Takayoshi Matsumura

Vascular Implications of the Krüppel-Like Family of Transcription Factors

The Krüppel-like factor (KLF) family is a recently highlighted group of zinc finger transcription factors given their important biological roles which include the vasculature. KLF2, KLF4, KLF5, and KLF6 are notable factors that have been implicated in developmental as well as pathological vascular processes. In this brief review, we provide an up-to-date summary of the physiological functions and cellular effects as well as transcriptional regulatory mechanisms of the vascular KLFs. Through such, we aim to provide a working view for understanding the pathological actions of KLFs in the vasculature.

Regulation of Platelet-derived Growth Factor-A Chain by Krüppel-like Factor 5 NEW PATHWAY OF COOPERATIVE ACTIVATION WITH NUCLEAR FACTOR-κB

The transcription factor Krüppel-like factor 5 (KLF5) and its genetically downstream target gene platelet-derived growth factor-A (PDGF-A) chain are key factors in regulation of cardiovascular remodeling in response to stress. We show that KLF5 mediates a novel distinct delayed persistent induction of PDGF-A chain in response to the model agonist, phorbol ester, through a cis-element previously shown to mediate phorbol ester induction on to PDGF-A chain through the early growth response factor (Egr-1). Interestingly, the nuclear factor-κB (NF-κB) p50 subunit further cooperatively activates PDGF-A chain through protein-protein interaction with KLF5 but not Egr-1. RNA interference analysis confirmed that KLF5 and p50 are important for induction of PDGF-A chain. Collectively, we identify a novel regulatory pathway in which PDGF-A chain gene expression, under the control of KLF5, is cooperatively activated by the NF-κB p50 subunit and a pathophysiological stimulus.

Two Susceptibility Loci to Takayasu Arteritis Reveal a Synergistic Role of the IL12B and HLA-B Regions in a Japanese Population

Takayasu arteritis (TAK) is an autoimmune systemic vasculitis of unknown etiology. Although previous studies have revealed that HLA-B*52:01 has an effect on TAK susceptibility, no other genetic determinants have been established so far. Here, we performed genome scanning of 167 TAK cases and 663 healthy controls via Illumina Infinium Human Exome BeadChip arrays, followed by a replication study consisting of 212 TAK cases and 1,322 controls. As a result, we found that the IL12B region on chromosome 5 (rs6871626, overall p = 1.7 × 10(-13), OR = 1.75, 95% CI 1.42-2.16) and the MLX region on chromosome 17 (rs665268, overall p = 5.2 × 10(-7), OR = 1.50, 95% CI 1.28-1.76) as well as the HLA-B region (rs9263739, a proxy of HLA-B*52:01, overall p = 2.8 × 10(-21), OR = 2.44, 95% CI 2.03-2.93) exhibited significant associations. A significant synergistic effect of rs6871626 and rs9263739 was found with a relative excess risk of 3.45, attributable proportion of 0.58, and synergy index of 3.24 (p ≤ 0.00028) in addition to a suggestive synergistic effect between rs665268 and rs926379 (p ≤ 0.027). We also found that rs6871626 showed a significant association with clinical manifestations of TAK, including increased risk and severity of aortic regurgitation, a representative severe complication of TAK. Detection of these susceptibility loci will provide new insights to the basic mechanisms of TAK pathogenesis. Our findings indicate that IL12B plays a fundamental role on the pathophysiology of TAK in combination with HLA-B(∗)52:01 and that common autoimmune mechanisms underlie the pathology of TAK and other autoimmune disorders such as psoriasis and inflammatory bowel diseases in which IL12B is involved as a genetic predisposing factor.

Krüppel-like factor 5 shows proliferation-specific roles in vascular remodeling, direct stimulation of cell growth and inhibition of apoptosis

Krüppel-like factor 5 (KLF5), originally isolated as a regulator of phenotypic modulation of vascular smooth muscle cells, induces pathological cell growth and is expressed in the neointima. Although induction of KLF5 up-regulates growth factors like platelet-derived growth factor-A chain, how KLF5 actually contributes to vascular remodeling, notably its direct effects on cell proliferation, had been poorly clarified. To investigate the effects of KLF5 on neointimal formation, we at first performed adenoviral overexpression of KLF5 to rats subjected to carotid balloon injury. Neointimal formation and proliferating cell nuclear antigen-positive rate were significantly increased at 14 days after injury in the KLF5-treated animals. At the cellular level, overexpression of KLF5 also resulted in markedly increased cell proliferation and cell cycle progression. As a molecular mechanism, we showed that KLF5 directly bound to the promoter and up-regulated gene expression of cyclin D1, as well as showing specific transactivation of cyclins and cyclin-dependent kinase inhibitors in cardiovascular cells. Conversely, knockdown of KLF5 by RNA interference specifically down-regulated cyclin D1 and impaired vascular smooth muscle cell proliferation. Furthermore, KLF5 attenuated cleavage of caspase-3 under conditions of apoptotic stimulation. Moreover, KLF5-administered animals exhibited a significant decrease in terminal deoxynucleotidyltransferase-mediated dUTP nick end-labeling-positive cells in the medial layer, suggesting inhibition of apoptosis in the early phase after denudation. These findings collectively suggest that KLF5 plays a central role in cardiovascular pathologies through direct and specific stimulation of cell growth as well as inhibition of apoptosis.

The Deacetylase HDAC1 Negatively Regulates the Cardiovascular Transcription Factor Krüppel-like Factor 5 through Direct Interaction

Transcription is regulated by a network of transcription factors and related cofactors that act in concert with the general transcription machinery. Elucidating their underlying interactions is important for understanding the mechanisms regulating transcription. Recently, we have shown that Krüppel-like factor KLF5, a member of the Sp/KLF family of zinc finger factors and a key regulator of cardiovascular remodeling, is regulated positively by the acetylase p300 and negatively by the oncogenic regulator SET through coupled interaction and regulation of acetylation. Here, we have shown that the deacetylase HDAC1 can negatively regulate KLF5 through direct interaction. KLF5 interacts with HDAC1 in the cell and in vitro. Gel shift DNA binding assay showed that their interaction inhibits the DNA binding activity of KLF5, suggesting a property of HDAC1 to directly affect the DNA binding affinity of a transcription factor. Reporter assay also revealed that HDAC1 suppresses KLF5-dependent promoter activation. Additionally, overexpression of HDAC1 suppressed KLF5-dependent activation of its endogenous downstream gene, platelet-derived growth factor-A chain gene, when activated by phorbol ester. Further, HDAC1 binds to the first zinc finger of KLF5, which is the same region where p300 interacts with KLF5 and, intriguingly, HDAC1 inhibits binding of p300 to KLF5. Direct competitive interaction between acetylase and deacetylase has been hitherto unknown. Collectively, the transcription factor KLF5 is negatively regulated by the deacetylase HDAC1 through direct effects on its activities (DNA binding activity, promoter activation) and further through inhibition of interaction with p300. These findings suggest a novel role and mechanism for regulation of transcription by deacetylase.

Giant tumorous lesions (correction of legions) surrounding the right coronary artery associated with immunoglobulin-G4-related systemic disease.

Immunoglobulin G4 (IgG4)-related systemic disease was first recognized as a clinicopathological entity about 10 years ago, and since then, it has attracted growing attention. It is an autoimmune disease which affects multiple organs including the pancreas, bile duct, salivary glands and retroperitoneum. Further, it was recently reported that it can be manifested as periarteritis, often as inflammatory abdominal aortic aneurysm. We describe the case of a 75-year-old man with autoimmune pancreatitis and parotitis who presented with angina. The serum concentration of IgG4 was significantly increased at 2,510 mg/dl. Coronary angiography showed multiple stenotic lesions and pronounced dilatation of the right coronary artery. Cardiac computed tomography disclosed increased wall thickness of the coronary arteries and focal tumorous lesions surrounding the right coronary artery. Treatment with steroids proved only marginally effective and he underwent surgical resection of the aneurysm and coronary artery bypass grafting. The diagnosis of IgG4-related systemic disease was confirmed by histological examination of the resected mass, which showed a massive infiltration of IgG4-positive plasma cells. This case emphasizes the importance of considering the diagnosis in any patient with abnormally increased wall thickness or ectatic lesions in the coronary arteries.

Promoter Region-Specific Histone Incorporation by the Novel Histone Chaperone ANP32B and DNA-Binding Factor KLF5

ABSTRACT Regulation of chromatin in eukaryotic transcription requires histone-modifying enzymes, nucleosome remodeling complexes, and histone chaperones. Specific regulation of histone incorporation/eviction by histone chaperones on the promoter (e.g., region specific) is still poorly understood. In the present study, we show that direct and functional interaction of histone chaperone and DNA-binding transcription factor leads to promoter region-specific histone incorporation and inhibition of histone acetylation. We report here that the DNA-binding transcription factor Krüppel-like factor 5 (KLF5) interacts with the novel histone chaperone acidic nuclear phosphoprotein 32B (ANP32B), leading to transcriptional repression of a KLF5-downstream gene. We further show that recruitment of ANP32B onto the promoter region requires KLF5 and results in promoter region-specific histone incorporation and inhibition of histone acetylation by ANP32B. Extracellular stimulus (e.g., phorbol ester) regulates this mechanism in the cell. Collectively, we have identified a novel histone chaperone, ANP32B, and through analysis of the actions of this factor show a new mechanism of promoter region-specific transcriptional regulation at the chromatin level as mediated by the functional interaction between histone chaperone and DNA-binding transcription factor.

Brief report: Takayasu arteritis and ulcerative colitis: High rate of co-occurrence and genetic overlap

Takayasu arteritis (TAK) is a systemic vasculitis affecting large arteries and large branches of the aorta. Ulcerative colitis (UC) is a prevalent autoimmune colitis. Since TAK and UC share HLA–B*52:01 and IL12B as genetic determinants, and since there are case reports of the co‐occurrence of these diseases, we hypothesized that UC is a common complication of TAK. We undertook this study to perform a large‐scale analysis of TAK, both to evaluate the prevalence of concurrent cases of TAK and UC and to identify and estimate susceptibility genes shared between the 2 diseases.

Functional interaction between the transcription factor Krüppel-like factor 5 and poly(ADP-ribose) polymerase-1 in cardiovascular apoptosis.

Krüppel-like factor 5 (KLF5) is a transcription factor important in regulation of the cardiovascular response to external stress. KLF5 regulates pathological cell growth, and its acetylation is important for this effect. Its mechanisms of action, however, are still unclear. Analysis in KLF5-deficient mice showed that KLF5 confers apoptotic resistance in vascular lesions. Mechanistic analysis further showed that it specifically interacts with poly(ADP-ribose) polymerase-1 (PARP-1), a nuclear enzyme important in DNA repair and apoptosis. KLF5 interacted with a proteolytic fragment of PARP-1, and acetylation of KLF5 under apoptotic conditions increased their affinity. Moreover, KLF5 wild-type (but not a non-acetylatable point mutant) inhibited apoptosis as induced by the PARP-1 fragment. Collectively, we have found that KLF5 regulates apoptosis and targets PARP-1, and further, for acetylation to regulate these effects. Our findings thus implicate functional interaction between the transcription factor KLF5 and PARP-1 in cardiovascular apoptosis.

Regulation of Transforming Growth Factor-β-dependent Cyclooxygenase-2 Expression in Fibroblasts

Abnormal transforming growth factor-β (TGF-β) signaling is a critical contributor to the pathogenesis of various human diseases ranging from tissue fibrosis to tumor formation. Excessive TGF-β signaling stimulates fibrotic responses. Recent research has focused in the main on the antiproliferative effects of TGF-β in fibroblasts, and it is presently understood that TGF-β-stimulated cyclooxygenase-2 (COX-2) induction in fibroblasts is essential for antifibroproliferative effects of TGF-β. Both TGF-β and COX-2 have been implicated in tumor growth, invasion, and metastasis, and therefore tumor-associated fibroblasts are a recent topic of interest. Here we report the identification of positive and negative regulatory factors of COX-2 expression induced by TGF-β as determined using proteomic approaches. We show that TGF-β coordinately up-regulates three factors, heterogeneous nuclear ribonucleoprotein A/B (HNRPAB), nucleotide diphosphate kinase A (NDPK A), and nucleotide diphosphate kinase A (NDPK B). Functional pathway analysis showed that HNRPAB augments mRNA and protein levels of COX-2 and subsequent prostaglandin E2 (PGE2) production by suppressing degradation of COX-2 mRNA. In contrast, NDPK A and NDPK B attenuated mRNA and protein levels of COX-2 by affecting TGF-β-Smad2/3/4 signaling at the receptor level. Collectively, we report on a new regulatory pathway of TGF-β in controlling expression of COX-2 in fibroblasts, which advances our understanding of pathophysiological mechanisms of TGF-β.