7, suggest their distinct signaling functions in mammalian cells. © 2003 Elsevier Science (USA). All rights reserved.

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A role of suppressor of cytokine signaling 3 (SOCS3/CIS3/SSI3) in CD28-mediated interleukin 2 production

Matsumoto Akira, Seki Yoh ichi, Watanabe Ryosuke, Hayashi Katsuhiko, Johnston James A., Harada Yohsuke, Abe Ryo, Yoshimura Akihiko, Kubo Masato

Journal of Experimental Medicine 197 ( 4 ) 425 - 436 2003.02

ISSN 0022-1007

Suppressor of cytokine signaling (SOCS)3 has been characterized as a negative feedback regulator in cytokine-mediated Janus kinase signal transducer and activator of transcription signaling. However, this study shows that T cells from transgenic mice expressing SOCS3 exhibit a significant reduction in interleukin (IL)-2 production induced by T cell receptor cross-linking when T cells are costimulated with CD28. Decreased protein expression in SOCS3^+/- mice enhanced CD28-mediated IL-2 production, clearly indicating the correlation between expression level of SOCS3 and IL-2 production ability. The SOCS3 protein interacted with phosphorylated CD28 through its SH2 domain but not the kinase inhibitory region. In addition, a point mutation in the SOCS3 SH2 domain attenuated the inhibition of CD28 function in IL-2 promoter activation. Committed T helper (Th)2 cells exclusively expressed SOCS3 and production of Th2 cytokines, such as IL-4 and IL-5, was much less dependent on CD28 costimulation compared with interferon γ and IL-2 production in Th1 cells. Consistent with this notion, the expression level of SOCS3 in early T cell activation influenced the ability of IL-2 production induced by CD28 costimulation. Therefore, the SOCS3 may play an alternative role in prohibiting excessive progression of CD28-mediated IL-2 production.

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The suppressor of cytokine signaling-1 (SOCS1) is a novel therapeutic target for enterovirus-induced cardiac injury

Yasukawa Hideo, Yajima Toshitaka, Duplain Hervé, Iwatate Mitsuo, Kido Masakuni, Hoshijima Masahiko, Weitzman Matthew D., Nakamura Tomoyuki, Woodard Sarah, Xiong Dingding, Yoshimura Akihiko, Chien Kenneth R., Knowlton Kirk U.

Journal of Clinical Investigation 111 ( 4 ) 469 - 478 2003.02

ISSN 0021-9738

Enteroviral infections of the heart are among the most commonly identified causes of acute myocarditis in children and adults and have been implicated in dilated cardiomyopathy. Although there is considerable information regarding the cellular immune response in myocarditis, little is known about innate signaling mechanisms within the infected cardiac myocyte that contribute to the host defense against viral infection. Here we show the essential role of Janus kinase (JAK) signaling in cardiac myocyte antiviral defense and a negative role of an intrinsic JAK inhibitor, the suppressor of cytokine signaling (SOCS), in the early disease process. Cardiac myocyte-specific transgenic expression of SOCS1 inhibited enterovirus-induced signaling of JAK and the signal transducers and activators of transcription (STAT), with accompanying increases in viral replication, cardiomyopathy, and mortality in coxsackievirus-infected mice. Furthermore, the inhibition of SOCS in the cardiac myocyte through adeno-associated virus-mediated (AAV-mediated) expression of a dominant-negative SOCS1 increased the myocyte resistance to the acute cardiac injury caused by enteroviral infection. These results indicate that strategies directed at inhibition of SOCS in the heart and perhaps other organs can augment the host-cell antiviral system, thus preventing viral-mediated endorgan damage during the early stages of infection.

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The N-terminal truncated isoform of SOCS3 translated from an alternative initiation AUG codon under stress conditions is stable due to the lack of a major ubiquitination site, Lys-6

Sasaki Atsuo, Inagaki-Ohara Kyoko, Yoshida Takafumi, Yamanaka Atsushi, Sasaki Mika, Yasukawa Hideo, Koromilas Antonis E., Yoshimura Akihiko

Journal of Biological Chemistry 278 ( 4 ) 2432 - 2436 2003.01

ISSN 0021-9258

The suppressor of cytokine signaling-3 (SOCS3/CIS-33/SSI-3) is an important negative regulator of cytokine signaling. Here, we show that an N-terminal truncated isoform (AN-SOCS3) translated from the internal AUG codon 12 was profoundly induced by endoplasmic reticulum (ER) stress- or active double-stranded RNA-activated protein kinase PKR, as a result of induction of eukaryotic initiation factor 2α phosphorylation. ΔN-SOCS3 exhibited a stronger cytokine-inhibitory activity and a higher stability than WT-SOCS3 in Ba/F3 hematopoietic cells. A potential ubiquitination residue, Lys-6, at the N terminus is evolutionary conserved among SOCS3 species. The K6Q-SOCS3 mutant showed a much longer half-life than WT-SOCS3 in Ba/F3 cells. Furthermore, inhibition of the 26 S proteasome pathway increased both ubiquitination and protein levels of WT-SOCS3 but had no effect on K6Q-SOCS3. SOCS3 mutant lacking the carboxyl-terminal SOCS-box exhibited the same stability as K6Q-SOCS3. These observations suggest that the short form of SOCS3 is a naturally occurring stabilized inhibitory protein, whereas WT-SOCS3 is a short-lived protein modulated by Lys-6 ubiquitination and proteasome-dependent degradation. Our findings provide strong evidence for the first time that translational control plays an important role in stabilization and function of SOCS3.

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Control mechanism of JAK/STAT signal transduction pathway

Yamada Satoshi, Shiono Satoru, Joo Akiko, Yoshimura Akihiko

FEBS Letters 534 ( 1-3 ) 190 - 196 2003.01

ISSN 0014-5793

Suppressor of cytokine signaling-1 (SOCS1) was identified as the negative regulator of Janus kinase (JAK) and signal transducer and activator of transcription (STAT) signal transduction pathway. However, the kinetics and control mechanism of the pathway have not yet been fully understood. We have developed the computer simulation of the JAK/STAT pathway. Without nuclear phosphatase, SOCS1's binding to JAK did not cause the decrease in nuclear phosphorylated STAT1. However, without SH2 domain-containing tyrosine phosphatase 2 (SHP-2) or cytoplasmic phosphatase, it did. So nuclear phosphatase is considered to be the most important in this system. By changing parameters of the model, dynamical characteristics and control mechanism were investigated. © 2002 Federation of European Microbiological Societies. Published by Elsevier Science B.V. All rights reserved.

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SHP2 and SOCS3 contribute to Tyr-759-dependent attenuation of interleukin-6 signaling through gp130

Lehmann Ute, Schmitz Jochen, Weissenbach Manuela, Sobota Radoslaw M., Hörtner Michael, Friederichs Kerstin, Behrmann Iris, Tsiaris William, Sasaki Atsuo, Schneider-Mergener Jens, Yoshimura Akihiko, Neel Benjamin G., Heinrich Peter C., Schaper Fred

Journal of Biological Chemistry 278 ( 1 ) 661 - 671 2003.01

ISSN 0021-9258

Interleukin-6 (IL-6) activates the Jak/STAT pathway as well as the mitogen-activated protein kinase cascade. Tyrosine 759 of the IL-6 signal-transducing receptor subunit gp130 has been identified as being involved in negative regulation of IL-6-induced gene induction and activation of the Jak/STAT pathway. Because this site is known to be a recruitment motif for the protein-tyrosine phosphatase SHP2, it has been suggested that SHP2 is the mediator of tyrosine 759-dependent signal attenuation. We recently observed that the suppressor of cytokine-signaling SOCS3 also acts through the tyrosine motif 759 of gp130. However, the relative contributions of SHP2 and SOCS3 to the repression of IL-6 signaling are not understood. Therefore, we designed experiments allowing the independent recruitment of each of these proteins to the IL-6-receptor complex. We show that receptor- and membrane-targeted SHP2 counteracts IL-6 signaling independent of SOCS3 binding to gp130. On the other hand, SOCS3 inhibits signaling in cells expressing a truncated SHP2 protein, which is not recruited to gp130. These data suggest, that there are two, largely distinct modes of negative regulation of gp130 activity, despite the fact that both SOCS3 and SHP2 are recruited to the same site within gp130.

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CIS1 interacts with the Y532 of the prolactin receptor and suppresses prolactin-dependent STAT5 activation

Endo Takaho, Sasaki Atsuo, Minoguchi Mayu, Joo Akiko, Yoshimura Akihiko

Journal of Biochemistry 133 ( 1 ) 109 - 113 2003.01

ISSN 0021-924X

Prolactin (PRL) interacts with a single-chain prolactin-specific receptor of the cytokine receptor superfamily. PRL triggers the activation of JAK2 kinase, which phosphorylates the PRL receptor itself, and of STAT5, a member of the family of signal transducers and activators of transcription (STAT). We have shown that the STAT5-dependent immediate early gene, CIS1 (Cytokine-Inducible SH2 domain-containing protein-1), suppresses PRL-induced STAT5 activation in vitro as well as in transgenic mice. To facilitate the study of the interactions between CIS1 and the PRL receptor, we have developed the yeast tri-hybrid system, a modification of the yeast two-hybrid system. We expressed CIS1 fused to the DNA-binding domain and PRL receptor cytoplasmic domain fused to the transcription activation domain in the presence or absence of the tyrosine kinase domain of JAK2 in yeast. CIS1 bound to the PRL receptor cytoplasmic domain in a JAK2-dependent manner. Moreover, we determined that the phosphorylated Y532 of the murine PRL receptor is the binding site for CIS1. Interestingly, Y532 has been shown to be unnecessary for STAT5 activation, although CIS1 overexpression suppressed PRL-induced STAT5 activation. These data suggest that the suppression of STAT5 activation by CIS1 is not due to a simple competition with STAT5 but rather to a modification of the receptor by CIS1 binding.

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Negative regulation of cytokine signaling by CIS/SOCS family proteins and their roles in inflammatory diseases.

Hanada T., Kinjyo I., Inagaki-Ohara K., Yoshimura A.

Reviews of physiology, biochemistry and pharmacology 149 72 - 86 2003

ISSN 0303-4240

Immune and inflammatory systems are controlled by multiple cytokines, including interleukins (ILs) and interferons. These cytokines exert their biological functions through Janus tyrosine kinases (JAKs) and STAT transcription factors. The CIS (cytokine-inducible SH2 protein) and SOCS (suppressors of cytokine signaling) are a family of intracellular proteins, several of which have emerged as key physiological regulators of cytokine responses, including those that regulate the inflammatory systems. In this review, we focused on the molecular mechanism of the action of CIS/SOCS family proteins and their roles in inflammatory diseases. Furthermore, we illustrate several approaches for treating inflammatory diseases by modulating extracellular and intracellular signaling pathways.

Combined hypermethylation and chromosome loss associated with inactivation of SSI-1/SOCS-1/JAB gene in human hepatocellular carcinomas

Nagai Hisaki, Kim Yong Sung, Konishi Noboru, Baba Masaru, Kubota Takeo, Yoshimura Akihiko, Emi Mitsuru

Cancer Letters 186 ( 1 ) 59 - 65 2002.12

ISSN 0304-3835

We previously demonstrated using restriction landmark genomic scanning-based 2-dimensional genome electrophoresis method decreased results of 16 primary hepatocellular carcinomas (HCCs) revealed reduction of intensity of 60 NotI-landmark spots, and increase in five spots that were frequently observed in HCCs. Most frequently decreased spot (14/16 HCCs) was identified to it corresponds to a gene encoding SSI-1, a JAK-binding protein (SSI-1/SOCS-1/JAB) that regulated the JAK/STAT signal transduction pathway. This signaling pathway is important for relaying signals from various cytokines outside the cell to the inside. Expression level of SOCS-1 messenger RNA was markedly suppressed in 50% of HCCs (4/8). Loss of heterozygosity at the SSI-1 gene, was found in all cases with aberrant expression. Methylation analysis of the CpG-rich regions of SSI-1 gene revealed hypermethylation of these regions. In an additional series of methylation analysis using 30 HCCs, 16 (53%) showed hypermethylation of the gene. These results indicate that the SSI-1 gene is silenced in a substantial portion of HCC though the combined mechanisms of methylation of either 5′ or exon CpG rich regions and by a chromosomal loss of the remaining allele. © 2002 Elsevier Science Ireland Ltd. All rights reserved.

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SOCS1/JAB is a negative regulator of LPS-induced macrophage activation

Kinjyo Ichiko, Hanada Toshikatsu, Inagaki-Ohara Kyoko, Mori Hiroyuki, Aki Daisuke, Ohishi Masanobu, Yoshida Hiroki, Kubo Masato, Yoshimura Akihiko

Immunity 17 ( 5 ) 583 - 591 2002.11

ISSN 1074-7613

Bacterial lipopolysaccharide (LPS) triggers innate immune responses through Toll-like receptor (TLR) 4. We show here that the suppressor of cytokine-signaling-1 (SOCS1/JAB) is rapidly induced by LPS and negatively regulates LPS signaling. SOCS1^+/- mice or SOCS1^-/- mice with interferon-γ (IFNγ)-deficient background were more sensitive to LPS-induced lethal effects than were wild-type littermates. LPS-induced NO₂^- synthesis and TNFα production were augmented in SOCS1^-/- macrophages. Furthermore, LPS tolerance, a protection mechanism against endotoxin shock, was also strikingly reduced in SOCS1^-/- cells. LPS-induced I-κB and p38 phosphorylation was upregulated in SOCS1^-/- macrophages, and forced expression of SOCS1 suppressed LPS-induced NF-κB activation. Thus, SOCS1 directly suppresses TLR4 signaling and modulates innate immunity.

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Expression of the suppressor of cytokine signaling-5 (SOCS5) negatively regulates IL-4-dependent STAT6 activation and Th2 differentiation

Seki Yoh Ichi, Hayashi Katsuhiko, Matsumoto Akira, Seki Noriyasu, Tsukada Jun, Ransom John, Naka Tetsuji, Kishimoto Tadamitsu, Yoshimura Akihiko, Kubo Masato

Proceedings of the National Academy of Sciences of the United States of America 99 ( 20 ) 13003 - 13008 2002.10

ISSN 0027-8424

The development of helper T (Th) cell subsets, which secrete distinct cytokines, plays an important role in determining the type of immune response. The IL-4-mediated Janus kinase-signal transducer and activator of transcription signaling pathway is crucial for mediating Th2 cell development. Notably, this pathway is selectively impaired in Th1 cells, although the molecular basis of this impairment remains unclear. We show here that during Th1 differentiation a reduction in the association of Janus kinase 1 with the IL-4 receptor (IL-4R) correlated with the appearance of the suppressor of cytokine signaling-5 (SOCS5). SOCS5 protein was preferentially expressed in committed Th1 cells and interacted with the cytoplasmic region of the IL-4Rα chain irrespective of receptor tyrosine phosphorylation. This unconventional interaction of SOCS5 protein with the IL-4R resulted in the inhibition of IL-4-mediated signal transducer and activator of transcription-6 activation. T cells from transgenic mice constitutively expressing SOCS5 exhibited a significant reduction of IL-4-mediated Th2 development. Therefore, the induced SOCS5 protein in Th1 differentiation environment may play an important role by regulating Th1 and Th2 balance.

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Activation of STAT3 by the hepatitis C virus core protein leads to cellular transformation

Yoshida Takafumi, Hanada Toshikatsu, Tokuhisa Takeshi, Kosai Ken Ichiro, Sata Michio, Kohara Michinori, Yoshimura Akihiko

Journal of Experimental Medicine 196 ( 5 ) 641 - 653 2002.09

ISSN 0022-1007

The signal transducer and activator of transcription (STAT) family proteins are transcription factors critical in mediating cytokine signaling. Among them, STAT3 is often constitutively phosphorylated and activated in human cancers and in transformed cell lines and is implicated in tumorigenesis. However, cause of the persistent activation of STAT3 in human tumor cells is largely unknown. The hepatitis C virus (HCV) is a major etiological agent of non-A and non-B hepatitis, and chronic infection by HCV is associated with development of liver cirrhosis and hepatocellular carcinoma. HCV core protein is proposed to be responsible for the virus-induced transformation. We now report that HCV core protein directly interacts with and activates STAT3 through phosphorylation of the critical tyrosine residue. Activation of STAT3 by the HCV core in NIH-3T3 cells resulted in rapid proliferation and up-regulation of Bcl-XL and cyclin-D1. Additional expression of STAT3 in HCV core-expressing cells resulted in anchorage-independent growth and tumorigenesis. We propose that the HCV core protein cooperates with STAT3, which leads to cellular transformation.

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Regulation of cytokine signaling and inflammation

Hanada Toshitkatsu, Yoshimura Akihiko

Cytokine and Growth Factor Reviews 13 ( 4-5 ) 413 - 421 2002.08

ISSN 1359-6101

Inflammation progresses by the action of pro-inflammatory cytokines, including interleukin-1 (IL-1), the tumor necrosis factor (TNF), gamma-interferon (IFNγ), IL-12, IL-18, and the granulocyte-macrophage colony-stimulating factor, and is resolved by anti-inflammatory cytokines such as IL-4, IL-10, IL-13, IFNα, and the transforming growth factor (TGF)β. The intracellular signal transduction pathways of these cytokines have been studied extensively, and these pathways ultimately activate transcription factors, such as NF-κB, Smad, and STATs. Recently, the negative -feedback regulation of these pathways has been identified. In this review, we provide examples of the relationship between cytokine signal transduction, negative-signal regulation, and inflammatory disease models. Furthermore, we illustrate several approaches for treating inflammatory diseases by modulating extracellular and intracellular signaling pathways. © 2002 Elsevier Science Ltd. All rights reserved.

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Involvement of suppressor of cytokine signaling-3 as a mediator of the inhibitory effects of IL-10 on lipopolysaccharide-induced macrophage activation

Berlato Chiara, Cassatella Marco A., Kinjyo Ichiko, Gatto Luana, Yoshimura Akihiko, Bazzoni Flavia

Journal of Immunology 168 ( 12 ) 6404 - 6411 2002.06

ISSN 0022-1767

Previous studies have shown that IL-10 can induce the expression of the suppressor of cytokine signaling 3 (SOCS-3) mRNA in human monocytes and neutrophils, suggesting that the capacity of IL-10 to inhibit the expression of LPS-inducible proinflammatory genes may depend on SOCS-3 induction. However, no direct experimental evidence has been provided to support such hypothesis. Herein, we show that stable transfection of SOCS-3 into the mouse macrophage cell line J774 resulted in an inhibition of NO, TNF-α, IL-6, and GM-CSF secretion in response to LPS at levels similar to those exerted by IL-10 in LPS-stimulated wild-type J774. Constitutive SOCS-3 expression also down-regulated the mRNA expression of inducible NO synthase and IL-6 and impaired the production of TNF-α, mainly at a post-transcriptional level. In addition, SOCS-3-transfected cells displayed a constitutive expression of the IL-1R antagonist gene, consistent with the observation that IL-10 enhances IL-1R antagonist mRNA in LPS-stimulated wild-type cells. Furthermore, in peritoneal macrophages harvested from mice carrying heterozygous disruption of the SOCS-3 gene, IL-10 was less effective in repressing LPS-stimulated TNF-α and NO production. Taken together, our data show that SOCS-3 inhibits LPS-induced macrophage activation, strongly supporting the idea that it plays a role in the molecular mechanism by which IL-10 down-modulates the effector functions of LPS-activated macrophages. Finally, we show that forced expression of SOCS-3 significantly suppresses the ability of IL-10 to trigger tyrosine phosphorylation of STAT3. Therefore, SOCS-3 functions both as an LPS signal inhibitor and as a negative feedback regulator of IL-10/STAT3 signaling.

Stat3β inhibits γ-globin gene expression in erythroid cells

Foley Heather A., Ofori-Acquah Solomon F., Yoshimura Akihiko, Critz Stuart, Surendra Baliga B., Pace Betty S.

Journal of Biological Chemistry 277 ( 18 ) 16211 - 16219 2002.05

ISSN 0021-9258

We demonstrated previously γ-globin gene inhibition in K562 cells and primary erythroid progenitors treated with interleukin-6. Although several cis-acting elements have been identified in the globin promoters, the precise mechanism for cytokine-mediated globin gene regulation remains to be elucidated. In this report we demonstrate inhibitors of Stat3 phosphorylation abrogate interleukin-6-mediated γ gene silencing in erythroid cells. DNA-protein binding studies established Stat3 interaction in the 5′-untranslated γ-globin promoter region. Furthermore, co-transfection experiments with Stat3β demonstrate γ promoter inhibition in a concentration-dependent manner, which was significantly reversed when the cognate Stat3-binding site in the 5′-untranslated region was mutated. These studies establish a novel mechanism for γ gene silencing through the STAT signal transduction pathway.

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Erratum

Hara Hiromitsu, Takeda Atsunobu, Takeuchi Michiyo, Wakeham Andrew C., Itié Annick, Sasaki Masafumi, Mak Tak W., Yoshimura Akihiko, Nomoto Kikuo, Yoshida Hiroki

Journal of Immunology 168 ( 8 ) 4241 2002.04

ISSN 0022-1767

The apoptotic protease-activating factor 1-mediated pathway of apoptosis is dispensable for negative selection of thymocytes

Hara Hiromitsu, Takeda Atsunobu, Takeuchi Michiyo, Wakeham Andrew C., Itié Annick, Sasaki Masafumi, Mak Tak W., Yoshimura Akihiko, Nomoto Kikuo, Yoshida Hiroki

Journal of Immunology 168 ( 5 ) 2288 - 2295 2002.03

ISSN 0022-1767

Negative selection is a process to delete potentially autoreactive clones in developing thymocytes. Programmed cell death or apoptosis is thought to play an important role in this selection process. In this study, we investigated the role of apoptotic protease-activating factor 1 (Apaf1), a mammalian homologue of CED-4, in programmed cell death during the negative selection in thymus. There was no developmental abnormality in thymocytes from newborn Apaf1^-/- mice in terms of CD4 and CD8 expression pattern and thymocyte number. Clonal deletion by endogenous male H-Y Ag of Apaf1-deficient thymocytes with transgenic expression of H-Y Ag-specific TCRs (H-Y Tg/Apaf1^-/- thymocytes) was normally observed in lethally irradiated wild-type mice reconstituted with fetal liver-derived hemopoietic stem cells. Clonal deletion induced in vitro by a bacterial superantigen was also normal in fetal thymic organ culture. Thus, Apaf1-mediated pathway of apoptosis is dispensable for the negative selection of thymocytes. However, H-Y Tg/Apaf1^-/- thymocytes showed partial resistance to H-Y peptide-induced deletion in vitro as compared with H-Y Tg/Apaf1^+/- thymocytes, implicating the Apaf1-mediated apoptotic pathway in the negative selection in a certain situation. In addition, the peptide-induced deletion was still observed in H-Y Tg/Apaf1^-/- thymocytes in the presence of a broad spectrum caspase inhibitor, z-VAD-fmk, suggesting the presence of caspase-independent cell death pathway playing roles during the negative selection. We assume that mechanisms for the negative selection are composed of several cell death pathways to avoid failure of elimination of autoreactive clones.

Induction of SOCS family proteins in inflammatory diseases and suppression of inflammatory arthritis model by the adenovirus gene transfer

Yoshimura Akihiko, Shouda Takanori, Yoshida Takafumi, Hanada Toshikatsu

Tanpakushitsu kakusan koso. Protein, nucleic acid, enzyme 47 ( 16 Suppl ) 2355 - 2361 2002

ISSN 0039-9450

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